http://www.ims.riken.jp/english/
RIKEN Yokohama Campus
1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama City,
Kanagawa, 230-0045, Japan
Tel : 045-503-9111
Fax : 045-503-9113
Email : yokohama@riken.jp

RIKEN IMS
Annual Report 2016

RIKEN IMS Annual Report 2016 RIKEN Center for Integrative Medical Sciences

RIKEN Center for
Integrative Medical Sciences

RIKEN IMS
Annual Report 2016
RIKEN Center for Integrative Medical Sciences

RIKEN 2017-030

RIKEN Center for Integrative Medical Sciences
Organization Chart

Access to RIKEN Yokohama Campus

Local Access

RIKEN President
By BusTake the #08 bus from Platform 8 at the East Exit of
Tsurumi Station (also accessible from the West Exitof Keikyu Tsurumi Station) and get off at the RIKEN
Shidai Daigakuin Mae bus stop. The institute is acrossthe street. All buses from this platform are bound for
Fureyu.

Hiroshi Matsumoto

Director

Buses depart Tsurumi every 5-15 minutes. It takes
about 15 minutes to arrive at RIKEN Yokohama. The
fare is 220 yen.

Tadashi Yamamoto

Integrative
Medical Sciences
Planning Office

By Train
A 15-minute walk from JR Tsurumi-Ono Station (JRTsurumi Line), which is directly accessible by transferfrom JR Tsurumi Station.

Senior Advisor

Trains run about every 10 minutes during morning andevening rush hour, but less frequently at other times.

Masaru Taniguchi
Shizuo Akira

Searchable train timetables in English are available at
http://www.hyperdia.com/en/

By Taxi
Use the taxi stand at the East Exit of JR Tsurumi Stationor the West Exit of Keikyu Tsurumi Station. The trip
takes about 10 minutes and costs around 1,200 yen.

RIKEN Center for
Integrative
Medical Sciences
Advisory Council
Deputy Director
Michiaki Kubo
Haruhiko Koseki

Max Cooper (chair)Mark Lathrop (vice chair)
Hiroyuki Aburatani
Rudi Balling
Riccardo Dalla-Favera
Michel Georges
Ronald N. Germain
Hajime Karasuyama
Yutaka Kawakami
Paul W. Kincade
Bernard MalissenJohn O’Shea
Fiona Powrie
Peter Sorger
Kiyoshi Takatsu
Katsushi Tokunaga
Dale Umetsu
Arthur Weiss

From the Airport
From Haneda Airport

From Narita Airport

Route 1
Take the Keikyu Railways Airport Express* (blue kanjisign) for Yokohama and get off at Keikyu Tsurumi
Station (27-29 minutes). Airport Express trains run
every 10-15 minutes between 9:30 a.m. and 9:30 p.m.
Next, follow the Local Access directions above to get to
RIKEN Yokohama.

From Narita Airport Station take the JR Sobu Line
(Rapid Express), Airport Limousine Bus or JR Narita
Express to JR Shinagawa Station. (JR Sobu Line is themost inexpensive option and takes about 1 hour and 15
minutes). From JR Shinagawa Station take the JR Keihin
Tohoku Line (Yokohama direction) to JR Tsurumi Station(18 minutes). Next, follow the Local Access directions
above to get to RIKEN Yokohama.

Route 2
Take any train marked with a green (express), red or
dark grey kanji sign to Keikyu Kamata Station. Transfer
to the Keikyu Main Line and take a local train* toward
Yokohama until Keikyu Tsurumi Station* (12 minutes).

* A reserved seat express that requires payment of asurcharge in addition to train fare.
Searchable train timetables in English are available at
http://www.hyperdia.com/en/*Only Airport Express (blue kanji sign) and local trains(dark grey kanji sign) stop at Keikyu Tsurumi Station.
Note that Keikyu Tsurumi Station and JR Tsurumi Station
are two different railway stations and are separated bya bus rotary (the stations are about 150 meters apart).

85

Core for Homeostatic Regulation
Lab. for Cell Signaling: Takashi Saito

Lab. for Immune Homeostasis: Shohei HoriLab. for Lymphocyte Differentiation: Tomohiro Kurosaki

Lab. for Skin Homeostasis: Masayuki Amagai

Lab. for Transcriptional Regulation: Ichiro Taniuchi

Lab. for Metabolic Homeostasis: Naoto Kubota

Lab. for Immune Cell Systems: Shigeo Koyasu

Lab. for Immune Crosstalk: Hilde Cheroutre

Lab. for Human Disease Models: Fumihiko Ishikawa

Lab. for Inflammatory Regulation: Takashi Tanaka

Lab. for Intestinal Ecosystem: Hiroshi Ohno

Lab. for Cytokine Regulation: Masato Kubo

Lab. for Mucosal Immunity: Sidonia FagarasanLab. for Innate Immune Systems: Kazuyo MoroLab. for Gut Homeostasis: Kenya Honda

Core for Precise Measuring and ModelingLab. for Developmental Genetics: Haruhiko KosekiLab. for Integrated Bioinformatics: Todd Duane Taylor

Lab. for Integrative Genomics: Osamu Ohara

Lab. for Tissue Dynamics: Takaharu Okada

Lab. for Disease Systems Modeling: Hiroaki KitanoLab. for Integrated Cellular Systems: Mariko Okada

Lab. for Medical Science Mathematics: Tatsuhiko Tsunoda

Lab. for Metabolomics: Makoto Arita

Lab. for Immunogenetics: Tadashi Yamamoto

Lab. for Microbiome Sciences: Masahira HattoriCore for Genomic Medicine
Lab. for Genotyping Development: Yukihide Momozawa

Lab. for Autoimmune Diseases: Kazuhiko Yamamoto

Lab. for Genome Sequencing Analysis: Hidewaki Nakagawa

Lab. for Digestive Diseases: Kazuaki Chayama

Lab. for Statistical Analysis: Yoichiro Kamatani

Lab. for Bone and Joint Diseases: Shiro Ikegawa

Lab. for Pharmacogenomics: Taisei Mushiroda

Lab. for Endocrinology, Metabolism and Kidney Diseases:

Lab. for International Alliance on Genomic Research:

Taisei Mushiroda

Momoko Horikoshi
Lab. for Respiratory and Allergic Diseases: Mayumi Tamari

Lab. for Cardiovascular Diseases: Kaoru Ito

Program for Medical Innovations
Lab. for Immune Regulation: Masaru Taniguchi

Drug Discovery Antibody Platform Unit: Toshitada TakemoriLab. for Immunotherapy: Shin-ichiro Fujii

Young Chief Investigator Program
YCI Laboratory for Immune Regeneration: Tomokatsu Ikawa

YCI Laboratory for Trans-omics: Katsuyuki Yugi

YCI Laboratory for Cellular Bioenergetic Network:

YCI Laboratory for Immunological Transcriptomics:

Toshimori Kitami

Hideyuki YoshidaiContents

Organization .......................................................................... iContents ................................................................................. ii

Director’s Report ................................................................... iv

Part 1
Research Highlights
Liver cancer’s genomic landscape ....................................... 2
Mutational signatures mark cancer's smoking gun .......... 3
New anti-cancer strategy mobilizes both innate
and adaptive immune responses ................................... 4
Hyperactivation of JAK1 tyrosine kinase induces
atopic dermatitis ............................................................. 5
Immune system overreaction may trigger eczema
into becoming a chronic disease ................................... 6

New mechanism of antiviral protection ............................. 7
Mechanism of memory B cell differentiation .................... 8
GWAS identifies seven novel loci for type 2 diabetes ....... 9
Sending bone growth down the spine ................................ 10
GWAS identifies a genetic risk factor associatedwith clozapine-induced agranulocytosis/
granulocytopenia ............................................................ 11
Keeping viral DNA at bay .................................................... 12Part 2Lab Activities
Core for Homeostatic Regulation ........................... 14
Laboratory for Cell Signaling ........................................ 15
Laboratory for Lymphocyte Differentiation ................ 16
Laboratory for Transcriptional Regulation .................. 17
Laboratory for Immune Cell Systems .......................... 18
Laboratory for Human Disease Models ....................... 19
Laboratory for Intestinal Ecosystem ............................ 20

Laboratory for Immunogenetics ................................... 35
Laboratory for Integrated Bioinformatics .................... 36
Laboratory for Tissue Dynamics .................................. 37
Laboratory for Integrated Cellular Systems ................. 38
Laboratory for Metabolomics ....................................... 39
Core for Genomic Medicine ....................................... 40
Laboratory for Genotyping Development ................... 41

Laboratory for Mucosal Immunity ............................... 21
Laboratory for Gut Homeostasis .................................. 22
Laboratory for Immune Homeostasis .......................... 23
Laboratory for Skin Homeostasis ................................. 24
Laboratory for Metabolic Homeostasis ........................ 25
Laboratory for Immune Crosstalk ................................ 26

Laboratory for Genome Sequencing Analysis ............. 42
Laboratory for Statistical Analysis ................................ 43
Laboratory for Pharmacogenomics .............................. 44
Laboratory for Cardiovascular Diseases ...................... 45
Laboratory for Autoimmune Diseases ......................... 46
Laboratory for Digestive Diseases ................................ 47

Laboratory for Inflammatory Regulation .................... 27
Laboratory for Cytokine Regulation ............................ 28
Laboratory for Innate Immune Systems ...................... 29
Core for Precise Measuring and Modeling ......... 30
Laboratory for Developmental Genetics ...................... 31

Laboratory for Bone and Joint Diseases ....................... 48
Laboratory for Diabetes, Metabolism andRenal diseases ........................................................... 49
Laboratory for Respiratory and Allergic Diseases ...... 50
Program for Medical Innovations .......................... 51

Laboratory for Integrative Genomics ........................... 32
Laboratory for Disease Systems Modeling .................. 33
Laboratory for Medical Science Mathematics ............. 34

Laboratory for Immune Regulation ............................. 52
Laboratory for Immunotherapy .................................... 53
Drug Discovery Antibody Platform Unit .................... 54ii

YCI Laboratory for Immune Regeneration ................. 55
YCI Laboratory for Cellular Bioenergetic Network ... 56
Central Facilities
FACS Laboratory ............................................................ 57
Confocal Laboratory ...................................................... 57
Genomics Laboratory ..................................................... 58
Animal Facility ................................................................ 58

Other Programs
RIKEN International Program Associate (IPA) .......... 60
RIKEN Foreign Postdoctoral Researcher (FPR)
Program ..................................................................... 60
RIKEN Junior Research Associate (JRA) Program .... 60
RIKEN Special Postdoctoral Researcher (SPDR)
Program ..................................................................... 60

Award Winners 2016 .................................................... 59

Part 3
Research Projects
Projects
iPS project ........................................................................ 62
Modeling skin diseases .................................................. 62
Integrated omics approach to elucidate
the impact of the gut microbiomeon the pathogenesis of type 2 diabetes ................... 63
Humanized mouse .......................................................... 63
NKT Cell Projects ........................................................... 64

Linkage to RIKEN Program for Drug Discovery
and Medical Technology Platforms (DMP) .......... 64
PGRN-RIKEN Project ................................................... 65
Collaboration with Asian institutes and SEAPharm ... 65
International Cancer Genome Consortium (ICGC) ... 66
Commissioned Research
The Biobank Japan project ............................................. 66

Part 4
EventsRIKEN IMS Summer Program (RISP) 2016 ..................... 68

RIKEN IMS-IITU Joint Workshop ..................................... 70

The IMS-JSI International Symposium on
Immunology 2016 .......................................................... 68

The Academia Sinica IBMS - RIKEN IMS
Joint Workshop 2016 ...................................................... 71

13th PGRN-RIKEN Strategic Alliance Meeting ............... 69
IMS Advisory Council 2016 ................................................ 69
Harvard Summer School 2016 ............................................ 70

Adjunct Professorship Programs ........................................ 71
Guest lectures 2016 ....................................................... 72

Part 5
Data and Statistics
Publlications 2016 ................................................................. 74

Access to RIKEN Yokohama Campus ..................... 85

Budget, personnel and patents ............................................ 84iii

Director’s ReportT

his is the fourth annual report of the RIKEN Center
for Integrative Medical Sciences (IMS), and one andhalf years have been passed since I became IMS Director.
IMS held its second Advisory Council Meeting (IMAC)
in September, 2016, and RIKEN’s Advisory Council (RAC)
met in December. It was my first experience to attend thesemeetings, but I was very much encouraged because in both
meetings, IMS was characterized as one of the key pillars of
RIKEN’s medical and life science research.
IMS has been making tremendous contributions to
RIKEN’s multidisciplinary projects: for example, Aging
Project, Epigenetics Project, Medical Innovation Hub Program, and Single Cell Project. In addition, IMS has started
a discussion with the Division of Genomic Technologies,
led by Dr. Piero Carninci, about our future collaborations
and common visions.
Our body constantly attempts to maintain its internalstability, but environmental and genetic factors can disrupt
this homeostasis. When the body responds to environmental changes, the result is always changes in gene expression.
Thus, we hope to understand how genetic and environmental factors regulate our body homeostasis and diseases, andthereby we will contribute to human health. To succeed inthis aim, it is necessary to collect data in multiple layers
and then to integrate them to understand and simulate thewhole system. I believe that IMS’s expertise in immunology
and genomics, together with DGT’s expertise in translational regulation, will facilitate elucidation of these mechanisms and ultimately result in translation of this knowledge
into practical applications to promote our health.
The IMAC, chaired by Dr. Max Cooper of Emory Uni-

iv

versity USA, encouraged IMS researchers to integrate
genomic and biological science at IMS, although their academic cultures are quite different. The former thrives onteam science to answer large scale questions, but on theother hand, the latter thrives on individual/small group science to investigate basic issues that may have clinical applications. The IMAC felt that the two groups could synergize
in a very powerful fashion. The comments from the IMAC
stimulated IMS researchers very much, regardless of theirfield of specialization.
In 2016 after the AC, IMS newly started an internal discussion group and a seminar series, PI Club and Researcher
Seminar, to facilitate the synergy between IMS laboratories.
At the PI Club, timely research topics at IMS are introduced
and PIs discuss the directions of the research or possibilities
for internal collaboration. At the Researcher Seminar, twoselected IMS researchers give presentations about their projects. Also, personally, I started to have lunch with several
young researchers, which I enjoy very much. I feel that gradually people are starting to recognize the Center’s direction
and are becoming enthusiastic about contributing to it.
IMS continued to publish papers in significant journals
in 2016. Hidewaki Nakagawa reported whole genome sequencing of 300 liver cancers in Japan and discovered 25
genes repeatedly mutated in different patients (Nature Genetics 2016). Masato Kubo discovered a novel mechanism
in which anti-viral antibody is generated in the absence of
TFH cells (Nature Immunology 2016). Hisahiro Yoshidapublished a paper (Journal of Clinical Investigation 201)
culminating his 15 years of work. He used a pioneering
integrative approach to collect data from multiple layers,
resulting from a collaboration between the former RIKEN
GSC and RCAI, and university hospitals. There were 289
papers published from IMS in 2016.
I believe that our unique approach will pioneer a newarea of life sciences, and hope that our challenges will bestrongly supported by RIKEN and the government.

Tadashi Yamamoto
Director,
RIKEN Center for Integrative Medical Sciences

Part 1
Research
Highlights

Liver cancer’s genomic landscapeResearch highlights

A massive DNA sequencing effort has exposed themutations behind liver cancer in Japanese patients

Figure: Liver tumors (like the one shown in redhere) are major killers. A new study has shownthat liver cancer can be classified into six different types.
©David Marchal/iStock/Getty Images Plus/Getty

I

n one of the largest ever genomic studies of a single-organcancer, scientists in Japan have discovered a multitude ofmutations responsible for causing liver cancer. The research,
led by a team from the RIKEN Center for Integrative Medical Sciences, could help doctors develop personalized drug
regimens tailored to each patient’s genetic signature.
“Whole-genome sequencing revealed that about 40% ofliver cancers had a mutation in genes related to therapeutictargets and that are hence expected to be actionable,” says
Hidewaki Nakagawa, senior author of the study.
The fifth most common cancer and the fourth leading
cause of cancer-related deaths in Japan, liver cancer was responsible for an estimated 19,000 deaths in the country lastyear alone. Chronic infection with hepatitis B and C viruses
can often lead to liver cancer, but so can alcohol abuse, metabolic diseases and certain environmental toxins, suggestingthat many different genetic drivers may interact with lifestyle factors to bring about liver tumors.
In search of mutations that could explain the diversityof the disease, Nakagawa and his colleagues sequenced theentire genomes of liver cancers from 300 Japanese patients.
That much sequencing yielded more than 300 terabytes ofdata, and the analysis required using the supercomputer
SHIROKANE—the fastest in the life-science research sector

This article was reproduced from RIKEN Research
http://www.riken.jp/en/research/rikenresearch/highlights/8220/

2

in Japan.
The researchers discovered 25 genes that were repeatedly mutated in different patients’ cancer samples, causingchanges in tumor suppressors or other regulatory proteins.
They also found a number of recurrently mutated sites in the
genome that did not code for a protein, as well as structural
rearrangements that popped up time and again with an effect on expression of nearby genes.
Looking across the mutational landscape, Nakagawa andhis team found that liver cancer among Japanese patients
could be broken down into six types—and that these types
were strongly linked to survival outcomes.
“Our study revealed that the prognosis of patients with
mutations in some genes is worse than in others,” says Akihiro Fujimoto, who worked on the research at RIKEN before
moving to the Kyoto University Graduate School of Medicine. “Although validation with an independent cohort isrequired, this result can contribute to predict patient prognosis in the future.”
Knowing the different liver cancer types and the mutations that underlie them could also help scientists develop
new, precision drug treatments. “Our analysis identified
novel driver gene candidates, which can be targets for therapies,” says Fujimoto.

Original Paper:Fujimoto, A., Furuta, M., Totoki, Y., Tsunoda, T., Kato,
M., Shiraishi, Y., Tanaka, H., Taniguchi, H., Kawakami,Y., Ueno, M. et al. Whole-genome mutational landscapeand characterization of noncoding and structural mutations in liver cancer. Nat Genet 48, 500–509 (2016)

Mutational signatures markcancer’s smoking gun

Research highlights

A study of cancer genome sequences identifies
telltale mutational signatures associated withsmoking tobacco.

Figure: Signature 4, extracted from cancer associated with tobacco smoking, is very similar
to the mutational signature induced in vitro byexposing cells to benzo[α]pyrene. Signature 4
was detected at significantly higher level in lungcancers, larynx cancers and (Japanese) liver cancers of smokers.

A

broad computational study of cancer genome sequences
identifies telltale mutational signatures associated withsmoking tobacco and demonstrates, for the first time, that smoking
increases cancer risk by causing somatic mutations in tissues directly and indirectly exposed to tobacco smoke. The international
study conducted by researchers representing 16 institutions in the
US, Europe, and Asia, including Los Alamos National Laboratory,
Wellcome Trust Sanger Institute and RIKEN Center for Integrative
Medicine, was published in the November 4 issue of Science.
Previous large-scale epidemiological studies have associated tobacco smoking with increased risk for 17 different types ofcancer, including cancer in tissues not directly exposed to smoke.
However, the mechanisms by which tobacco smoke causes cancer
have previously remained elusive. This study demonstrates thatsmoking increases cancer risk by causing somatic mutations thatboth directly damage DNA and increase the speed of an endogenous molecular clock.
“This research brings together Big Data generated by international cancer consortia and supercomputing and machine-learning capabilities to address one of the leading public health issuesof our time,” said Dr. Charlie McMillan of Los Alamos. “The work
leverages pattern-recognition software in genomic screening andrepresents a creative breakthrough in cancer research.”
The study focused on identifying the mutation signatures andDNA methylation changes in 5,243 genome sequences of smoking-related cancers by comparing the cancers of smokers to thoseof non-smokers. All mutations–harmless or cancer-promoting–This article was redacted from Los Alamos National
Laboratory Press Release
http://www.lanl.gov/discover/news-release-archive/2016/November/11.03-mutational-signtres-mark-cancers-smoking-gun.php

are due to the activity of endogenous or exogenous mutation processes, each one of which leaves a signature of scrambled DNA
code on the base pairs of that cell᾽s genome. The new study foundmore than 20 mutational signatures across the 17 cancer types associated with tobacco smoking. However, only five of these signatures were elevated in cancers from smokers. Some cancer types
had only a single mutational signature elevated in smokers, whileothers had multiple.
One signature, called signature 4, can be traced to DNA being
damaged by direct exposure to tobacco smoke and it was detected
at significantly higher level in the smokers᾽ lung cancers, larynx
cancers and (Japanese) liver cancers, interestingly. Signature 4 islikely the direct mutational consequence of misreplication of DNA
damage induced by tobacco carcinogens, particularly benzo[α]pryrene, according to the study. Signature 5, found by previous
research to occur in all cells and to trigger mutations with clocklike regularity, also correlated with increased mutations in smokers versus non-smokers. Dr. Ludmil Alexandrov of Los Alamos
explains that smoking accelerates the clock function, mostly likely by altering the molecular machinery underlying this signature.
The study modeled the cancer mutational processes as ablind-source-separation problem to distinguish coherent signals
from a noisy background, a methodology used in other areas of
Los Alamos research related to its nuclear-security program, taking advantage of high-performance computing resources and expertise, as well as expertise in numerical optimization problems.

Original paper:
Alexandrov LB, Ju YS, Haase K, Van Loo P, Martincorena
I, Nik-Zainal S, Totoki Y, Fujimoto A, Nakagawa H,
Shibata T, Campbell PJ, Vineis P, Phillips DH, Stratton
MR. Mutational signatures associated with tobaccosmoking in human cancer. Science 354, 618–622(2016)

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New anti-cancer strategy
mobilizes both innate andadaptive immune responses

Research highlights

Figure: The antitumor effect and the mechanism of aAVC
A. C57BL/6 mice were injected with OVA-expressing B16 melanoma cells s.c. and thentreated with or without aAVC-OVA on day 12. Images are representative of pre- (d12)and post-aAVC-OVA treatment. B. The infiltration of CD8+ T cells into the tumor was
analyzed by immunohistochemistry. Scale bar, 100 µm. Closely-aggregated structures
composed of CD11c+ DCs (green) and antigen-specific CD8+ T cells (red) around theblood vessels (blue) were seen in aAVC-OVA treated tumor sites. C. The frequency of
OVA antigen-specific CD8+ T cells in the spleen and tumor (TIL) in the untreated oraAVC-OVA treated, tumor-bearing mice was analyzed using OVA-tetramer binding.
The frequency of OVA-specific CD8+ T cells in both tumor (TIL) and spleen was higher
in aAVC-OVA treated mice, and in the treated mice was much higher in the tumor, thesite of tumor destruction, than in the spleen (30% vs 1% respectively).

Scientists from the RIKEN Center for Integrative Medical
Sciences have developed a new vaccine that involves injecting
cells that have been modified so that they can stimulate both aninnate immune response and the more specific adaptive responseto the tumor. This approach allows the body to develop “memory”
of the original cancer and then attack any new tumor cells as they
form or spread. In the study published in Cancer Research, the
RIKEN scientists found that the vaccine made it possible for killer
CD8+ T-cells—important players in the immune response against
cancer—to enter the tumor and target cancerous cells.
According to Shin-ichiro Fujii, leader of the Laboratory for Immunotherapy, who led the study, “Cancer cells each have different
sensitivities to the innate and adaptive response, so it is important to target both responses in order to eradicate them. We havedeveloped a special type of modified cell-based vaccine, called
aAVC, which we found can do this.”
The aAVC are not taken from the subject’s own body but areforeign cells, which are modified by adding a natural killer T cell
ligand, which permits them to stimulate innate-like natural killer
T cells (NKT), along with an antigen associated with the particular cancer. The group found that when NKT cells are activated,
they in turn promote the maturation of dendritic cells, which act
as coordinators of the innate and acquired responses. Dendritic
cells are key because they allow the activation of immune memory, where the body remembers and responds to a threat evenyears later.
To find out whether the aAVC actually worked in the body, theyThis article was reproduced fromhttp://www.riken.jp/en/pr/press/2016/20160701_2/RIKEN Press Release4

conducted experiments in mice inoculated with a virulent form of
melanoma that also expresses a model antigen called OVA. Tests
in mice showed that aggressive tumors could be shrunken by vaccinating the animals with aAVC cells that were programmed tocarry OVA antigen. Following this treatment, the tumors in thetreated animals were smaller and necrotic in the interior—a signthat the tumor was being attacked by killer CD8+T-cells.
Fujii continues, “We were interested in discovering a mechanism, and found that the aAVC treatment led to the developmentof blood vessels in the tumors that expressed a pair of importantadhesion molecules, ICAM-1 and VCAM-1, which are not normally expressed in tumors. This allowed the killer CD8+T cells topenetrate into the tumor.”
Importantly, they also found that in animals that had undergone the treatment, cancer cells injected even a year later were
eliminated. “This indicates,” says Fujii, “that we have successfully
created an immune memory that remembers the tumor and attacks it even much later.”
Looking to the future, Fujii says, “Our therapy with aAVC is
promising because typical current immunotherapies have to be
tailor-made with the patient’s own cells. In our case we use foreign cells, which can be produced in a uniform quality and usedin any patient. Because we found that our treatment can lead tothe maturation of dendritic cells, immunotherapy can move fromlocal treatment to more systemic treatment based on immune
memory.”Original article:
Shimizu K, Yamasaki S, Shinga J, Sato Y, Watanabe T,
Ohara O, Kuzushima K, Yagita H, Komuro Y, Asakura M,
Fujii S. Systemic DC activation modulates the tumor microenvironment and shapes the long-lived tumor-specific memory mediated by CD8+ T cells. Cancer Res 76,
3756–3766 (2016)

Hyperactivation of JAK1 tyrosine
kinase induces atopic dermatitis

Research highlights

Figure: Spade mutant (right) and a wildtype littermate (left) at 20 weeks of age.

A

topic dermatitis (AD) is an inflammatory skin disease
that primarily affects children. AD can be triggered byenvironmental factors such as stress, detergents, weather,and food allergies, but genetic factors also contribute to AD
pathogenesis. Hisahiro Yoshida and his team at RIKEN IMS
discovered that JAK1 hyperactivation affects the skin barriersystem and induces atopic dermatitis.
Yoshida’s team worked on phenotype screening of N-ethyl
N-nitrosourea (ENU)-induced mutant mice to find genetically-based allergic diseases. Among 3,000 mice examined,
they found an interesting line where the mice showed continuous scratching and eventually developed dermatitis. The
disease onset was from 8 to 12 weeks after birth and, interestingly, serum IgE and IgG1 elevation and serum histamine elevation were not detected until about three weeks after thedermatitis onset. Because of those characteristics, they named
this mutant “Stepwise progressive atopic dermatitis (Spade).”By genetic mapping of the Spade mutant, they found apoint mutation in the coding region of the Jak1 gene, which
encodes a tyrosine kinase used in signaling through manycytokine receptors. This amino acid change caused hyperactivation of JAK1, therefore, Yoshida decided to test whether
treating the mice with a JAK inhibitor suppressed dermatitis. As they predicted, painting JAK inhibitors on the skindelayed the onset of dermatitis.
Because Jak1 is a well-known signal transduction molecule for various cytokine receptors and strongly affects im-

mune cells, they decided to assess the role of hematopoietic
cells in the dermatitis. When Spade mouse bone marrow wastransferred into a normal mouse, the mouse did not develop dermatitis, suggesting that non-hematopoietic/immune
cells contribute to the dermatitis. In fact, dermatitis onset
was caused by a skin barrier dysfunction. When they applied
petrolatum to the Spade mouse skin, dermatitis was prevented for more than a month.
“The result was surprising to us. Some immunologists said
that this mutant cannot be an atopic dermatitis disease model because it develops independently of acquired immune
function. However, many dermatologists encouraged us, saying that such a phenotype sounded similar to juvenile typeatopic dermatitis development in human,” Yoshida commented. “In particular, the human patients usually have a defect in skin barrier function, and the disease starts with itchy
dermatitis independent of serum IgE or histamine elevation.”
To test whether JAK1 hyperactivation is also seen in human atopic dermatitis, they examined six independent atopic dermatitis skin biopsies and detected increased JAK1 activation in four of them.
“The results indicate that signal transduction by JAK1
plays an important role in the skin barrier and this can be
modified by environmental conditions. We hope that theSpade mouse will be a good model to investigate humanatopic dermatitis and to better understand the disease onset,” said Yoshida.

Original paper:
Yasuda T, Fukada T, Nishida K, Nakayama M, Matsuda
M, Miura I, Dainichi T, Fukuda S, Kabashima K, Nakaoka
S, Bin BH, Kubo M, Ohno H, Hasegawa T, Ohara O, Koseki H, Wakana S, Yoshida H. Hyperactivation of JAK1
tyrosine kinase induces stepwise, progressive pruritic
dermatitis. J Clin Invest. 2016;126 (6):2064–76.

5

Immune system overreaction
may trigger eczema into
becoming a chronic disease

Research highlights

Figure: Double-switch mechanism for the onset
and progression of AD

A

topic dermatitis (AD) is a medical condition primarily affecting children, where patches of skin become rough andinflamed with blisters, causing itching and bleeding. It is a chronic condition triggered by a combination of factors such as stress,
detergents, weather, allergies to certain foods, and genetic factors
also play a role.
Now, bioengineers from Imperial College London, RIKEN Center for Integrative Medical Sciences and Trinity College Dublin
have collated data from previous studies on AD and developed
a mathematical model that suggests how AD may progress to become chronic. The team’s model showed that repeated flare-upsof AD trigger an immune system overreaction, which, once triggered, cannot be reversed. This creates a cycle where the thresholdfor triggering further AD outbreaks becomes lower, the flare-ups
are more severe, and the condition progresses to becoming longterm. Severe flare-ups happen as a result of the complex interactions between the body’s immune system, the skin’s protective
barrier, and environmental factors such as stress.
The team examined approximately 500 clinical and experimental studies, and carried out research involving mice, to develop their mathematical model, which modelled four different
types of AD, ranging from no symptoms to severe flare-ups. They
also looked at how genetic factors affect the skin and the immune
system.
The researchers stress that theirs is a general model of AD.
However, they believe it suggests that preventing an immune system overreaction from being triggered in the first place may beThis article was redacted from Imperial College London
Press Release
http://www3.imperial.ac.uk/newsandeventspggrp/imperialcollege/newssummary/news_24-11-2016-15-14-54

6

the key to preventing AD from becoming more severe.
Two recent clinical trials demonstrated that babies who received moisturizing treatments, known as emollients, which wereapplied directly to the skin to reduce water loss and cover it witha protective film, were less like to develop AD. The authors of
this study also explained the possible mechanism behind theseobserved preventive effects. The emollients provide an effective
barrier, preventing the itching and scratching cycle that can progress the condition further. They also demonstrated that thesepreventive effects are applicable not only for babies with genetic
markers that indicate a predisposition to AD, but also for those
without genetic markers.
Dr. Reiko Tanaka at Imperial College London, said: “Ourmathematical modeling is helping us to make a clearer connection between a systemic reaction in AD and its progression. Morework needs to be done to verify our results in patients. However,we think a crucial preventative measure may be the use of ointments on all babies early on. It doesn’t matter which moisturizing
treatment is used as long as a barrier on the skin is created, which
may stop the AD cycle from triggering a systemic reaction.”
This approach of using mathematical models to understandthe underlying causes of diseases or conditions is called systems
medicine. Dr. Tanaka and her colleagues now hope to develop amodel that can be tailored to each patient based on their individual clinical data. This could enable doctors to tailor each treatment to the specific needs of the patients, optimizing its effectiveness and thus lessening the severity and impact of the condition.

Original paper:
Domínguez-Hüttinger E, Okada-Hatakeyama M, Kubo
M, Tanaka RJ et al. Mathematical modeling of atopic
dermatitis reveals “double switch” mechanisms underlying four common disease phenotypes. J Allergy Clin
Immunol S0091–6749(16)31433–6 (2016)

New mechanism of antiviralprotection

Research highlights

Figure: Antiviral protection by IgG2 antibody.

Influenza pandemics have become a serious public healthconcern in recent history and vaccination is critical for
preventing the spread of the influenza virus in the population. However, there is no vaccine that offers adequate
protection against emerging flu viruses. Thus, a better understanding of the mechanism of neutralizing antibody induction during influenza infection is essential to develop
alternative vaccines against emerging pandemic influenza.
It has been long believed that generation of high-affinity neutralizing antibody is critical for antiviral humoral
immunity. For the production of high-affinity antibodies,
formation of germinal centers (GC) in lymph nodes andthe presence of a subset of T cells called follicular helper T(TFH) cells in the GC are essential. Because of this, efficient
activation of TFH cells in GC was thought to be an essential
strategy for the development of antiviral vaccines.
However, a research team led by Masato Kubo of the
RIKEN Center for Integrative Medical Sciences discovered
a novel pathway that regulates induction of neutralizing antibodies independently of TFH cells and GC.
“Generation of not only TFH but also helper T1 (TH1)cells has been reported in the mouse model of acute influenza infection. So we decided to investigate the role of TH1
cells in protection against influenza virus,” says Kubo. They
used genetically modified mice lacking TFH cells or GC. To

their surprise, after vaccination these mutant mice were still
protected from a lethal dose of pandemic H1N1 or highly pathogenic H5N1 influenza A virus. The production ofhigh-affinity IgG1 antibody was massively reduced as a consequence of the loss of TFH cells and GC, however, the function of TH1 cells remained normal, and there was no effecton the production of IgG2 antibody. Although the avidityof the IgG2 antibody was reduced in the TFH/GC-deficient
mice, its neutralizing ability was not affected.
To confirm the role of TH1 cells in the production of IgG2
antibody, Kubo and colleagues isolated TH1 cells from thevaccinated mice and transferred them into genetically engineered mice that have a defect in IgG antibody production
due to the lack of helper T cells. Mice with transferred TH1
cells generated protective anti-viral IgG2 antibodies after influenza virus infection.
“Activation of TH1 cells resulted in production of cytokines such as IFN-γ and IL-21, and we found that thesecytokines were essential for production of IgG2 antibodies.
The IgG2 antibodies induced by TH1 cells have high neutralizing activity although their affinity is low. This insightshould contribute to new strategies to develop novel typesof vaccines against newly emerging influenza viruses” commented Kubo.

Original paper:
Miyauchi K, Sugimoto-Ishige A, Harada Y, Adachi Y,
Usami Y, Kaji T, Inoue K, Hasegawa H, Watanabe T, Hijikata A, Fukuyama S, Maemura T, Okada-Hatakeyama
M, Ohara O, Kawaoka Y, Takahashi Y, Takemori T, Kubo
M. Protective neutralizing influenza antibody responsein the absence of T follicular helper cells. Nat Immunol
17, 1447–1458 (2016)7

Mechanism of memory B celldifferentiation

Research highlights

Figure: Schematic view of memory B cell and plasma
cell development from GC B cells
The affinity of the B cell receptor for antigen and the expression level of Bach2 are inversely correlated in germinal center
B cells. Low affinity germinal center B cells, which receive
weaker stimulation from antigen and antigen-specific CD4 T
cells, maintain a relatively high level of Bach2 expression andtend to be induced to form memory B cells in the germinal center. On the other hand, high affinity cells, which receive strong
stimulation from antigen and antigen-specific CD4 T cells, havedifficulty maintaining Bach2 expression at high levels and tend
to be induced to form plasmablasts (plasma cells).

W

hen our body encounters a pathogen during an infection, our immune system remembers it even after
the infection has cleared. If we are re-exposed to the same
infection, a robust antibody response is quickly induced to
get rid of the pathogen. The “memory” B cell is the main
player in remembering pathogens and inducing faster protection from re-infection, however, it has been unclear how
memory B cells are generated and selected during the first
infection.
Ryo Shinnakasu and Tomohiro Kurosaki of RIKEN
Center for Integrative Medical Sciences and IFReC, Osaka
University, published a paper in Nature Immunology showing that memory B cells are predominantly generated during
the early phase of the germinal center (GC) response after
immunization. This was a surprising finding since it had
been thought that these cells arose later.
To understand the mechanism behind the memory B cell
induction, they first generated a transgenic mouse in which
they could irreversibly mark B cells in the GC, the site where
the B cells are responding to antigen, by administration of adrug called tamoxifen. After immunizing the mice, they ad-

Original paper:
Shinnakasu R, Inoue T, Kometani K, Moriyama S, Adachi
Y, Nakayama M, Takahashi Y, Fukuyama H, Okada T,
Kurosaki T. Regulated selection of germinal-center cells
into the memory B cell compartment. Nat Immunol
17, 861–869 (2016)8

ministered tamoxifen for three days starting at different time
points, days 5–7, 9–11, 19–21, and 29–31. This allowed themto analyze the generation of GC B cells and memory B cells
in a time-dependent manner. The results clearly indicatedthat memory B cells were generated mainly at the early stage,
9-11 days after immunization.
“Traditionally, it was believed that memory B cells induced late in the immune response have an advantage because they are derived from B cells with high-affinity for theantigen,” said Kurosaki. “Therefore, we analyzed the affinity
maturation of GC B cells and memory B cells 20 days after immunization. To our surprise, the results indicated that
low-affinity GC B cells are preferentially selected to become
memory B cells.”
They further found that high expression of the transcription factor Bach2 in GC B cells is important for memory
B cell differentiation. “Our findings have the potential tochange current vaccine strategies,” said Kurosaki. “Bach2
might be a novel target for the development of new vaccinesthat induce efficient differentiation of memory B cells from
low-affinity B cells.”

GWAS identifies seven novel locifor type 2 diabetes

Research highlights

Figure: Connection of type 2 diabetes risk genes
to drug targets. PPI, protein-protein interactions.

D

iabetes is one of the largest global health problems. Thereare currently an estimated 415 million adults in the world
with diabetes, and this figure will reach 642 million by 2040. The
Ministry of Health, Labour and Welfare estimates that more than
9.5 million Japanese are already diabetic or pre-diabetic, and 90%of them are affected by Type 2 diabetes due to insulin resistance
and/or insufficient insulin secretion.
Genome-wide association studies (GWAS) have identified
more than 80 susceptibility loci for type 2 diabetes, but mostof its heritability still remains to be elucidated. The relative importance of the causative mechanisms appears to differ between
Eastern and Western populations. Accumulating clinical evidence
suggests that defective insulin secretion contributes more to thepathogenesis of Japanese type 2 diabetes, whereas insulin resistance seems more important for European type 2 diabetes. Thus,
it is important to know the genetic differences in diabetes pathogenesis in Japanese and Europeans.
Minako Imamura and Shiro Maeda at RIKEN Center for Integrative Medical Sciences, in collaboration with Takashi Kadowaki and Toshimasa Yamauchi at Tokyo University Hospital, analyzed DNA samples of 15,463 Japanese type 2 diabetes patientsand 26,183 controls for 5.8 million SNPs. “This was the largest
sample size ever for GWAS studies of Japanese type 2 diabetes,”says Imamura. They subjected 17 candidate SNPs identified inthis GWAS to a second round of analysis, using a cohort of 7,936
patients and 5,536 controls. Based on these data, they identified
seven novel loci (CCDC85A, FAM60A, DMRTA1, ASB3, ATP8B2,

Original paper:
Imamura M, Takahashi A, Yamauchi T, Hara K, Yasuda
K, Grarup N, Zhao W, Wang X, Huerta-Chagoya A, Hu
C, Moon S, Long J, Kwak SH, Rasheed A, Saxena R, Ma
RC, Okada Y, Iwata M, Hosoe J, Shojima N, Iwasaki M,
Fujita H, Suzuki K, Danesh J, Jørgensen T, Jørgensen
ME, Witte DR, Brandslund I, Christensen C, Hansen T,
Mercader JM, Flannick J, Moreno-Macías H, Burtt NP,

MIR4686, and INAFM2) with a statistically significant linkage totype 2 diabetes.
They next analyzed the association of these seven loci withdisease susceptibility in populations other than Japanese. Whenthey examined the association of these loci with type 2 diabetes
on 65,936 patients and 158,030 controls from East Asian, South
Asian, European and Mexican populations, they found that five of
the seven loci were also linked to type 2 diabetes susceptibility inthese non-Japanese populations.
To search for potential drug targets for type 2 diabetes, they investigated 752 genes, 40 genes which lie on the 90 risk loci (the 7
new loci identified in this study and 83 previously identified loci)and 712 genes which have direct protein-protein interaction withthe products of the 40 genes, that overlap with drug target genes
corresponding to approved, clinical trial stage or experimental
drugs for various human diseases. Among the overlapping genes,
indeed, three genes that are targets for already-approved drugs
widely used for the treatment of type 2 diabetes. In addition,
they found drug targets that are under clinical trials: KIF11 fortreatment of cancer, GSK3B for leukemia and JUN for rheumatoid
arthritis; these drugs may be useful for the treatment of type 2
diabetes. “We could propose several new potential drug targets
for type 2 diabetes treatment using a systematic bioinformatics
approach. GWAS is still useful to identify novel susceptibility lociand, by integrating the findings with other genetic, biological andpharmacological studies, we hope to develop new treatments for
type 2 diabetes,” says Imamura.

Zhang R, Kim YJ, Zheng W, Singh JR, Tam CH, Hirose H,
Maegawa H, Ito C, Kaku K, Watada H, Tanaka Y, Tobe
K, Kawamori R, Kubo M, Cho YS, Chan JC, Sanghera D,
Frossard P, Park KS, Shu XO, Kim BJ, Florez JC, Tusié-Luna T, Jia W, Tai ES, Pedersen O, Saleheen D, Maeda S,
Kadowaki T. Genome-wide association studies in theJapanese population identify seven novel loci for type 2
diabetes. Nat Commun 7, 10531 (2016)

9

Sending bone growth down
the spine

Research highlights

Genetic variant causes unwanted bone growth
in a spinal disorder common among Asians

Figure: Patients with the spinal disorderossification of the posterior longitudinal ligament (OPLL) of the spine have aberrant bone
growth in the spine.
© Spondylolithesis/iStock/Getty Images PlusFor the first time, RIKEN researchers have shown how
a genetic variant increases the risk of a spinal disorderprevalent throughout Asia.
Ossification of the posterior longitudinal ligament of thespine (OPLL) is a degenerative disease that afflicts morethan 2% of people in Japan. It results when bone starts developing in the soft tissue of the spinal cord, leading to nerve
compression, pain and numbness—debilitating symptoms
that can severely affect daily life.
Over the years, scientists have found several gene variants that occur more often in people with OPLL, but it was
not known if or how any of these genes directly contributed
to the patients’ aberrant bone growth. Now a team led by
Shiro Ikegawa at the RIKEN Center for Integrative Medical
Sciences tested the role of one potential susceptibility gene,
known as RSPO2.
The team previously conducted a comprehensive gene
sleuthing study, which found six sites in the genome that
seemed to confer OPLL disease risk. One of these genomic segments included the gene RSPO2, which carries the
instructions for making a protein called R-spondin 2. The
protein was known to play a critical role in skeletal development, but the researchers wanted to drill deeper into how
RSPO2 works at a molecular level. They used mouse and human bone precursor cells to examine gene expression in alab dish.
They showed that RSPO2 normally acts to put the brakes

This article was reproduced from RIKEN Research
http://www.riken.jp/en/research/rikenresearch/highlights/8262/

10

on bone formation by inhibiting the development of the early cartilage cells, which are later replaced by bone cells. However, the gene variant linked to OPLL decreases gene activity by changing the promoter region—the ‘on switch’—of
RSPO2. Specifically, the disease-associated variant alters thebinding of a transcription factor that would otherwise allow
RSPO2 expression and thereby halt bone growth. Without
this binding, stem cells in the spinal ligament get misdirected into bone.
“Our findings provide new insights into the etiology andpathogenesis of OPLL, as well as a new target for treatmentof the disease,” says Masahiro Nakajima, who conducted thestudy. If drug companies can find a way to block the pathway
unleashed by low RSPO2 expression, Nakajima explains,
they could help improve the lives of people with the disease.
The team plans to investigate the role of other potential
OPLL genes in search of additional drug targets. “We believe
it is possible to find other causal variants and susceptibility
genes using the same approach,” Nakajima says.

Original article:
Nakajima M, Kou I, Ohashi H, Genetic Study Group of
the Investigation Committee on the Ossification of Spinal Ligaments, Ikegawa S. Identification and functional
characterization of RSPO2 as a susceptibility gene forossification of the posterior longitudinal ligament of
the spine. Am J Hum Genet 99, 202–207 (2016)

GWAS identifies a genetic risk
factor associated with clozapineinduced agranulocytosis/
granulocytopenia

Research highlights

Figure: A Manhattan plot of GWAS results for
CIAG in the Japanese population. The horizontal
axis indicates each SNP’s chromosomal location,
while the vertical axis indicates the -log10 (Pvalue) of the association. SNPs positioned higher
on the plot have a more reliable association with
CIAG.

S

chizophrenia is a chronic and serious mental disorderthat affects how a person thinks, feels and behaves.
Its lifetime prevalence is 1% of the world population, andthere are 700–800 thousand patients in Japan sufferingfrom schizophrenia. The most useful therapeutic option for
schizophrenia is antipsychotics; however, approximately one
third of patients do not respond adequately to these medications, resulting in treatment-resistant schizophrenia.
Clozapine, a second generation “atypical” antipsychotic,
is a gold standard drug for treatment-resistant schizophrenia. It has a lower incidence of movement abnormalities andits efficacy is superior to that of other atypical antipsychotics for treatment-resistant schizophrenia. Despite these advantages, clozapine may have side effects, some serious andpotentially fatal, and therefore the use of clozapine is significantly restricted.
Agranulocytosis or granulocytopenia is an acute and severe leukopenia, potentially life threatening. About 1–3% of
people who take clozapine develop the side effect of clozapine-induced agranulocytosis (CIA)/granulocytopenia (CIG)(CIAG). Several pharmacogenetic/pharmacogenomic studies in European populations suggested that certain genetic

Original paper:
Saito T, Ikeda M, Mushiroda T, Ozeki T, Kondo K,
Shimasaki A, Kawase K, Hashimoto S, Yamamori H,
Yasuda Y, Fujimoto M, Ohi K, Takeda M, Kamatani Y,
Numata S, Ohmori T, Ueno S, Makinodan M, Nishihata
Y, Kubota M, Kimura T, Kanahara N, Hashimoto N,
Fujita K, Nemoto K, Fukao T, Suwa T, Noda T, Yada Y,
Takaki M, Kida N, Otsuru T, Murakami M, Takahashi

polymorphisms in the MHC (HLA) region may be associated with CIAG.
Taisei Mushiroda of the RIKEN Center for Integrative
Medical Sciences, in collaboration with researchers at Fujita Health University and Osaka University, analyzed 900
thousand SNPs in genomic DNA samples from 50 Japanesepatients with CIAG and 2,905 controls. They identified four
SNPs in the HLA region on chromosome 6 associated with
CIAG (Figure). To determine the associated HLA allele, theyfurther conducted HLA typing. As a result, they found that a
specific allele named HLA-B*59:01 was significantly associated with CIAG in Japanese.
In addition, they explored the risk of developing CIA inclozapine rechallenge treatment. “Our results indicated thatclozapine rechallenge is a possible option for patients who
respond only to clozapine,” say the research group. “Our explorative estimates showed that about 60% of CIG patients
without the HLA-B*59:01 allele have the potential to avoid
future CIA. Of course, frequent monitoring of the whiteblood cells and absolute neutrophil count is essential during
the rechallenge.”

A, Kubo M, Hashimoto R, Iwata N. Pharmacogenomic
Study of Clozapine-Induced Agranulocytosis/Granulocytopenia in a Japanese Population. Biol Psychiatry 80,
636–642 (2016 )11Research highlights

Keeping viral DNA at bay

Scientists find that an epigenetic interplay keeps
virus-derived DNA sequences repressed in embryos
but not in the surrounding placenta

Figure: Wild-type (left), Dnmt1-deficient (center)and Np95-deficient (right) mouse embryos. Only
the Dnmt1 mutant shows activity of endogenous
retroviruses, indicated by the blue stain.
© ElsevierT

he human genome is riddled with virus-derived sequences considered to be remnants of viral infections
during our evolutionary past. A new study by RIKEN researchers explains how these so-called endogenous retroviruses (ERVs) are repressed in the developing embryo, butalso finds that they spring into action in the cells of the placenta.
“In my field, people mostly think that ERVs are harmful
and therefore should be silenced in all cells,” says Jafar Sharifof the RIKEN Center for Integrative Medical Sciences, who
led the study with colleagues at RIKEN and collaborators inJapan, Canada and the US. “But our work shows that ERV
expression takes place even under perfectly normal conditions, such as in the placenta during development.”
Sharif and his colleagues first wanted to clarify how theseviral sequences are silenced in the embryonic lineage.
An enzyme called DNMT1 was known to help to inactivate these rogue genetic elements by ensuring that methyl
tags are placed in the right locations on the DNA backbone
to maintain proper gene expression and regulation.
When functioning DNMT1 is absent, the viral elements
kick into action, which was thought to be due to a lack of
DNA methylation. But that turned out to be only part of thestory. The RIKEN team has shown that the partially methylated DNA wrought by depletion of DNMT1 actually at-

This article is reproduced from RIKEN Research
http://www.riken.jp/en/research/rikenresearch/highlights/8230/

12

taches itself to a second protein called NP95, which triggers
the regulatory cascade responsible for releasing the brakes
on viral repression.
The researchers discovered that prolonged binding with
NP95 disrupted the way DNA was packaged into chromatin fibers and this allowed ERVs to be expressed. They confirmed the crucial role of Np95 by removing the protein inmouse embryos, either alone or in combination with Dnmt1
removal. In both cases, the viruses remained dormant(Figure, the Dnmt1/Np95 double-deficient results are notshown).
“I was very surprised,” says Sharif of this unexpected finding. “In fact, at first I thought my experiments were not going well when I found deleting the Np95 gene together with
the Dnmt1 gene inexplicably extinguished the activation of
ERVs.”
But more surprising was that the Np95 protein and ERVs
are highly expressed in placental cells.
This discrepancy between ERV activity in embryonic andextra-embryonic tissues remains a mystery. “There is still nophysiological explanation for why these ERVs are expressedin the placenta,” notes Sharif, who intends to find out why. “Ilike to think that ERV expression must have some biologicalmeaning, or else it would not be tolerated in the placenta,”
Sharif says.

Original paper:
Sharif J, Endo TA, Nakayama M, Karimi MM, Shimada
M, Katsuyama K, Goyal P, Brind'Amour J, Sun MA, Sun
Z, Ishikura T, Mizutani-Koseki Y, Ohara O, Shinkai Y,
Nakanishi M, Xie H, Lorincz MC, Koseki H.
Activation of endogenous retroviruses in Dnmt1−/− ESCs
involves disruption of SETDB1-mediated repression by
NP95 binding to hemimethylated DNA. Cell Stem Cell

19, 81-94 (2016)

Part 2Lab Activities

Core for
Homeostatic
RegulationT

he ultimate goal of the Core for Homeostatic Regulation is to elucidate
the mechanisms of onset of human diseases and to create new scientificparadigms. This Core clarifies the regulation of homeostasis in individuals,
focusing on their immune, metabolic and environmental response systems.
In addition, the Core for Homeostatic Regulation will validate the diseasemodels established by the Core for Precise Measuring and Modeling in a multitier timeframe from before to after the onset of diseases.
There are 15 laboratories in the Core for Homeostatic Regulation which
are divided into four areas;[1] Immune homeostasis
Cell signaling (T. Saito), Lymphocyte differentiation (T. Kurosaki), Immune
homeostasis (S. Hori), Metabolic homeostasis (N. Kubota)[2] Lymphocyte development
Transcriptional regulation (I. Taniuchi), Human disease models (F. Ishikawa)[3] Mucosal immunity
Intestinal ecosystem (H. Ohno), Mucosal immunity (S. Fagarasan), Immune
cell systems (S. Koyasu), Gut homeostasis (K. Honda), Immune crosstalk (H.
Cheroutre)[4] Allergy and inflammation
Skin homeostasis (M. Amagai), Inflammatory regulation (T. Tanaka), Cytokine
regulation (M. Kubo), Innate immune systems (K. Moro)
All of these areas elucidate the basic mechanisms of immune regulation
at cellular tissue and systemic levels. We ultimately aim to analyze the onset
of autoimmune diseases, metabolic disorders [1], primary immunodeficiency[2], inflammatory bowel disease and colitis [3], and atopic dermatitis andallergic diseases [4].

14

Laboratory for

Lab activities

Cell Signaling
Group Director: Takashi Saito

Figure: The microsynapse, composed of a microadhesion ring surrounding a TCR microcluster, isessential for T cell activation[A] The Pyk2 focal adhesion molecule (green) forms
a micro-adhesion ring around TCR-microclusters (red)upon T cell stimulation[B] Schematic diagram of a microsynapse. The microsynapse, composed of a micro-adhesion ring around a
TCR-microcluster, is formed during early activation andfacilitates formation of a microcluster and cSMAC andin T cell activation, particularly under weak stimulation.
[C] An adhesion-ring in a microsynapse induces outside-in signals through LFA1/focal adhesion molecules,
thus helping induce activation signals through TCR-microclusters.

Recent Major Publications
Ishikawa E, Kosako H, Yasuda T, Ohmuraya M, Araki K,
Kurosaki T, Saito T, Yamasaki S. Protein kinase D regulates positive selection of CD4+ thymocytes throughphosphorylation of SHP-1. Nat Commun 7, 12756(2016)
Hashimoto-Tane A, Sakuma M, Ike H, Yokosuka T,
Kimura Y, Ohara O, Saito T. The Micro adhesion-ringsurrounding each TCR microclusters forms synapse-likestructure essential for T cell activation. J Exp Med 213,
1609–1625 (2016)
Takeuchi A, Badr MESG, Miyauchi K, Ishihara C, Onishi
R, Guo Z, Sasaki T, Ike H, Takumi A, Tsuji NM, Murakami
Y, Katakai T, Saito T. CRTAM determines the CD4+ cytotoxic T lymphocyte lineage. J Exp Med 213, 123–138(2016)

Invited Presentations
Saito T. “Single cell molecular imaging for T cell activation” The 54th Annual Meeting of the Biophysical Society of Japan (Tsukuba, Japan) November, 2016
Saito T. “Microsynapse composed of micro-adhesion
ring surrounding TCR microcluster is essential for T cell
activation” EMBO conference (Siena, Italy) September,
2016
Saito T, Hashimoto-Tane A. “Microsynapse composed offocal adhesion molecules surrounding TCR microcluster
is essential for T cell activation” International Congressof Immunology 2016 (Melbourne, Australia) August,
2016
Saito T. “Dynamic Regulation of T cell Activation at
Immune Synapse” POSTEC Symposium (Pohang, Korea)
July, 2016
Saito T. “Development and function of CD4+CTL in
inflammation diseases” SICORP Japan-New Zealand
Joint Research on Functional Foods (Wellington, New
Zealand) February, 2016T

cells play central roles in immune regulation. They initiate immune responses, induce activation and generate various effector T cells, which protect against infection
and oncogenesis. Aberrant T cell function results in infectious, cancer and autoimmune
diseases. Our group aims to determine the molecular mechanism of T cell activation,differentiation and homeostasis, particularly from a signaling perspective.
T cell activation is induced through TCR-microclusters (MC), the signaling clusters
generated by recruiting TCR and proximal signaling molecules. We found that TCR-MCs
at the initial activation stage are surrounded by an adhesion-ring composed of integrin and focal adhesion molecules. This structure resembles an immune synapse in micro-scale, and is termed the microsynapse (Figure). The microsynapse supports integrin
outside-in signaling, which facilitates formation of the MC and cSMAC and then T cell
activation. The microsynapse is particularly important for activation under weak stimulation condition. Thus, the microsynapse and MCs seem to be the major membrane structures for inducing an activation signal in T cells.
We also investigate the regulation of T cell activation by innate signals. We have analyzed the function of TLRs and nucleic acid recognition in T cells. STING is known to
be a major intracellular DNA sensor in innate cells. Since STING is highly expressed in
T cells, its function in T cell activation was analyzed. Surprisingly, STING stimulationinduced inhibition of T cell proliferation and anti-viral responses. Thus, STING ligands
may serve as specific inhibitors of activated T cells.
The ultimate aim of our diverse approaches is to elucidate the mechanisms of inducing
autoimmune diseases through aberrant T cell function in order to be able to modulatethem and inhibit/prevent autoimmunity and allergic inflammation. We are analyzing
the function and regulation of T cells by phosphatases (PTPN22, PTPN2), whose deficiency is associated with induction of autoimmune diseases. The onset of autoimmunity
will be analyzed in gene-targeted mouse models.

15

Laboratory for

Lab activities

Lymphocyte
Differentiation
Group Director: Tomohiro Kurosaki

Figure: Foxo1 controls GC B cell proliferation in
response to T cell help
(Left) Schematic illustration of Foxo1 function in GC B
cells. Foxo1 controls GC B cell proliferation (1), partly by
mediating BATF up-regulation in the LZ B cells (2). In
the LZ B cells, Foxo1 is also required for antigen presentation to TFH cells (3) and up-regulation of the DZ program genes in response to T cell help (4), which triggers
the switch from the LZ to the DZ. (Right) Immunohistochemical analysis of spleen sections showing the GC
compartmentalization defect resulting from Foxo1-ablation. Expression of CD45.1(Foxo1+/+ or Foxo1-/- derived
donor cells), CD35 (FDC network), and IgD (follicular B
cells) are shown.

Recent Major Publications
Inoue T, Shinnakasu R, Ise W, Kawai C, Egawa T, Kurosaki T. The transcription factor Foxo1 controls germinal
center B cell proliferation in response to T cell help. J
Exp Med 214, 1181–1198 (2017)
Igarashi K, Kurosaki T, Roychoudhuri R. BACH familytranscription factors in innate and adaptive immunity.
Nat Rev Immunol (in press)
Shinnakasu R, Inoue T, Kometani K, Moriyama S, Adachi
Y, Nakayama M, Takahashi Y, Fukuyama H, Okada T,
Kurosaki T. Regulated selection of germinal-center cells
into the memory B cell compartment. Nat Immunol
17, 861–869 (2016)

Invited Presentations
Kurosaki, T. “Fate decisions of germinal center B cells
into the memory B cell or plasma cell compartment”
4th Antibody Symposium (Singapore, Singapore) November, 2016
Kurosaki, T. “Overview of germinal center immunity”
The 13th International Workshop on Autoantibodies
and Autoimmunity (IWAA2016) (Kyoto, Japan) October,
2016
Kurosaki, T. “Transcriptional regulation in memory B cell
development” EMBO Conference: Lymphocyte Antigen
Receptor Signaling (Siena, Italy) September, 2016
Kurosaki, T. “Instructive selection of germinal center
B cells into the memory compartment” International
Congress of Immunology 2016 (Melbourne, Australia)
August, 2016
Kurosaki, T. “Instructive selection of germinal center B
cells into the memory compartment” Antibody and Fc
Receptor Biology: Bench to Bedside, A Special Symposium to Honor Jeffrey V. Ravetch on his 65th Birthday(New York, USA) May, 201616

H

umoral memory relies on the development of memory B cells and longlived plasma cells, and the vast majority of these cells are derived from germinal center (GC) B cells. Therefore, our lab has been focusing on characterizingof these three cell types and clarifying how memory B and plasma cells are generated through GC reactions.
GC B cells cycle between two states, the light zone (LZ) and the dark zone (DZ)
and, in the latter, they proliferate and hypemutate their immunoglobulin V genes(Igs). How this functional transition takes place is still controversial. We foundthat ablation of the transcription factor Foxo1 after GC development led to loss of
the DZ GC B cells and disruption of the GC architecture (Figure). Mechanistically,
even upon provision of adequate T cell help, Foxo1-deficient GC B cells manifested less proliferative expansion than controls. Moreover, we found that the transcription factor BATF was transiently induced in a small subset of GC B cells in a
Foxo1-dependent manner and that deletion of BATF similarly led to GC disruption. Thus, the switch from the LZ to the DZ is triggered after receipt of T cell help,
and Foxo1-mediated BATF up-regulation is at least partly involved in this switch.
In regard to differentiation of GC B cells into long-lived plasma cells, only highaffinity GC B cells are selected into the plasma cell pool, however the underlying
mechanism is still obscure. We found that the IRF4hiBcl6lo LZ GC B cell subset
with high affinity BCRs favored the plasma cell fate over GC cycling. Given thatIRF4 acts to facilitate differentiation towards plasma cells in a dose-dependent
manner, our data suggest that a strong BCR signal and T cell help modulate theexpression of IRF4 and Bcl6, thereby facilitating plasma cell differentiation andantagonizing the GC program.

Laboratory for

Lab activities

Transcriptional
Regulation
Group Director: Ichiro Taniuchi

Figure: The evolutionarily conserved Cbfβ2 variant confers thymus-homing capacity.
In mutant mice lacking Cbfβ2, which is one of two RNA
splice variants encoded by the Cbfb gene, differentiation of IL7R+PIR+ fetal liver cells and induction of Ccr9 in
those cells are impaired (A). Cbfβ2 binds predominantly
to two upstream regions in the Ccr9 gene (B). Removalof these two regions eliminated CCR9 expression specifically in PIR+ but not PIR– cells (C).

Recent Major Publications
Seo W, Muroi S, Akiyama K, Taniuchi I. Distinct requirement of Runx complexes for TCRβ enhancer activation
at distinct developmental stages. Sci Rep 7, 41351(2017)
Kitagawa Y, Ohkura N, Kidani Y, Vandenbon A, Hirota
K, Kawakami R, Yasuda K, Motooka D, Nakamura S,
Kondo M, Taniuchi I, Kohwi-Shigematsu T, Sakaguchi S.
Guidance of regulatory T cell development by Satb1-dependent super-enhancer establishment. Nat Immunol
18, 173–183 (2017)
Taniuchi I. Views on helper/cytotoxic lineage choicefrom a bottom-up approach. Immunol Review 271,
98–113 (2016)

Invited Presentations
Taniuchi I. “Regulation of Runx Complex Function by
Modulating C-terminal Sequences in Its Two Components” Cancer Science Institute of Singapore. (Singapore, Singapore) November, 2016
Taniuchi I. “Regulation of T cell development in thethymus by transcription factors.” CELL AND DEVELOPMENTAL BIOLOGY SEMINAR SERIES at University ofMichigan. (Arbor, USA) October, 2016
Taniuchi I. “Regulation of Langerhans cell development
by Runx transcription factor complexes.” Immunological
Diseases and Translation Research Forum 2016 (Shanghai, China) October, 2016
Taniuchi I. “Transcription factors in regulation of T cell
development and function.” The first RIKEN-IITU Works(Beijing, China) September, 2016
Taniuchi, I. “T cell development in the thymus” The 26th
Japan Metrology Association Academic Conference y(Fukuoka, Japan) July, 2016T

he vertebrate immune system consists of two components, innate and acquired. The acquired immune system is responsible for antigen-specific recall
responses, is the principle for vaccination, and appeared latter during evolution, atleast by acquisition of the system for generating pools of lymphocytes with a broad
variety of antigen-specificities. Thus, one of the major questions in developmental
biology is how the genetic program that generates a primary lymphoid organ, thethymus, for supporting T lymphocyte development has been exploited.
My laboratory has been addressing functions of Runx transcription factor complexes, which consist of a Runx protein and a non-DNA binding Cbfβ protein. Our
recent work provided novel insight into how modulation of Runx complexes contributed to formation of lymphoid organs/tissues. Two functional Cbfβ variants,
Cbfβ1 and Cbfβ2, are generated from the mammalian Cbfb gene by alternative
RNA splicing. We found that the Cbfβ2 variant is essential not only for differentiation of thymic progenitors, defined as IL7R+PIR+ fetal liver cells, but also for endowing them with thymus-homing capacity through activating cell-type specificenhancers in the Ccr9 gene, which encodes the primary thymus homing receptor.
Our comparative genomics analyses revealed that RNA splicing, which produces
Cbfβ2, was likely to have emerged at some point in the evolution toward bony
fish, when the thymus appeared in its current form. Thus, acquisition of novel
splice variants of the Cbfb gene established regulation that allows firm interaction
between hematopoietic and stromal cells to generate a primary lymphoid organ,
the thymus. In addition, our results suggest that this ancestral regulation was likely co-opted later to exploit development of lymphoid tissue inducer (LTi) cells
to generate secondary lymphoid tissues. These results illustrate how an increasein the functional diversity of transcription factors was adapted to modulate theimmune system.

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Immune Cell Systems
Group Director : Shigeo Koyasu

Figure: ILC2 in the small intestine are involved in
the induction of diet-induced obesity.
While Rag2 –/– mice lacking acquired immune cells gain
weight upon high-fat diet feeding, Il2rg –/–Rag2 –/– micelacking all lymphocytes including innate lymphoid cells(ILCs) are resistant to diet-induced obesity. Adoptive
transfer of ILC2 from small intestine but not white
adipose tissue restored diet-induced obesity in Il2rg –/–Rag2–/– mice.

Recent Major Publications
Morita H, Moro K, Koyasu S. Innate lymphoid cells inallergic and non-allergic inflammation. J Allergy Clin
Immunol 138, 1253–1264 (2016)
Yamazumi Y, Sasaki O, Imamura M, Oda T, Ohno Y,
Shiozaki-Sato Y, Nagai S, Suyama Kamoshita S, Funato
Y, Yasui K, Kikutani T, Yamamoto K, Dohi M, Koyasu S,
Akiyama T. The RNA binding protein Mex-3B is required
for IL-33 induction in the development of allergic airway inflammation. Cell Rep 16, 2456–2471 (2016)

Invited Presentations
Koyasu S. “IL-7 and Notch conditionally control the commitment of ILC2 differentiation” 2nd EMBO Conference“Innate lymphoid cells” (Berlin, Germany) December,
2016
Koyasu S. “Regulatory mechanisms for ILC2 activationin allergic inflammation” 4th International Cytokine andInterferon Society (San Francisco, USA) October, 2016
Koyasu S. “Innate lymphoid cells and inflammation”
Cold Spring Harbor Asia Symposium on “Frontiers of
Immunology in Health and Disease” (Awaji, Hyogo)
October, 2016
Koyasu S. “Innate lymphoid cells and inflammation” 5thInternational GK Symposium on Regulators of Adaptive
Immunity (Erlangen, Germany) September, 201618

W

e have been working on the role of the natural helper (NH) cell, one of
the group 2 innate lymphoid cells (ILC2). Because we originally discovered this population in the mesentery, one of largest adipose tissues in the body,
we have been interested in the role of ILC2 in adipose tissue homeostasis. Obesity induces adipose tissue inflammation associated with infiltration of immune
cells, such as T cells, B cells and macrophages, into adipose tissues and reduction of Tregs. Although it is believed that complex interactions of immune cells
are involved in the induction of obesity, the involvement of the immune system,
specifically lymphocytes, has not been fully explored before. We found that Il2rg–/–
Rag2–/– mice, lacking all lymphocytes, but not Rag2–/– mice, lacking only acquired
immune cells, were resistant to diet-induced obesity. Transplantation of Rag2–/–
bone marrow cells into Il2rg–/–Rag2–/– mice abolished this resistance, indicating the
involvement of ILCs. Mice lacking ILC2 or ILC3 but not NK cells were resistant toobesity, with stronger phenotypes in mice lacking only ILC2. Adoptive transfer ofnaïve ILC2 isolated from small intestine (SI) but not white adipose tissue (WAT)
restored the induction of diet-induced obesity in Il2rg–/–Rag2–/– mice. These results
suggest a role of small intestinal ILC2s in the induction of obesity.

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Human Disease Models
Group Director & Chief Scientist: Fumihiko Ishikawa

Figure: Creating new therapeutic strategies
targeting poor prognosis leukemia through amulti-faceted approach
To understand the in vivo behavior of normal and leukemic stem cells, we have developed humanized mousesystems. Using the humanized mice, we identified
where human stem cells home, how human AML develops, and how human hematopoietic stem cells generate myeloid and lymphoid progeny in vivo. Throughthis multi-faceted approach integrating medicinal
chemistry, computational science, and protein structural analysis, we have been identifying new drug candidates targeting human leukemia. We hope to translate
humanized mouse research into future medicine.

Recent Major Publications
Najima Y, Tomizawa-Murasawa M, Saito Y, Watanabe
T, Ono R, Ochi T, Suzuki N, Fujiwara H, Ohara O, Shultz
LD, Yasukawa M, Ishikawa F. Induction of WT1-specifichuman CD8+ T cells from human HSCs in HLA class I Tg
NOD/SCID/IL2rgKO mice. Blood 127, 722–734 (2016)
Aoki Y, Watanabe T, Saito Y, Kuroki Y, Hijikata A, Takagi
M, Tomizawa D, Eguchi M, Eguchi-Ishimae M, Kaneko
A, Ono R, Sato K, Suzuki N, Fujiki S, Koh K, Ishii E, Shultz
LD, Ohara O, Mizutani S, Ishikawa F. Identification of
CD34+ and CD34– leukemia-initiating cells in MLL-rearranged human acute lymphoblastic leukemia. Blood
125, 967–980 (2015)

Invited Presentations
Ishikawa F. “Exploring complexity and heterogeneity ofhuman hematopoiesis in vivo” (Hawaii, USA) February,
2017
Ishikawa F. “Targeting acute myeloid leukemia withgenetic complexity and heterogeneity” 75th Annual
Meeting for Japan Cancer Association (Yokohama,Japan) October, 2016
Ishikawa F. “Toward understanding human immunityand diseases” Merck Meeting for Preclinical Models for
Immunotherapy (San Francisco, USA) September, 2016T

he specific aim of our laboratory has been to understand normal and diseased
human hematopoiesis and immunity and to apply our research findings toclinical medicine. To this end, we have developed humanized mouse models byintravenously injecting human normal and disease-initiating stem cells into immune-compromised NOD/SCID/Il2rgKO (NSG) newborns. By injecting normal
human hematopoietic stem cells, we have succeeded in reconstituting multiple
organs of the recipient mice with human acquired and innate immunity. Furthermore, to better understand human hematopoiesis and immunity, we have createda new generation humanized mice expressing human molecules that serve essential roles in mediating environmental signaling. Using these new humanized mice,
we are examining roles of human cytokines and adhesion molecules in differentiation, maturation, and function of human immune cell types.
Among various human diseases, we have been focusing on poor prognosis
leukemia. Through identification of leukemia stem cells responsible for disease
initiation and relapse, we have been searching for small molecules and antibodies
that can eradicate human leukemia at the stem cell level. We found that RK-20449,
a dual kinase inhibitor for HCK and FLT3, could effectively target FLT3-ITD mutated acute myeloid leukemia in bone marrow, spleen, and peripheral blood. We
will further assess the therapeutic and adverse effects of RK-20449 and its related compounds toward establishment of a new therapeutic strategy for FLT3-ITD
mutated AML.

Ishikawa F. “Exploring complexity and heterogeneityof human acute myeloid leukemia” 2016 the JapaneseSociety of Medical Oncology Annual Meeting (Kobe,
Japan) July, 2016

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Intestinal Ecosystem
Group Director: Hiroshi Ohno

Figure: Schematic representation of the mechanism for tunneling nanotube-mediated HIV-1
spreading by direct intercellular transmissionFrom top to bottom, i) HIV-1 infects a macrophage, ii)
HIV-1 replication, iii) formation of a tunneling nanotube, an intercellular connection of the plasma membranes via interaction of HIV-1 Nef protein and host cell
M-Sec, iv) transmission of HIV-1 from an infected to a
non-infected macrophage, v) replication of HIV-1 in the
newly infected macrophage.

Recent Major Publications
Okai S, Usui F, Yokota S, Hori-I Y, Hasegawa M, Nakamura T, Kurosawa M, Okada S, Yamamoto K, Nishiyama
E, Mori H, Yamada T, Kurokawa K, Matsumoto S, Nanno
M, Naito T, Watanabe Y, Kato T, Miyauchi E, Ohno H,
Shinkura R. High-affinity monoclonal IgA regulates gut
microbiota and prevents colitis in mice. Nat Microbiol
1, 16103 (2016)
Nakato G, Hase K, Sato T, Kimura S, Sakakibara S,
Sugiyama M, Obata Y, Hanazato M, Iwanaga T, Ohno H.
Epithelium-Intrinsic MicroRNAs Contribute to Mucosal
Immune Homeostasis by Promoting M-Cell Maturation.
PLoS One 11, e0150379 (2016)
Hashimoto M, Bhuyan F, Hiyoshi M, Noyori O, Nasser H,
Miyazaki M, Saito T, Kondoh Y, Osada H, Kimura S, Hase
K, Ohno H, Suzu S. Potential Role of the Formation of
Tunneling Nanotubes in HIV-1 Spread in Macrophages.
J Immunol 196, 1832–1841 (2016)

Invited Presentations
Ohno H. “Gut microbiota and human diseases” International Society of Cardiomyopathies and Heart Failure(ISCHF) Congress 2016 (Kyoto, Japan) December, 2016
Ohno H. “Regulation by the gut microbiota of the intestinal immune system: the largest immune system in thebody” The 6th Chemical Festa of the Chemical Society ofJapan (Tokyo, Japan) November, 2016
Ohno H. “Human gut microbiota as novel therapeutictargets-An overview-” BioJapan 2016 (Yokohama, Japan) October 2016
Ohno H. “Integrated multi-omics approach for understanding the gut ecosystem” International Conference
on Beneficial Microbes (ICOBM) 2016 (Phuket, Thailand) May-June, 2016
Ohno H. “Gut Microbiota, Host Defense and Immunity”
The 5th NIF Winter School on Advanced Immunology(Awaji, Japan) January, 201620

E

normous numbers of bacteria, collectively called the gut microbiota, reside in
our intestines; nevertheless, the gut does not unconditionally accept commensal microorganisms. The intestinal immune system somehow senses the typeand quantity of bacteria in the gut lumen and tries to contain them. Reciprocally,
the gut microbiota shape the host immune system, and the host-gut microbiota
interaction deeply impacts host physiology and pathology. The aim of our laboratory is to understand the mechanisms by which the host and its gut microbiota
interact, especially focusing on how gut microbes are delivered across the intestinal epithelial barrier to be recognized by the intestinal immune system, how thegut microbiota shape host defense and immune system, and how host-gut microbiota interactions affect host health and disease status.
The delivery of particulate antigens such as bacteria is thought to be mainly
achieved by a unique subset of intestinal epithelial cells, M cells. These reside in alimited region of the epithelial layer called the follicle-associated epithelium(FAE), which covers the lymphoid follicles of gut-associated lymphoid tissue(GALT), such as Peyer’s patches (PP). In 2016, we reported the identification ofepithelium-intrinsic microRNAs important for M-cell maturation.
Regarding the host-gut microbiota interactions, we are employing an integrated multiple omics approach, combining exhaustive (meta)genomic, (meta)transcriptomic, and metabolomic analyses. In collaboration with Professor Shinkura,
we have reported that high-affinity IgA regulates the gut microbiota and prevents
colitis in mice. The integrated multiple omics approach is currently being applied
to understand the impact of the gut microbiota on human diseases such as infantile allergic diseases and type 2 diabetes.
We have also shown that tunneling nanotubes are important for HIV-1 tospread directly through intercellular connections among macrophages (Figure).

Laboratory for

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Mucosal Immunity
Team Leader: Sidonia Fagarasan

Figure: System immunology
A scheme portraying the systemic effects of T cell activation in PD-1 deficient mice. Activation of T cells in theabsence of PD-1 leads not only to abnormal expansionof T cells with effector/memory phenotype but also toa series of modifications leading to microbiota, serum
metabolome and metabolic changes as well as biochemical shifts in the central nervous system. We aim
to uncover the mechanisms leading to such systemic
effects of immune activation.

Recent Major Publications
Chamoto K, Chowdhury PS, Kumar A, Fagarasan S,
Honjo T. Mitochondria activation chemicals synergize
with PD-1 checkpoint blockade for T cell-dependentanti-tumor activity. Proc Natl Acad Sci U S A 114,
E761–E770 (2017)
Zhang B, Chikuma S, Hori S, Fagarasan S, Honjo T.
Nonoverlapping roles of PD-1 and FoxP3 in maintaining
immune tolerance in a novel autoimmune pancreatitis mouse model. Proc Natl Acad Sci U S A 113,
8490–8495 (2016)
Sutherland DB, Suzuki K, Fagarasan S. Fostering ofadvanced mutualism with gut microbiota by Immunoglobulin A. Immunol Rev 270, 20–31 (2016)

Invited Presentations
Fagarasan S. “PD-1, a key regulator of immune-brain
communication” The 2016 Kyoto Prize Workshop in
Basic Sciences: “From the Molecular Immunity to theSuppression of Cancer” (Kyoto, Japan) November, 2016
Fagarasan S. “Involvement of PD-1 in antibody diversification and body homeostasis” Japan-UK ClinicalTranslational Research Symposium: Metabolic profilingfor precision medicine (Tokyo, Japan) October, 2016
Fagarasan S. “IgA and T cells: Regulators of gut andsystemic homeostasis” The 5th International GK symposium Regulators of adaptive immunity (Erlangen,Germany) September, 2016
Fagarasan S. “On germinal centers, IgA synthesis andthe intestinal ecosystem”, International Congress of
Immunology ICI 2016: B cells (Melbourne, Australia)
August, 2016
Fagarasan S. “On germinal centers, IgA synthesis andthe intestinal ecosystem” Keystone Symposia: T Follicular Helper Cells and Germinal Centers (Monterey, USA)
March, 2016A

ctivation of T cells is accompanied by metabolic reprogramming for energy andbiosynthesis necessary for sustaining expansion, survival and effector functions.
The consequences of such T cell reprogramming for immunity are widely accepted. However, it is currently unknown whether T cell activation impacts on systemic metabolic
homeostasis, which would then affect the function of other major physiological systemsof the body. This is the fundamental question our laboratory aims to answer.
Blockade of the inhibitory receptor PD-1 by antibodies reinvigorates T cell immunityand boosts antitumor activity, making PD-1 an important target in cancer immunotherapy. We exploit PD-1 deficiency as a mouse model of sustained immune activation andused a systemic approach to evaluate the metabolic outcome of T cell activation.
We find that persistent T cell activation in mice lacking PD-1 results in dramatic
systemic metabolome changes and that such changes affect other major physiologicalsystem of the body, such as the central nervous system. Strikingly, in a tumor model wealso find that activation of T cells in the presence of PD-1 blockade, even for a short period of time, reproduces the metabolic phenotype of PD-1 deficient mice.
We are interested in further exploring the mechanisms leading to the systemic metabolic shift as a basis to understand the systemic pathologies associated with chronic activation of the immune system. Such fundamental knowledge may contribute to understanding the processes associated with aging.
In addition to dissecting the immune system impact on various physiological systems
we aim to understand how the microbiota impact on the immuno-metabolic phenotype
in PD-1-deficient mice and the contribution of gut bacteria to the efficacy of PD1 antibody blockade.
In our quest to decipher the basic principles of systems immunology we employ
approaches that combine genetic and immunologic methods with metagenomic, mass
spectrometry-based proteomic and metabolomic methodologies together with extensive
phenotypic and functional in vivo studies.

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Gut Homeostasis
Team Leader: Kenya Honda

Figure: Identification and isolation of a colonic
TH1cell-inducing K. pneumoniae strain derived
from human saliva microbiota
FACS analysis of colonic LP T cells from exGF mice inoculated with saliva samples from patients with Crohn’sdisease (CD).

Recent Major Publications
Honda K, Littman DR. The microbiota in adaptive immune homeostasis and disease. Nature 535, 75–84(2016)
Saito T, Nishikawa H, Wada H, Nagano Y, Sugiyama D,
Atarashi K, Maeda Y, Hamaguchi M, Ohkura N, Sato
E, Nagase H, Nishimura J, Yamamoto H, Takiguchi S,
Tanoue T, Suda W, Morita H, Hattori M, Honda K, Mori
M, Doki Y, Sakaguchi S. Two FOXP3+CD4+ T cell subpopulations distinctly control the prognosis of colorectal
cancers. Nat Med 22, 679–684 (2016)
Tanoue T, Atarashi K, Honda K. Development andmaintenance of intestinal regulatory T cells. Nat Rev
Immunol 16, 295–309 (2016)

Invited Presentations
Honda K. “Regulation of T Cells by the Gut Microbiota”
Gordon Research Conference (GRC) in Immunochemistry and Immunobiology (Lucca (Barga), Italy) June,
2016
Honda K. “T cell stimulatory bacteria colonizing in thehuman oral cavity” CSH ASIA: Frontiers of Immunology
in Health & Disease (Awaji, Japan) October, 2016
Honda K. “Gut microbiota in immune activation” 4th
Annual meeting of the International Cytokine and Interferon Society (San Francisco, USA) October, 201622

T

he Laboratory for Gut Homeostasis has been working on the identification of specific members of the gut microbiota that have a deep impact on the immune system.
We have succeeded in isolation of 17 human gut-associated commensal bacterial strains
belonging to the class Clostridia that can potently trigger accumulation of colonic CD4+
Foxp3+ regulatory T (Treg) cells. The induced Treg cells expressed RORγt and IL-10 andhad strong suppressive activity for effector T cells. Introducing the 17 Clostridia strains
reduced GVHD severity through production of short chain fatty acids.
We have also focused on Th17 cells, which are known to accumulate in response
to colonization with a subgroup of intestinal microbes such as segmented filamentous
bacteria (SFB). Our previous study showed that SFB bound to small intestinal epithelial
cells (ECs) and induced Th17 cells strictly in a host-specific manner. Further analyses ofTh17 cell induction by the intestinal pathogens Citrobacter rodentium, Escherichia coli
O157:H7 and their adhesion-defective mutants revealed a strong correlation between epithelial adhesion and Th17 induction. Moreover, 20 bacterial strains isolated from feces ofan Ulcerative Colitis patient were able to induce Th17 cells in the colonic lamina propria
of germ-free mice, and these strains also exhibited EC-adhesive characteristics. These
results support our hypothesis that the physical interaction of the microbiota with thegut epithelium is essential for Th17 cell differentiation. Currently, we are in the process of
elucidating the nature of microbiota-inducing Th17 cells from healthy human donors bymetagenomic 16S rRNA sequencing.
Intestinal colonization of bacteria of oral origin correlates with several negative
health outcomes, such as inflammatory bowel diseases (IBD). In our recent project, wehave identified a strong intestinal Th1 cell-inducing Klebsiella pneumoniae strain froma saliva sample of an IBD patient (Figure). The K. pneumoniae strain produces innate
immune ligands, possibly packaged in outer membrane vesicles, to induce Th1 cells. Our
work suggests that there may be a sub-type of IBD in which bacteria of oral origin contribute to the pathogenesis.

Laboratory for

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Immune Homeostasis
Team Leader: Shohei Hori

Figure: Differential impacts of null versus“edgetic” Foxp3 alleles on Treg cells and self-tolerance
A Foxp3null allele globally compromises Treg cell physiology by removing not only Foxp3 itself but also all
the interactions between Foxp3 and its target genes aswell as partner proteins, and thereby leads to severe
systemic autoimmunity. On the other hand, an “edgetic” Foxp3 allele, which perturbs only certain molecularinteractions with its specific target genes or partner
proteins, could impair only specific facets of Treg cell
physiology and thereby result in more focused or deviated autoimmunity.

Recent Major Publications
Zhang B, Chikuma S, Hori S, Fagarasan S, Honjo T.
Nonoverlapping roles of PD-1 and FoxP3 in maintaining
immune tolerance in a novel autoimmune pancreatitis
mouse model. Proc Natl Acad Sci U S A 113, 84908495 (2016)
Ono T, Okamoto K, Nakashima T, Nitta T, Hori S, Iwakura Y, Takayanagi H. IL-17-producing gammadelta T cells
enhance bone regeneration. Nat Commun 7, 10928(2016)
Nishio J, Baba M, Atarashi K, Tanoue T, Negishi H, Yanai
H, Habu S, Hori S, Honda K, Taniguchi T. Requirementof full TCR repertoire for regulatory T cells to maintainintestinal homeostasis. Proc Natl Acad Sci U S A 112,
12770-12775 (2015)

Invited Presentations
Hori S. “Functional dissection of the Foxp3-centeredmolecular network in regulatory T cells” The 45th Annual Meeting of the Japanese Society for Immunology(Okinawa, Japan) December, 2016
Hori S. “Dissecting Foxp3-dependent mechanisms ofregulatory T cell function” The 39th Annual Meetingof the Molecular Biology Society of Japan (Yokohama,Japan) December, 2016

Regulatory T (Treg) cells expressing the transcription factor Foxp3 play an indispensable role in the establishment and maintenance of immunological self-tolerance
and tissue homeostasis, a concept well illustrated by the discovery that defective generation or function of Treg cells underlies the development of a fatal autoimmune diseasein Foxp3-mutant mice and in humans suffering from the IPEX syndrome. Findings that
Foxp3+ Treg cells exert tissue-protective or immune-suppressive functions under diverse
circumstances have raised the question of what mechanisms ensure the robustness ofTreg cell functions, and thus of immunological self-tolerance, in the face of unpredictable
perturbations in the extracellular environment. To answer this question, we have focusedon the mechanisms that control stability and adaptability of Treg cells in changing environments.
We have previously shown that, although Treg cells exhibit phenotypic plasticity,
they retain epigenetic memory of, and thus remain committed to, Foxp3 expression andregulatory functions. We are now addressing the mechanisms underlying this epigenetic
memory of Treg cell phenotype and function.
Another focus of our research is to understand how Foxp3 and Treg cells control immunological self-tolerance and tissue homeostasis in changing environments. To address
this question, we have addressed how Foxp3 mutations found in human IPEX impingeon Treg cells in vivo using knock-in mutagenesis in mice. Our analysis revealed that,while many mutations are loss-of-function mutations, one particular mutation acts as
an “edgetic” perturbation that specifically alters Foxp3 interactions with specific target
genes by broadening its DNA-binding specificity. Furthermore, this mutation preferentially impairs the ability of Treg cells to adapt to certain non-lymphoid tissue environments and thereby leads to tissue-restricted autoimmunity. By taking advantage of thisunique edgetic Foxp3 allele, we are currently investigating how Treg cells adapt to diverseand fluctuating tissue environments.

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Skin Homeostasis
Team Leader: Masayuki Amagai

Figure: Comprehensive analysis of skin barrier
homeostasis
Our team is trying to clarify the mechanisms of skinbarrier homeostasis by focusing on stratum corneum(SC), tight junction (TJ), and SG1 cells.

Recent Major Publications
Amagai M. Modulating immunity to treat autoimmune
disease. New Eng J Med 375, 1487–1489, 2016
Yokouchi M, Atsugi T, Logtestijn MV, Tanaka RJ, Kajimura M, Suematsu M, Furuse M, Amagai M, Kubo A.
Epidermal cell turnover across tight junctions based on
Kelvin's tetrakaidecahedron cell shape. eLife 5, e19593(2016)
Kobayashi T, Glatz M, Horiuchi K, Kawasaki H, Akiyama
H, Kaplan DH, Kong HH, Amagai M, Nagao K: Dysbiosis
and Staphyloccus aureus Colonization Drives Inflammation in Atopic Dermatitis. Immunity 42, 756–766(2015)

Invited Presentations
Amagai M. “Atopic diseases as a result of skin barrier
failure” 41st Annual Meeting of the Japanese Society
for Investigative Dermatology (Sendai, Japan) December, 2016
Amagai M. “The skin as a barrier in a dirty world”
136th Annual Meeting of American Dermatological
Association (Santa Barbara, USA) October, 2016
Amagai M. “Skin barrier homeostasis and its failure in
atopic dermatitis” Plenary Lectures at the 25th Congress of European Academy of Dermatology and Venereology (Vienna, Austria) September–October, 2016
Amagai M. “Skin barrier function and dysfunction in atopic diseases” 4th Taiwan Dermatology Aesthetic Conference/2016 Taiwanese Dermatological Association
Spring Meeting (Kaohsiung, Taiwan) April–May, 2016
Amagai M. “Skin barrier homeostasis by stratum
corneum and tight junction” Keystone Symposia Conference: Immunity in Skin Development, Homeostasis andDisease (Tahoe, USA) February–March, 201624

S

kin is the site where immunity meets external antigens. Cutaneous sensitization is now thought to be the initial key step for many allergic disorders, notonly atopic dermatitis (AD), but also asthma, food allergy and anaphylaxis. Skin
harbors several barriers to prevent easy penetration of external antigens into thebody; however, the exact molecular mechanisms by which the skin barriers form
and are maintained are largely unknown. The epidermis is keratinized stratified
squamous epithelia and is the outermost component of the skin. From bottom totop, the epidermis is composed of the stratum basale, stratum spinosum, stratum
granulosum (SG) and stratum corneum (SC). Our group has been focusing on the
SC barrier as an air-liquid barrier, and on tight junctions (TJ) as a liquid-liquidbarrier formed between SG2 cells, among many other skin barriers. There is afundamental biophysical paradox regarding the function of the epidermis, namely, how it can maintain the barrier but still constantly replace and shed cells. Ourgroup is trying to clarify how epidermal barrier homeostasis in maintained under normal conditions and how impaired barrier function occurs and affects microenvironments of the skin in various disease conditions. We use comprehensive
approaches combining molecular biology, biochemistry, ultrastructural anatomy,
live cell imaging, microbiology, and systems biology. For example, we have recently succeeded in isolating and characterizing SG1 cells and also in visualizing
SC-pH in living mice, which enables us to perform several unique experiments tounderstand SC homeostasis. Another of our strengths is to be able to go back andforth between the findings in basic science in mice and those in clinical science inhumans with various skin diseases. Our goal is to understand skin barrier homeostasis in health and diseases, and to provide patients suffering from severe allergic
diseases with more targeted, ideal therapeutic approaches with fewer side effects.

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Metabolic Homeostasis
Team Leader: Naoto Kubota

Figure: The mechanism of ‘selective insulin
resistance’ in type 2 diabetes with obesity
In type 2 diabetes with obesity, hepatocyte IRS-1
expression levels remain unaffected by thehyperinsulinemia, but the expression of IRS-2 isdownregulated in both the periportal (PP) andperivenous (PV) zones. Thus, in the PP zone, where IRS-1
is less abundantly expressed and IRS-2 expression is
downregulated, insulin signaling is impaired despite thehyperinsulinemia, leading to the impaired suppressionof gluconeogenesis and hyperglycaemia. By contrast, in
the PV zone, where IRS-1 is abundantly expressed,
insulin signaling is rather stimulated in the presence ofhyperinsulinemia despite the downregulation of IRS-2,
resulting in increased lipogenesis and development ofsteatosis.

Recent Major Publications
Sato H, Kubota N, Kubota T, Takamoto I, Iwayama
K, Tokuyama K, Moroi M, Sugi K, Nakaya K, Goto
M, Jomori T, Kadowaki T. Anagliptin increases insulin-induced skeletal muscle glucose uptake via anNO-dependent mechanism in mice. Diabetologia 59,
2426–2434 (2016)
Kubota N, Kubota T, Kajiwara E, Iwamura T, Kumagai H,
Watanabe T, Inoue M, Takamoto I, Sasako T, Kumagai
K, Kohjima M, Nakamuta M, Moroi M, Sugi K, Noda
T, Terauchi Y, Ueki K, Kadowaki T. Differential hepatic
distribution of insulin receptor substrates causes selective insulin resistance in diabetes and obesity. Nat
Commun 7, 12977 (2016)
Kubota T, Kubota N, Sato H, Inoue M, Kumagai H, Iwamura T, Takamoto I, Kobayashi T, Moroi M, Terauchi Y,
Tobe K, Ueki K, Kadowaki T. Pioglitazone Ameliorates
Smooth Muscle Cell Proliferation in Cuff-Induced Neointimal Formation by Both Adiponectin-Dependent and-Independent Pathways. Sci Rep 6, 34707 (2016)

Invited Presentations
Kubota N. “Diabetes and Cardiovascular diseases” Diabetes Scientific Update Meeting in Tokyo 2016 (Tokyo,
Japan) October, 2016
Kubota N. “Diabetes, obesity and gut microbiota” BioJapan 2016 (Yokohama, Japan) October, 2016
Kubota N. “Molecular mechanism of ’selective insulin
resistance’ in the liver” The 10th Diabetes Leading-edge
Conference (Tsukuba, Japan) August, 2016
Kubota N. “Molecular mechanisms in the dysregulationof glucose and lipid homeostasis in the liver” Annual
Scientific Meeting of Insulin Resistance and Metabolic
Syndrome Study Group 2016 (Tokyo, Japan) June, 2016
Kubota N. “Elucidation of the molecular mechanismof ‘selective insulin resistance’ in the liver” The 59th
Annual Meeting of the Japan Diabetes Society (Kyoto,
Japan) May, 2016

I

n recent years, there has been a rapid increase in the incidence of type 2 diabetes inboth Western and Asian countries; however, the precise molecular mechanisms underlying the progression of type 2 diabetes remain poorly understand. The goal of ourteam is to identify molecular mechanisms of insulin secretion and insulin resistance.
1) Molecular mechanism of insulin secretion
Impaired insulin secretion leads to the development of type 2 diabetes. This impaired
insulin secretion is thought to be partially caused by genetic factors. Most of the common variant single-nucleotide polymorphisms (SNPs) identified by genome-wide association study (GWAS) have been reported to be associated with defective pancreatic islet
function. However, the functional role of genes identified by GWAS remains unclear. To
elucidate the physiology and pathophysiological role of such genes in vivo, we have generated genetically engineered mice (Diabetologia 2014). Specifically, we are investigating
common variants in potassium voltage-gated channel subfamily Q member 1 (KCNQ1)
and ubiquitin-conjugating enzyme E2 E2 (UBE2E2), which confer the largest effect on
the risk of type 2 diabetes in Asians.
2) Molecular mechanism of insulin resistanceObesity-induced insulin resistance plays a crucial role in the pathogenesis of lifestyle-related diseases, including metabolic syndrome, nonalcoholic fatty liver disease (NAFLD),
and type 2 diabetes. Insulin resistance is defined as a condition in which physiologicalinsulin signals are impaired for some reason, such as chronic inflammation. Once insulinbinds to the insulin receptor, insulin receptor substrate (IRS)-1 and IRS-2 are activatedand mediate intracellular insulin signaling. Until now, we have been studying the role of
IRS-1 and IRS-2, which show ubiquitous expression patterns (Diabetes 2000, Circulation 2003, J Clin Invest 2004, Cell Metab 2008, Cell Metab 2011, Cell 2012, Nat Commun
2016). We will clarify the underlying mechanisms of insulin resistance by studying therole of IRS-1 and IRS-2.
25

Laboratory for

Lab activities

Immune Crosstalk
Team Leader: Hilde Cheroutre

Figure: Simplified model of the interaction
between Themis and T cell receptor signaling
molecules
Themis is constitutively associated with Grb2 whichbinds both LAT and SHP1. This association requires
two tandem tyrosine residues as well as a proline-richsequence (PRS) in the C terminal region of Themis. Although Themis translocates to the nucleus in response
to TCR stimulation, some amount of Themis also exists
in the nucleus at steady state. The Themis mutant
knock-in mouse, in which Themis cannot reside in the
nucleus, has a deficiency in T cell development indicating an essential role for Themis in the nucleus.

B
Recent Major Publications
Larange A, Cheroutre H. Retinoic Acid and Retinoic Acid
Receptors as Pleiotropic Modulators of the Immune
System. Annu Rev Immunol 34, 369–394 (2016)
Satoh R, Kakugawa K, Yasuda T, Yoshida H, Sibilia M,
Katsura Y, Levi B, Abramson J, Koseki Y, Koseki H, vanEwijk W, Hollander GA, Kawamoto H. Requirement of
Stat3 Signaling in the Postnatal Development of Thymic
Medullary Epithelial Cells. PLoS Genet 12, e1005776(2016)
Krause P, Morris V, Greenbaum JA, Park Y, BjoerhedenU, Mikulski Z, Muffley T, Shui JW, Kim G, Cheroutre H,
Liu YC, Peters B, Kronenberg M, Murai M. IL-10-producing intestinal macrophages prevent excessive antibacterial innate immunity by limiting IL-23 synthesis. Nat
Commun 6, 7055 (2015)

Invited Presentations
Cheroutre H. KAI-IBS Joint International Conference(Seoul, Korea) November, 2016
Cheroutre H. “Food for Thought: Mucosal Immune Protection and Regulation Controlled by the Diet” Institutefor Medical Science, University of Tokyo (Tokyo, Japan)
September, 2016
Cheroutre H. Yale University School of Medicine (NewHaven, USA) September, 2016
Cheroutre H. Inaugural Chiba University- Universityof California, San Diego Symposium (San Diego, USA)
February, 2016
Cheroutre H. The 55th Midwinter Conference of Immunologists (Pacific Grove, USA) January, 201626

y phenotype screening of ENU-induced mutant mice, we newly identified Themis
as an essential gene for T cell development. Themis functions by interacting with
Grb2 and SHP1, thus reducing TCR signals and allowing positive selection of thymocytes that further mature to naïve T cells. We found that, although Themis deletion impaired conventional selection, it did not affect positive agonist selection of self-specific
T cells. By generating specific nuclear localization signal mutants we now find that thenuclear localization of Themis is critical and essential for its function. Therefore, in addition to its function in proximal TCR signaling, we speculate that Themis also plays arole in the nucleus to control gene expression. In order to elucidate its genomic function,
we established a x3 FLAG tagged Themis knock-in mouse by CRISPR/Cas9 gene editing in order to perform highly precise ChIP seq analysis. Since GWAS studies identified
Themis as associated with celiac disease and atopic dermatitis, and since Themis has been
linked with inflammatory bowel disease, and furthermore, since Themis is essential forthe pathogenesis of C. Malaria and for protection from P. Tuberculosis, it appears that
Themis plays central roles in regulating peripheral T cells. To examine the genomic roleof Themis in mature T cells in vivo, we generated mutant mice that contained one allele ofthe nuclear translocation mutation and one allele of floxed wild type (WT) Themis. These
mice were crossed onto various Cre-transgenic mice to delete the WT Themis allele inspecific cells or with specific kinetics. This allows us to study the genomic role of Themis
during thymic development and selection as well as in various disease settings. This approach will allow us to understand the role of Themis and other factors in controlling
and translating pre-TCR and TCR signal strength during thymic selection and fate decisions of immature and mature T cells. We are also using unique and novel loss-and
gain-of-function approaches, designed in collaboration with Dr. Taniuchi’s laboratory. In
another study we discovered cytoplasmic retinoic acid receptor alpha (RARα) as a newmember of the TCR signalosome and a ZAP70 interacting partner. We found that cytoplasmic RARα plays a critical non-nuclear role in the proximal TCR signaling complex
as well as in TCR-induced NOTCH activation, c-Myc expression and activation-induced
proliferation. Furthermore, we characterized cytoplasmic RARα at the molecular level
and identified it as encoded by an alternative splice form of the nuclear RARα1 and -2,
with a conserved C-terminus. Cytoplasmic RARα is expressed by mature T cells as wellas immature thymocytes, indicating that it might also serve non-nuclear roles during
thymic development and selection. We are now designing various in vitro and in vivo
strategies to fully characterize cytoplasmic RARα at the molecular and cellular level andunder physiological and disease settings.

Laboratory for

Lab activities

Inflammatory Regulation
Team Leader: Takashi Tanaka

Figure: MKRN2 is a novel ubiquitin E3 ligase forthe p65 subunit of NF-κB and negatively regulates inflammatory responses
MKRN2 and PDLIM2 are ubiquitin E3 ligases for the
p65 subunit of NF-κB, form heterodimers and cooperatively promote polyubiquitination and degradationof p65, thereby negatively regulating NF-κB-mediatedinflammatory responses.

Recent Major Publications
Shin C, Ito Y, Ichikawa S, Tokunaga M, Sakata-Sogawa
K, Tanaka T. MKRN2 is a novel ubiquitin E3 ligase forthe p65 subunit of NF-κB and negatively regulates inflammatory responses. Sci Rep 7, 46079 (2017)
Tanaka T. Clarification of the molecular mechanismsthat negatively regulate inflammatory responses. In:
Miyasaka M, Takatsu K. (eds.), Chronic Inflammation:
Mechanisms and Regulation, 1st edition, Tokyo,
Japan: Springer Japan KK, pp109–118 (2016)
Ono R, Kaisho T, Tanaka T. PDLIM1 inhibits NF-κB-mediated inflammatory signaling by sequestering the p65
subunit of NF-κB in the cytoplasm. Sci Rep 5, 18327(2015)

Invited Presentations
Tanaka T. “The roles of LIM protein family in the regulation of inflammatory responses” The 45th Annual Meeting of the Japanese Society for Immunology (Okinawa,
Japan) December, 2016
Tanaka T. “Negative regulation for inflammatory responses and its association with autoimmune diseases”
The 2nd RIKEN-IITU Joint Symposium (Yokohama,Japan) December, 2016T

he inflammatory response is an important host defense mechanism to sense
and eliminate invading microbial pathogens. However, these inflammatory
responses must be terminated at the appropriate time point, otherwise excessive
responses can damage normal tissue and may lead to autoimmune diseases. Our
research goal is to identify a series of key negative regulators of inflammation-related signal transduction pathways and clarify the complete picture of the molecularmechanisms for regulating inflammatory responses. We predict that dysfunctionof these negative regulators might be the cause of human autoimmune diseases.
We previously identified PDLIM2 (PDZ and LIM domain-containing protein-2), a nuclear protein that belongs to a large family of LIM proteins, as one of
the key factors negatively regulating inflammatory responses. PDLIM2 is a ubiquitin E3 ligase for the p65 subunit of NF-κB in dendritic cells, negatively regulating
NF-κB-mediated inflammation. Since both PDZ and LIM domains are generally
thought of as structures involved in protein-protein interaction, we assumed thatPDLIM2 controls inflammatory responses through its interaction with other intracellular molecules that are also important for regulating inflammatory signaling. We therefore sought to isolate PDLIM2-interacting proteins that are criticalfor suppressing NF-κB signaling. We recently identified MKRN2, a RING finger
domain-containing protein that belongs to the makorin ring finger gene family, asa novel p65 ubiquitin E3 ligase. We have demonstrated that MKRN2 binds to PDLIM2 and that the heterodimer then synergistically promotes polyubiquitinationand degradation of p65, thereby negatively regulating NF-κB-mediated inflammatory responses. Moreover, to elucidate the mechanisms by which initially helpful
inflammation can lead to pathogenic chronic inflammation, we also analyzed theregulation of in vivo inflammatory responses using PDLIM2-deficient mice. Thesestudies should help clarify the pathogenesis of human autoimmune and inflammatory diseases and may lead to the development of new therapeutic tools forthese diseases.
27

Laboratory for

Lab activities

Cytokine Regulation
Team Leader: Masato Kubo

Figure: TH1 cells control protective IgG2 antibody
responses
Influenza virus preferentially induces development of
two different type of IFN-γ producing effector T cells,
CXCR3+ CXCR5+ TFH and CXCR3+ TH1 cells. TH1 cells are
the dominant subset responsible for the IFN-γ-producing CD4 T cells and the production of IgG2 protective
antibodies. These TH1 cells secreted IL-21 and IFN-γ,
both of which were required for low affinity IgG2 antibody production.

Recent Major Publications
Domínguez-Hüttinger E, Christodoulides P, Miyauchi K,
Irvine AD, Okada-Hatakeyama M, Kubo M, Tanaka RJ.
Mathematical modeling of atopic dermatitis reveals“double-switch” mechanisms underlying 4 common
disease phenotypes. J Allergy Clin Immunol S0091–
6749(16)31433-6 (2016)
Miyauchi K, Sugimoto-Ishige A, Harada Y, Adachi Y,
Usami Y, Kaji T, Hasegawa H, Watanabe T, Hijikata A,
Fukuyama S, Maemura T, Ohara O, Kawaoka Y, Takahashi Y, Takemori T, Kubo M. Protective neutralizing influenza antibody response in the absence of T follicularhelper cells. Nat Immunol 17, 1447–1458 (2016)
Yasuda T, Fukada T, Nishida K, Nakayama M, Matsuda
M, Miura I, Dainichi T, Fukuda S, Kabashima K, Nakaoka S, Bin BH, Kubo M, Ohno H, Hasegawa T, Ohara
O, Koseki H, Wakana S, Yoshida H. Hyperactivation of
JAK1 tyrosine kinase induces stepwise, progressive pruritic dermatitis. J Clin Invest 126, 2064–2076 (2016)

Invited Presentations
Kubo M. “Cytokine signaling in skin homeostasis” NEXT
Lecture Meeting in Chiba (Chiba, Japan) October, 2016
Kubo M. “Role of T follicular helper (TFH) in humoral
immunity” The 13th International Workshop on Autoantibodies and Autoimmunity (IWAA2016) (Kyoto,
Japan) October, 2016
Kubo M. “T dependent antibody responses in influenza
virus vaccination” Seminar series of new wave of virology (Kyoto, Japan) September, 2016
Kubo M. “Cross talk between innate aquired immunity
in allergic responses” The 65th Annual Meeting of Japanese Society Allergology (Tokyo, Japan) August, 2016
Kubo M. “New therapeutic approach for atopic dermatitis (AD) using comprehensive analysis and system
biology” 2016 International Biomedical Interface Symposium (Taipei, Taiwan) March, 201628

T

cells play a central role in the effector and regulatory functions of the immunological surveillance system, and aberrations in these functions can lead tovarious immunological disorders. Cytokines are critical factors in the transmission of information from the receptor to the nucleus as well as in the establishmentof communication networks among cells. Our overall long-term goal is to understand how effector cytokines are controlled during cell lineage commitment.
We have focused on antibody responses in systemic vaccination against Influenza A virus (IAV). Follicular helper T (TFH) cells are critical for developmentof the germinal center (GC) environment. Although TFH cells are important inanti-viral humoral immunity, IgG2 antibodies predominate in the response tovaccination with inactivated IAV and were responsible for protective immunity tolethal challenge with pathogenic H5N1 and pandemic H1N1 IAVs, even in B or
T cell-specific Bcl6-deficient mice. IL-21 and IFN-γ secreted from TH1 cells were
essential for greater persistence and higher titers of IgG2 protective antibodies.
These results suggest that TH1 induction could be a promising strategy to induce
effective neutralizing antibodies against emerging influenza viruses.

Laboratory for

Lab activities

Innate Immune Systems
Team Leader: Kazuyo Moro

Figure: Group 2 innate lymphoid cells and thecytokine network
Group 2 innate lymphoid cells (ILC2) are known to be
activated by epithelial-related cytokines such as IL33, IL-25 and TSLP, which are produced by apoptotic
epithelial cells, tuft cells, and activated epithelial cells,
respectively. Production of type-2 cytokines, includinginterleukin IL-4, IL-5, IL-9, and IL-13 promote alternative activation of eosinophilia, goblet-cell hyperplasia
and smooth-muscle contractility that contribute toexpulsion of helminth parasites but also worsen allergic
inflammation. ILC2 are tissue-resident cells and arestrongly suppressed by IFNγ and IL-27 in a STAT1-dependent manner.

Recent Major Publications
Morita H, Moro K, Koyasu S. Innate lymphoid cells inallergic and nonallergic inflammation. J Allergy Clin
Immunol 138, 1253-1264 (2016)
Moro K, Kabata H, Tanabe M, Koga S, Takeno N, Mochizuki M, Fukunaga K, Asano K, Betsuyaku T, Koyasu S.
Interferon and IL-27 antagonize the function of group
2 innate lymphoid cells and type 2 innate immune
responses. Nat Immunol 17, 76-86. (2016)
Vasanthakumar A, Moro K, Xin A, Liao Y, Gloury R,
Kawamoto S, Fagarasan S, Mielke LA, Afshar-Sterle S,
Masters SL, Nakae S, Saito H, Wentworth JM, Li P, Liao
W, Leonard WJ, Smyth GK, Shi W, Nutt SL, Koyasu S,
Kallies A. The transcriptional regulators IRF4, BATF and
IL-33 orchestrate development and maintenance of adipose tissue-resident regulatory T cells. Nat Immunol
16, 276-285 (2015)

Invited Presentations
Moro K. “The role of type 2 innate lymphoid cells in
the immune systems” The 44th Annual Meeting of theJapan Society for Clinical Immunology (Tokyo, Japan)
August, 2016
Moro K. “Suppression mechanism of Group 2 innate
lymphoid cells” FASEB: IgE and Allergy, 50 Years & Onward (West Palm Beach, USA) July, 2016
Moro K. “Mechanism for ILC2-mediated eosinophilia inallergic inflammation” The 65th Annual Meeting of Japanese Society of Allergology (Tokyo, Japan) June, 2016
Moro K. “IFNs and IL-27 regulate type 2 innate lymphoid cells” The 89th Annual Meeting of Japanese Society for Bacteriology (Osaka, Japan) March, 2016
Moro K. “Group 2 Innate lymphoid cell, a novel targetof the treatment for allergic diseases” The 34th Annual
Meeting of the Japan society of Immunology & Allergology in Otolaryngology (Mie, Japan) February, 2016

R

ecent studies have revealed new types of lymphocytes functioning in innate
immune responses that are collectively called innate lymphoid cells (ILC).
Unlike T and B lymphocytes, ILCs lack Rag-dependent antigen-specific receptors
and are activated by cytokines produced by other innate immune cells or epithelial cells. ILCs have been divided into three groups based on their cytokine production profiles; group 1 ILC including NK cells and ILC1 produce IFNγ, group 2
ILC (ILC2) including natural helper cells, nuocytes and innate helper type 2 cells
produce type 2 cytokines such as IL-5, IL-6 and IL-13, and group 3 ILC including
lymphoid tissue inducer (LTi) cells and ILC3s produce IL-17 and IL-22.
Our research group focuses on group 2 innate lymphoid cells (ILC2), an innate
lymphocyte lineage that we identified in 2010. ILC2 localize to a variety of tissues
such as fat, lung, intestine, liver and skin, and mediate immunity to helminth andfungal infections via strong type 2 cytokine production. Infection with helminths
or fungi induces IL-25 and IL-33 production by epithelial cells or endothelial cells
and activates ILC2, leading to eosinophilia and goblet cell hyperplasia resulting
from the production of IL-5 and IL-13, respectively. These immune responses are
extremely important for defense against infection, but recent studies have shownthat ILC2 can also exacerbate allergic inflammation using the same mechanisms.
We think that it is particularly important to determine the ILC2 activation andsuppression mechanisms during infection and inflammation in order to develop effective treatments for ILC2-related allergic diseases. Recently, we found that
ILC2 are involved in obesity, which leads to a variety of metabolic syndromes. To
gain a comprehensive understanding of the role of ILC2, our lab focuses on theirdevelopment, cytokine signaling pathways, miRNA regulation as well as theirfunction in adipose tissue inflammation, autoimmune disease, helminth infection, and a number of allergic diseases.

29Core for
Precise Measuringand ModelingToward the ultimate goal of obtaining a comprehensive understandingof the pathogenesis of human diseases, the functions of the Core forPrecise Measuring and Modeling are three pronged: production of mousemodels, multiomics measurements and quantitative bioimaging, and bioinformatics/modeling of human disease processes. Through close interactions
among these three branches of the core, we aim to collect a wide varietyof quantitative data in order to build a computational and predictive network of the disease process. As for the production of genetically engineered
mice that will be used as models of human diseases, the laboratory for Developmental Genetics has begun to apply recent advances in genome engineering technology, e.g., CRISPR/Cas9-based genome editing, and thereby considerably enhance the production capacity and power of the diseasemodels. Regarding the precise quantitative measurements branch, we haverecently enhanced the power of metabolite analysis by the Laboratory for
Metabolomics. Together with mRNA/protein profiling by the Laboratory for
Integrative Genomics, the enhanced multiomics measurements and bio-imaging (Laboratory for Tissue Dynamics) will greatly contribute to explorationof the etiology of human diseases. After being processed by bioinformatics(Laboratory for Integrated Bioinformatics), the datasets are used for modeling (Laboratories for Disease Systems Modeling and Integrated Cellular
Systems). The laboratory for Medical Science Mathematics is working to fill
the gap between humans and mice. As a leading IMS project, an atopic
dermatitis model mouse, provided by the Laboratory for Immunogenetics,
has been extensively analyzed from several different angles, fully exploiting
the analysis powers of this core. These efforts should enable us to identify
new biomarkers for early diagnosis and prevention of atopic dermatitis in
the very near future.

30

Laboratory for

Lab activities

Developmental Genetics
Group Director: Haruhiko Koseki

Figure: EPC1/2-mediated epigenetic inheritancethat is coupled with the replication machinery
EPC1/2 are coupled with the replication machinery andare expected to contribute to chromatin replication bylinking replication and epigenetic mechanisms.

Recent Major Publications
Yamada D, Iyoda T, Vizcardo R, Shimizu K, Sato Y, Endo
TA, Kitahara G, Okoshi M, Kobayashi M, Sakurai M,
Ohara O, Taniguchi M, Koseki H, Fujii SI. Efficient regeneration of Human Vα24+ invariant NKT cells and theiranti-tumor activity in vivo. Stem Cells 34, 2852–2860(2016)
Sharif J, Endo TA, Nakayama M, Karimi MM, Shimada
M, Katsuyama K, Goyal P, Brind’Amour J, Sun M-A, Sun
Z, Ishikura T, Mizutani-Koseki Y, Ohara O, Shinkai Y,
Nakanishi M, Xie H, Lorincz MC, Koseki H. Activationof Endogenous Retroviruses in Dnmt1−/− ESCs Involves
Disruption of SETDB1-Mediated Repression by NP95
Binding to Hemimethylated DNA. Cell Stem Cell 19,
81–94 (2016)
Yakushiji-Kaminatsui N, Kondo T, Endo TA, Koseki Y,
Kondo K, Ohara O, Vidal M, Koseki H. RING1 proteins
contribute to early proximal-distal specification of theforelimb bud by restricting Meis2 expression. Development 143, 276–285 (2016)

Invited Presentations
Koseki, H. “The role of PRC1 variants during activationof Meis2” Seminar at Max-Planck Institute of Immunobiology and Epigenetics (Freiburg, Germany) November, 2016
Koseki, H. “Regeneration of NKT cell using iPS cell technology and its adoptive immunotherapy for cancer” The25th Annual Meeting of Japanese Society of Strategies
for Cancer Research and Therapy (Chiba, Japan) June,
2016
Koseki, H. “Polycomb in transcriptional phase transitionof developmental genes” Mini Symposium, Genetic
Control of Mouse embryonic Development at Hubrecht
Institute (Utrecht, the Netherlands) June, 2016
Koseki, H. “PCGF6-PRC1 to suppress premature activation of meiosis/germ cell related genes in ES cells”
Seminar at Trinity College, Oxford University (Oxford,
UK) June, 2016
Koseki, H. “Personalized Preventive Medicine: chronic
dermatitis” Workshop on Personalised Medicine,
Tübingen University Hospital, Center for Personalised
Medicine (Tübingen, Germany) May, 2016T

he Developmental Genetics Research Group is pursuing a research program toelucidate the molecular mechanisms underlying organ development and stem cellfunctions. Particular emphasis has been put on epigenetic regulation mediated by the
combinatorial actions of Polycomb group (PcG) gene products, DNA methylation mechanisms and cell cycle regulation.
EPC1/2 (Enhancer of Polycomb) are orphan Polycomb factors. We found that EPC1/2
are essential for proliferation of ES cells by generating conditional double knockout(Epc-dKO) ES cells. By using immunofluorescence staining and isolation of proteins on
nascent DNA (iPOND), we observed that EPC1/2 accumulate at nascent chromatin andmove along with the replication fork similar to PCNA, a processivity factor for DNA polymerase. Further, mass spectrometry analysis of the purified EPC1-Flag complex revealed
its association with the NuA4 complex, Ino80 chromatin remodeling complex, and cohesin complex, supporting its functions in DNA replication. Indeed, depletion of EPC
significantly impaired DNA replication, and cells became more sensitive to replicationstress. To dissect how EPC depletion influences replication, we monitored the binding of
replication factors to nascent chromatin. We observed a clear reduction in histone chaperones CAF-1 and ANP32e, FACT, Polδ, and PCNA in Epc-dKO cells. These results indicated that EPC plays a role as a platform to stabilize the replication fork.
EPC1/2 have been reported to bind to nucleosomes via their N-terminal regions,
therefore we presumed that EPC1/2 would contribute to nucleosome assembly at the replication fork. We indeed found that both parental and newly generated histones were lessefficiently incorporated in Epc-dKO. Consistently, the chromatin structure at promoter
regions was not sufficiently reassembled and loosened. Further, we found alternative
deposition of histone variants, an impaired incorporation of H2A.Z while H3.3 was increased, as well as defects in epigenetic features at pericentric heterochromatin (PHC),
reduced trimethylation levels at H3K9 and H3K27, loss of HP1 binding and DNA methylation. Based on these results together with other data, we suggest that EPC1/2 contributeto mediate epigenetic inheritance mechanisms during replication.
31

Laboratory for

Lab activities

Integrative Genomics
Group Director: Osamu Ohara

Figure: Development of A Real-Time Monitoring
Platform for Single-cell Omics
In a previous study, we used a single-hole chip formeasurement of protein secretion by single cells (SciRep, 2014, 4:4736). To improve the throughput of themeasurements, we developed a four-hole chip, termed“Quad Chip”, which enabled us to measure single-cellbehaviors under four different conditions simultaneously (Panel A). Together with improvements in the opticalsystem, the new platform equipped with a “Quad Chip”
greatly enhances measurement performance. In addition, we also improved a device for retrieval of single
cells at a time point of interest (Panel B). Consequently,
the new platform allows us to monitor the behavior
of intracellular and secreted proteins in real-time byfluorescent microscopy and obtain mRNA or epigenetic
profiles of single cells at their endpoint.

Recent Major Publications
Kagawa R, Fujiki R, Tsumura M, Sakata S, Nishimura S,
Itan Y, Kong XF, Kato Z, Ohnishi H, Hirata O, Saito S,
Ikeda M, El Baghdadi J, Bousfiha A, Fujiwara K, Oleatro
M, Yancoski J, Perez L, Danielian S, Ailal F, Takada H,
Hara T, Puel A, Boisson-Dupuis S, Bustamante J,
Casanova JL, Ohara O, Okada S, Kobayashi M. Alaninescanning mutagenesis of human signal transducer
and activator of transcription 1 to estimate loss- or
gain-of-function variants. J Allergy Clin Immunol pii:
S0091–6749, 31281–31287 (2016)
Miyauchi K, Sugimoto-Ishige A, Harada Y, Adachi Y,
Usami Y, Kaji T, Inoue K, Hasegawa H, Watanabe T, Hijikata A, Fukuyama S, Maemura T, Okada-Hatakeyama
M, Ohara O, Kawaoka Y, Takahashi Y, Takemori T, Kubo
M. Protective neutralizing influenza antibody responsein the absence of T follicular helper cells. Nat Immunol
17, 1447–1458 (2016)
Yasuda T, Fukada T, Nishida K, Nakayama M, Matsuda
M, Miura I, Dainichi T, Fukuda S, Kabashima K, Nakaoka S, Bin BH, Kubo M, Ohno H, Hasegawa T, Ohara
O, Koseki H, Wakana S, Yoshida H. Hyperactivation of
JAK1 tyrosine kinase induces stepwise, progressive pruritic dermatitis. J Clin Invest 126, 2064–2076 (2016)

Invited Presentations
Ohara O. “Snapshot vs Real-Time Monitoring: Toward
an Understanding of Population Dynamics of Immune
Cells at the Single-Cell Resolution” The 6th International Multidisciplinary Conference on Optofluidics (Beijing,
China) July, 2016
Ohara O. “From genotype to phenotype: Mind the gap
in multiscale structure of the phenotype” 5th Sardinian
Summer School: From genome-wide association studies(GWAS) to function (Pula, Italy) June, 201632

W

hile the original mission of the Laboratory for Integrative Genomics atthe beginning of the center was to function as a “Gateway” to genomics,
genomics has become the approach of choice to tackle complex problems in medical sciences. Thus, we are currently involved in many intramural and extramural collaborations and various strategic projects organized by the center, besides
providing our colleagues in the center with technical support as a central support
function. However, it is not our intent to be overwhelmed by these activities: Everyeffort has also been made to continuously update new technologies with the least
delay, but only after they become mature and robust enough to be useful in a general laboratory setting. This is still the basic mission of our laboratory.
Technology development has had a big impact on the design of biological experiments and greatly deepens our understanding of biological systems. As wehave experienced it first hand by the recent emergence of next-generation DNA
sequencing technology, new technology drastically changes the experimental approaches in biology; even a data-driven approach can now be personalized. In
this context, the most critical issue for our laboratory is to correctly decide on the
direction of technology development. As one layer in the multi-scale structure ofa biological system, our current focus is on the development of single-cell measurement technologies and their applications. In particular, we are most interested
in the temporal dynamics of cellular responses at single-cell resolution. As a fruitof studies in this direction, through collaboration with Dr. Uemura’s Laboratory
at the University of Tokyo (Department of Biological Sciences, Graduate Schoolof Science), we have recently developed a single-cell omics platform shown in the
Figure. Together with snapshot omics data, either in bulk or at single-cell resolution, the datasets of temporal dynamics of cellular responses obtained by thisplatform greatly contribute to a comprehensive understanding of the systems behaviors of a cellular ensemble of interest.

Laboratory for

Lab activities

Disease Systems
Modeling
Group Director: Hiroaki Kitano

Figure: Comprehensive maps of the stress response pathways in budding yeast
This is the most comprehensive signaling map of anorganism ever produced. Pathways were categorized
into six groups and contain multiple layers of information, which are not included in simple interaction
networks such as protein-protein interaction (PPI). Network structure analysis revealed a novel mechanism toensure robustness of signaling networks.

Recent Major Publications
Kawakami E, Nakaoka S, Tazro Ohta T, Kitano H. Kawakami E, Nakaoka S, Tazro Ohta T, Kitano H. Weighted
enrichment method for prediction of transcription
regulators from transcriptome and global chromatin
immunoprecipitation data. Nucleic Acids Res 44,
5010–5021 (2016)
Kitano H. Artificial intelligence to win the Nobel Prizeand beyond: Creating the engine of scientific discovery.
AI Magazine 37, 39–49 (2016)
Kawakami E, Singh VK, Matsubara K, Ishii T, Matsuoka
Y, Hase T, Kulkarni P, Siddiqui K, Kodilkar J, Danve N,
Subramanian I, Katoh M, Shimizu-Yoshida Y, Ghosh
S, Jere A, Kitano H. Network analyses based on comprehensive molecular interaction maps reveal robust
control structures in yeast stress response pathways.
NPJ Syst Biol Appl 2, 15018 (2016)

Invited Presentations
Kitano H. “Nobel Turing Challenge” The 39th Annual
Meeting of the Molecular Biology Society of Japan (Yokohama, Japan) December, 2016
Kitano H. “AI-Driven biomedical research – Grand Challenge of AI and Systems Biology” Precision Medicine in
Metabolic Disease: A new data driven approach to prevent the onset of metabolic disease (Sydney, Australia)
October, 2016T

he Laboratory for Disease Systems Modeling (LDSM) aims at in-depth understanding of several biological processes relevant to disease systems, andpossible applications to clinical practice. There are three interrelated projects: (1)Skin Homeostasis, (2) Aging, and (3) Precision systems biology using budding
yeast. Skin homeostasis is linked with the IMS center project on atopic dermatitis.
We are striving to create computational models combined with a rage of cell-based
experiments at LDSM together with mouse data from other IMS teams. Aging is
a new topic with the long-range objectives to understand possible control of theaging process and uncovering molecular mechanisms behind it. We are focusing
on the effect of SIRT1 (mammalian) and Sir2 (budding yeast), and nicotinamidemononucleotide as a chemical substance to affect these genes. We are linking skinhomeostasis and aging to low-grade chronic inflammation as the common thread.
Precision systems biology using budding yeast strengthens our understanding of
key biological process by providing deeper insights into detailed molecular processes with our original gTOW genome-wide assay system.
In addition, the LDSM has extensive involvement with the Medical Innovation
Hub project in its feasibility study stage, but that will not be reported here.

Kitano H. “Healthy Aging in the context of Systems
Biology, Digital Health, and Artificial Intelligence” JST(/
AMED)-Leibniz Association Joint Workshop on Healthy
Ageing (Tokyo, Japan) June, 2016
Kitano H. “Artificial Intelligence for Scientific Discovery
in Biomedical Sciences” GET Conference (Boston, USA)
April, 2016
Kitano H. “Cellular systems” The 3rd EMBO Conference
on Visualizing Biological Data (VIZBI 2016) (Heidelberg,
Germany) March, 201633

Laboratory for

Lab activities

Medical Science
Mathematics
Group Director: Tatsuhiko Tsunoda

Recent Major Publications
Fujimoto A+, Furuta M+, Totoki Y+, Tsunoda T+, Kato
M+ (+: co-first), Shiraishi Y, Tanaka H, Taniguchi H,
Kawakami Y, Ueno M, Gotoh K, Ariizumi S, Wardell CP,
Hayami S, Nakamura T, Aikata H, Arihiro K, Boroevich
KA, Abe T, Nakano K, Maejima K, Sasaki-Oku A, Ohsawa A, Shibuya T, Nakamura H, Hama N, Hosoda F, Arai
Y, Ohashi S, Urushidate T, Nagae G, Yamamoto S, Ueda
H, Tatsuno K, Ojima H, Hiraoka N, Okusaka T, Kubo M,
Marubashi S, Yamada T, Hirano S, Yamamoto M, Ohdan
H, Shimada K, Ishikawa O, Yamaue H, Chayama K,
Miyano S, Aburatani H, Shibata T, Nakagawa H. Whole
genome mutational landscape and characterization of
non-coding and structural mutations in liver cancer. Nat
Genetics 48, 500–509 (2016)
Sharma A, Boroevich KA, Shigemizu D, Kamatani Y,
Kubo M, Tsunoda T. Hierarchical Maximum Likelihood
Clustering Approach. IEEE Trans Biomed Eng 64,
112–122 (2016).
Fujimoto A, Furuta M, Shiraishi Y, Nguyen HH, Shigemizu D, Gotoh K, Kawakami Y, Nakamura T, Ueno M,
Ariizumi S, Shibata T, Abe T, Boroevich KA, Nakano K,
Sasaki A, Kitada R, Maejima K, Yamamoto Y, Tanaka H,
Shibuya T, Ojima H, Shimada K, Hayami S, Shigekawa
Y, Aikata H, Arihiro K, Ohdan H, Marubashi S, Yamada
T, Ishikawa O, Kubo M, Hirano S, Yamamoto M, Yamaue
H, Chayama K, Miyano S, Tsunoda T*, Nakagawa H*(*: co-last). Whole-genome mutational landscape ofliver cancers displaying biliary phenotype reveals hepatitis impact and molecular diversity. Nat Commun 6,
6120 (2015)

Invited Presentations
Lysenko A and Kamola P. “Strategies for discovering
disease-associated modules in integrated biologicalnetworks” RECOMB/ISCB Conference on Regulatory and
Systems Genomics (Phoenix, USA) November, 2016
Shigemizu D. “Current status of exome analysis for
identifying disease gene mutation” Grant-in-Aid for
Scientific Research A Symposium (I) Large scale data
analysis and modeling to understand complicated biological phenomena (Kurume, Japan) November, 2016
Tsunoda T. “Medical big data analysis drives precisionand preemptive medicine” Symposium ‘Basics andMedical Application of Statistical Genetics’, 23rd Annual Meeting of the Japanese Society for Gene Diagnosis
and Therapy (Tokyo, Japan) October, 2016
Tsunoda T. “Disease multi-omics analysis utilizing medical big data” CREST Symposium ‘Elucidation of the life
system by trans-omics’ (Tokyo, Japan) March, 2016
Tsunoda T. “Big data analysis frontiers personalized
medicine” Software Japan 2016 (Tokyo, Japan) February, 201634

Figure: Medical Big Data Analysis for Precision Medicine.
(a) Common analysis steps and methodologies. (b) Application to omics data from 300
hepatocellular carcinoma cases revealed six clusters, which showed significantly different disease-free survival rates (Nat Genet 48, 500–509 (2016)).

T

he application of rapidly progressing omic profiling technologies and, inparticular, the promotion of personalized/precision/preventive medicine
have recently become major goals of medical research. Traditional therapies do
not adequately take into account the individuality of each patient. Our laboratory
develops strategies to overcome such medical science issues through a combination of mathematics and computational sciences. Nowadays, biomedical big data,consisting of both clinical and omic profiles, are collected from hospitals and medical institutions. First, driven by need for integrative analysis of clinical and omic
data, we explore etiologies of intractable diseases, e.g., cancer, common diseases,
and neurodegenerative diseases. Next, we classify each disease into finer categories through molecular profiles, and clarify disease causing mechanisms througha systems approach. Last, we apply mathematical methods, e.g., machine learningtechniques, to optimize therapy prediction for each patient when she/he visits ahospital or medical institute. We can also apply these methods to disease prevention based on an individual’s medical history. Our laboratory does biomedical
research and genomic medicine research driven by prediction through complete
utilization of advanced mathematics and computational sciences with the goal ofdeveloping personalized/precision/preventive medicine strategies and methodologies. Recently, we developed statistical and computational methodologies foranalyzing up-to-date omics data, as well as for understanding and predicting cancer progression based on genomic data, with a final aim of establishing precision
medicine and applying it to real data [Nat Genet 48, 500–509 (2016); IEEE Trans
Biomed Eng 64, 112–122 (2016); J Theor Biol 393, 67–74 (2016); BMC Bioinformatics 17, 319 (2016); BMC Med Genomics 9(Suppl 3), 74, 2016; IEEE Trans Nanobioscience 14, 915–926 (2015)]. On the basis of these methodologies, we succeeded inobserving a significant correlation between the molecular clustering of omic dataand clinical information, e.g. survival time, from liver cancer omic analysis, andhave recently published these results (Nat Genet 48, 500–509 (2016)).

Laboratory for

Lab activities

Immunogenetics
Team Leader: Tadashi Yamamoto

Figures: Research projects in the laboratory
A. Defects in the mRNA decay system result inabnormal tissue development
The CNOT complex-mediated mRNA decay pathway
contributes to tissue development by regulating gene
expression in the transition from undifferentiated todifferentiated cells. When mRNA decay is deregulated
by inhibition of the CNOT complex or by an ectopic
increase in mRNA stabilizing molecules (green), abnormal tissues and various diseases develop. Photos:
necrosis in liver (upper) and reduction of adipocyte size
and macrophage infiltration in white adipose tissue(lower) in mice lacking the CNOT3 component of the
CNOT complex.
B. Activation dynamics lead to specific gene
expression
Signaling networks generate the distinct activation
dynamics induced by different stimuli. The differencesin activation dynamics may determine specific gene
expression profiles and, hence, biological output.
C. The kinetics of Ag-specific CD8 T cell numbers
following infection

Recent Major Publications
Li X, Morita M, Kikuguchi C, Takahashi A, Suzuki T*
and Yamamoto T*. (*corresponding author) Adipocyte-specific disruption of mouse Cnot3 causes lipodystrophy. FEBS Lett 591, 358–368 (2017).
Shinohara H*, Inoue K, Yumoto N, Nagashima T,
Okada-Hatakeyama M. (*corresponding author) Stimulus-Dependent Inhibitor of Apoptosis Protein Expression Prolongs the Duration of B Cell Signaling. Sci Rep
6, 27706 (2016)
Setoguchi R. IL-15 boosts the function and migrationof human terminally differentiated CD8+ T cells by
inducing a unique gene signature. Int Immunol 28,
293–305 (2016)

Invited Presentations
Yamamoto T. “The CCR4-NOT dedenylase: its roles incontrolling cells’ survival and differentiation in various
systems” The 39th Annual Meeting of the Molecular
Biology Society of Japan (Yokohama, Japan) December,
2016
Yamamoto T. “The CCR4-NOT deadenylase: a key playerof novel paradigm of gene regulation” The 89th Annual
Meeting of the Japanese Biochemical Society (Sendai,
Japan) September, 2016T

hree projects are ongoing in our team to understand the molecular basis of tissue
development and the dynamics of signal transduction.
1) Tissue development involves dramatic changes in gene expression. Control of mRNAstability as well as transcriptional regulation contribute to these gene expression changes.
We have found that suppression of the deadenylase activity of the CNOT complex, whichmediates mRNA degradation, leads to abnormal development of liver and adipose tissues
concomitant with cell death and inflammation. Augmentation of mRNA binding proteins that stabilize target mRNAs are also observed in various diseases. We are investigating how those molecules regulate tissue development and function in a developmental
stage-specific manner, and how they are deregulated in diseases.
2) By analyzing protein modification that controls signal dynamics as a drug target, it ispossible to develop therapeutic agents without side effects. Therefore, the importance ofunderstanding the dynamics of signal transduction has become increasingly clear. In B
cells, cell differentiation is determined by signals from the antigen receptor and cooperative receptors such as CD40. The mechanisms that produce different cellular responses
despite the qualitatively identical molecules activated by various stimuli will be mediatedby changes in active kinetics controlled by temporal information. We have quantitatively
analyzed the induced protein complexes and dynamics of signaling activity and cellularresponses by time-controlled ubiquitin modification. This has helped to clarify the signaling networks and control mechanisms of induced cell differentiation.
3) Memory CD8 T cells are long-lived antigen (Ag)-specific cells that can respond quickly, proliferate robustly, and exert effector functions faster than naïve CD8 T cells uponreencounter with their cognate Ags. Memory CD8 T cells are maintained as a stable population over a long period, but the mechanisms by which their population size is maintained remain elusive. One of objectives of our study is to reveal how memory CD8 T cells
are maintained in vivo. This research can provide new vaccination strategies for induction of protective immunity against tumor cells and in chronic infections.

35

Laboratory for

Lab activities

Integrated Bioinformatics
Team Leader: Todd D. Taylor

Figure: iCLiKVAL Overview
iCLiKVAL is a web-based tool that uses the power ofcrowdsourcing to accumulate annotation information
for all scientific media found online. Annotations in
the form of key-relationship-value tuples are added byusers through a variety of methods. Users can createor join common interest groups to work as part of acommunity. Controlled vocabulary lists can be created, edited and shared. Media can be bookmarked,
followed, reviewed and searched. Annotations can besorted, filtered and edited. The database is completely
searchable, without registration, and all of the data arefreely available to registered users via our application
programming interface.

Recent Major Publications
Lau N-S, Makita Y, Kawashima M, Taylor TD, Kondo S,
Othman AS, Shu-Chien AC, Matsui M. The rubber tree
genome shows expansion of gene family associatedwith rubber biosynthesis. Sci Rep 6, 28594 (2016)
Hebrard M, Taylor TD. MetaTreeMap: an alternativevisualization method for displaying metagenomic phylogenic trees. PLOS ONE 11, e0158261 (2016)
Jinda W, Poungvarin N, Taylor TD, Suzuki Y, Thongnoppakhun W, Limwongse C, Lertrit P, Suriyaphol P,
Atchaneeyasakul L. A novel start codon mutation of
the MERTK gene in a patient with retinitis pigmentosa.
Mol Vis 22, 342-351 (2016)

Invited Presentations
Taylor TD. “Improving the connectivity, searchability
and discoverability of scientific media through annotations with iCLiKVAL” Faculty of Medicine Siriraj Hospital,
Mahidol University (Bangkok, Thailand) June, 2016
Taylor TD. “Turning ‘big data’ into ‘small data’ throughcrowdsourced curation: integrating all types of medical
and scientific knowledge” 3rd AIST-RIKEN Joint Bioinformatics Seminar, RIKEN (Yokohama, Japan) June, 201636

W

e are developing an integrated database and sample-tracking system tohandle both small- and large-scale datasets of various types of experimental outputs. The system brings together an array of wet-lab experimental types being generated by various labs. Our goal is to develop a flexible system that makes
it easy for users to manage, access, analyze, integrate, and visualize their own data
as per their requirements.
We also develop general-purpose bioinformatic tools capable of efficiently processing and analyzing data from a variety of sources, with an emphasis on metagenomic data (e.g., taxonomic classification, phylogenic tree visualization, 16S rRNA
curation), host-microbiome interactions, and scientific discovery through big data
curation. We have constructed a highly-curated genomic-based 16S ribosomal
RNA gene database, which is continually being updated as new sequences appear
in the public databases. Because metagenomic samples can contain hundreds or
thousands of different species and are not easy to visualize or quantify, we developed an alternative visualization method for displaying phylogenic trees thatallows users to, at a glance, comprehend the distribution of the species within theirsamples. Big data in the form of scientific media comes in many languages andformats: journal articles, books, images, videos, etc. While there are many resources for browsing, searching and annotating some of this media, there is no single
place to search them all at once, and generalized search engines do not allow for
the comprehensive and precise searches researchers require. To address these issues, we have developed a web-based tool that uses the power of crowdsourcing toaccumulate annotation information for all scientific media found online. This willallow for richer data searches and discovery of novel connections by integrating all
forms of scientific knowledge through common terminology.

Laboratory for

Lab activities

Tissue Dynamics
Team Leader: Takaharu Okada

Figure: Multicolor imaging of dendritic cells andT cells in the mouse model of contact dermatitis
by intravital multiphoton microscopy. (Modified
from Okada et al. Pflugers Arch 468, 1793–1801,
2016).
Chicken ovalbumin-reactive T cell receptor transgenic
CD4+ T cells expressing tdTomato plus EGFP (pink topurple) and CD8+ T cells expressing tdTomato (red)were transferred to an Xcr1gfp/+ CD11c-YFP mouse forvisualization of their interactions with XCR1+ dendritic
cells (light blue) and other dendritic cells (green). The
mouse was subcutaneously immunized in the flankwith ovalbumin plus poly (I:C). Four days after immunization, the mice were then intradermally injected in
the dorsum of the foot with ovalbumin alone. Sevendays later, the mouse was anesthetized, and the skinof the dorsum of the foot was imaged on an inverted
multiphoton microscope with four external detectors.
Excitation wavelength was 910 nm. (A) Projection
images of ten z-slices of the dermis (33–60-µm depth
from the skin surface) at the beginning and end of the
2-h recording. Scale bar: 80 µm. Collagen fibers (white)were also visualized by second harmonic generation.
(B) Projection images of four z-slices of the epidermis(12–21-µm depth from the skin surface) at the samex-y position as (A). The scattered epidermal dendritic
cells in green are mostly Langerhans cells. (C and D)
Time-lapse images of the region indicated by dottedlines in (A) and (B). Scale bar: 40 µm. Yellow lines in(C) are paths of dendritic cell migration tracked every
minute. Yellow arrowheads indicate starting positionsof the tracks.

Recent Major Publications
Okada T, Takahashi S, Ishida A, Ishigame H. In vivo multiphoton imaging of immune cell dynamics. Pflugers
Arch 468, 1793–1801 (2016)
Shinnakasu R, Inoue T, Kometani K, Moriyama S, Adachi
Y, Nakayama M, Takahashi Y, Fukuyama H, Okada T,
Kurosaki T. Regulated selection of germinal-center cells
into the memory B cell compartment. Nat Immunol
17, 861–869 (2016)
Kitano M, Yamazaki C, Takumi A, Ikeno T, Hemmi H,
Takahashi N, Shimizu K, Fraser SE, Hoshino K, Kaisho
T, Okada T. Imaging of the cross-presenting dendritic
cell subsets in the skin-draining lymph node. Proc Natl
Acad Sci U S A 113, 1044–1049 (2016)T

he goal of the laboratory is to mechanistically understand the in vivo cellular dynamics that shape immune responses and inflammation. To this end,
we conduct multi-dimensional fluorescent imaging analysis by two-photon microscopy. This microscopy method has been revealing the striking dynamics ofimmune cells in various organs, underlining the importance of this approach toresolve the complexity of the immune system. By applying the imaging strategy to
relevant mouse models, we aim to reveal immune cell dynamics that are criticalfor generation of immunological memory and tolerance. We have continued to
study various mouse strains, in which (1) dynamics of helper T cells specialized
for B cell immune responses are perturbed, (2) cross-presenting dendritic cells(DCs) are fluorescently labeled for in vivo imaging, and (3) dynamics and function of regulatory T (Treg) cell subsets are perturbed to break peripheral toleranceof autoreactive B cells and CD8+ T cells.
In addition, we are studying the roles for the peripheral nervous system inmodulation of inflammation. Particularly, we are interested in structural andfunctional changes in sensory neurons in dermatitis. In this study as well, the imaging approaches are very powerful for deciphering the diversity and dynamics ofcellular activities. Through this study, we aim to find strategies to suppress chronic
inflammation based on the modulation of neuronal activities.

Invited Presentations
Okada T. “Cellular dynamics shaping adaptive immune
responses in the lymph node” The 54th Annual Meetingof The Biophysical Society of Japan (Tsukuba, Japan)
November, 2016
Okada T. “Imaging of cellular dynamics shaping theadaptive immune system” The 68th Annual Meetingof The Japan Society for Cell Biology (Kyoto, Japan)
June, 201637

Laboratory for

Lab activities

Integrated Cellular
Systems
Team Leader: Mariko Okada

Figure: A mathematical model of the BCR- NF-κB
network
Left: A model overview. Right: Detailed diagram of themodel. Black arrows indicate functional modifications
or association/dissociation of proteins. Gray dashedarrows indicate links of inter-modules (CBM complex,
TAK1 phosphorylation, IKK phosphorylation, IKK-IκBNF-κB binding, transport, transcription and translation).

Recent Major Publications
Inoue K, Shinohara H, Behar M, Yumoto N, Tanaka G,
Hoffmann A, Aihara K, Okada-Hatakeyama M. Oscillation dynamics underlies functional switching of NF-κB
for B cell activation. NPJ Syst Biol Appl 2, 16024(2016)
Yoshida K, Maekawa T, Zhu Y, Renard-Guillet C, Chatton
B, Inoue K, Uchiyama T, Ishibashi K, Yamada T, Ohno N,
Shirahige K, Okada-Hatakeyama M, Ishii S. The transcription factor ATF7 mediates lipopolysaccharide-induced epigenetic changes in macrophages involved
in innate immunological memory. Nat Immunol 16,
1034–1043 (2015)
Mina M, Magi S, Jurman G, Itoh M, Kawaji H, Lassmann
T, Arner E, Forrest ARR, Carninci P, Hayashizaki Y, DaubCO, the FANTOM Consortium, Okada-Hatakeyama M,
Furlanello C. Promoter-level expression clustering identifies time development of transcriptional regulatorycascades initiated by ErbB receptors in breast cancer
cells. Sci Rep 5, 11999 (2015)

Invited Presentations
Okada M. “Multiple-scale cooperativity in signal-transcription network for cellular commitment” Trans-Omics: New Approaches in Biology and Medicine 2016
(Fukuoka Japan) November, 2016
Okada-Hatakeyama M.” Experimental and modeling
analysis of signal transduction network in mammaliancells” 2016 A3 Workshop on Interdisciplinary Research
Connecting Mathematics and Biology (Beijing, China)
April, 2016
Okada-Hatakeyama M. “Experimental and mathematical analysis of signaling network”. National Taiwan
University (Taipei, Taiwan) April, 216
Okada-Hatakeyama M. “Switch-like activation of transcription factors in cell decision program”. Academia
Sinica, Taiwan (Taipei, Taiwan) April, 2016
Okada-Hatakeyama M. “Modeling Cellular Signaling
Functions in Mammalian Cells” The 4th Bioscience andBiotechnology International Symposium. Multifaceted
Approaches to Disease Intervention (Yokohama, Japan)
January, 201638

T

he aims of the laboratory are to define the general regulatory rules in signal transduction-transcriptional networks in cell determination processes and to apply
this knowledge of regulatory principles to the understanding and treatment of humandiseases. Using systems biology approaches, we uncovered several unique properties insignal-transcription networks in immune cell development and cancer. For example, thepotency and duration of signaling responses are controlled by self-regulatory mechanisms. One of such examples is negative feedback regulation in signaling cascades. We
recently found that transcriptionally-induced PHLDA1 suppresses activation of ErbB
receptors and downstream kinases after growth factor stimulation of breast cancer cells
and thereby acts as a negative feedback regulator. Our LC-MS analysis showed thatPHLDA1 binds to ErbB3. Mathematical modeling followed by single molecule analysis
of fluorescent-labeled ligand binding to the receptors indicated that PHLDA1 inhibits
high-order oligomerization of ErbB receptors, which suggested a novel inhibitory mechanism of ErbB receptor signaling (Manuscript in revision). On the other hand, intracellular signaling pathways often act as amplifiers to activate transcription factors forthresholding setting during cellular commitment. Our earlier study indicated that IKK
activity is regulated by positive feedback from IKK to TAK1, mediated by the scaffolding
protein CARMA1 at Ser-578, and that this feedback regulation could induce switch-like
activation of NF-κB in B cells (Shinohara et al. Science, 2014). Our recent mathematicalanalysis further indicated that this switch activation of NF-κB is important for induction of target genes, which are co-regulated by NF-κB oscillation dynamics (Inoue, etal. npj Syst. Biol. Appl. 2016). Thus, our studies showed that this signaling-transcription
network is controlled in the short term by post-transcriptional regulation and in thelong term by transcription-mediated regulation. Moreover, the resulting dynamics itself
contains important messages for cell regulatory mechanisms. Our lab also contributed
to research society in terms of large-scale data analysis and mathematical modeling.

Laboratory for

Lab activities

Metabolomics
Team Leader: Makoto Arita

Figure: Untargeted multi-lipidomics platform

Recent Major Publications
Hirabayashi T, Anjo T, Kaneko A, Senoo Y, Shibata A,
Takama H, Yokoyama K, Nishito Y, Ono T, Taya C, Muramatsu K, Fukami K, Muñoz-Garcia A, Brash AR, Ikeda
K, Arita M, Akiyama M, Murakami M. PNPLA1 has a
crucial role in skin barrier function by directing acylceramide biosynthesis. Nature Commun (in press)
Sakayori N, Kikkawa T, Tokuda H, Kiryu E, Yoshizaki K,
Kawashima H, Yamada T, Arai H, Kang JX, Katagiri H,
Shibata H, Innis SM, Arita M, Osumi N. Maternal dietary balance between omega-6 and omega-3 polyunsaturated fatty acids impairs neocortical development
via epoxy metabolites. Stem Cells 34, 470–482 (2016)
Tsugawa H, Cajka T, Kind T, Ma Y, Higgins B, Ikeda
K, Kanazawa M, VanderGheynst J, Fiehn O, Arita M.
MS-DIAL: data-independent MS/MS deconvolution forcomprehensive metabolome analysis. Nature Methods 12, 523–526 (2015)

Invited Presentations
Arita M. “Lipidomics & Discovery of Novel Bioactive
Omega-3 Metabolites” ISOR Inaugural Symposium“Omega-3 in Health Promotion & Disease Management” (Boston, USA) January, 2017
Arita M. “Advanced lipidomics to understand the quality
difference of fatty acids in biological systems” The 39th
Annual Meeting of the Molecular Biology Society ofJapan (Yokohama, Japan) November, 2016
Arita M. (Award Lecture) “Omega-3 fatty acid metabolism in controlling inflammation and related diseases”
The 25th Annual Meeting of Japan Society for Lipid
Nutrition (Akita, Japan) September, 2016
Arita M. “Emerging roles of lipid metabolism in phagocyte function” The 24th International Symposium on
Molecular Cell Biology of Macrophages (Tokyo, Japan)
June, 2016

L

ipids are recognized as extremely diverse molecules. Precise determinationof each molecular species of lipid, namely Lipo-Quality (Quality of Lipids),
becomes a prerequisite not only to understand their biological functions in physiology and disease, but also to discover novel bioactive lipids that may link lipid
metabolism and biological phenotypes. A powerful method for the analysis oflipid metabolites is liquid chromatography tandem mass spectrometry (LC-MS/
MS). Our research is aimed at elucidating structure and function of endogenous
lipid metabolites that regulate inflammation and tissue homeostasis.
A triple quadrupole (TripleQ) mass spectrometer is capable of carrying out
a targeted MS method called multiple reaction monitoring (MRM). A quadrupole Time-of Flight (Q-TOF) mass spectrometer provides the ability to perform
untargeted analyses with high resolution and accurate MS/MS information. The
MS/MS trigger is set at a low threshold level in order to detect as globally as
possible, and lipid structures are unbiasedly annotated by lipid database screening. By taking advantage of Q-TOF (global lipid screening) and TripleQ (quantitative analyses) mass spectrometry, our new approach has a strong potential
to search for lipids of interest globally, and to identify unknown lipid species ina non-biased fashion. We are developing a measured MS/MS library of diverse
lipid molecular species for Q-TOF-based untargeted analyses, especially focusing
on bioactive lipid mediators, oxidized phospholipids, skin barrier lipids such as
acylceramides, and unique lipid metabolites produced by gut microbiota. We arealso developing software that enables us to search for lipid structures more precisely and comprehensively.

Arita M. (Plenary Lecture) “Lipidomic approach to uncover anti-inflammatory properties of omega-3 polyunsaturated fatty acids” 2016 Korea-Japan Bioactive Lipid
Joint Symposium (Jeju, Korea) May, 2016

39

Core for
Genomic Medicine
T

he Core for Genomic Medicine is performing genomic research on humandiseases, especially the common diseases. The aims of Core for Genomic
Medicine are 1) to identify genetic variations related to disease susceptibility,
disease outcome and drug responses (efficacy/adverse reaction), 2) to provide
useful information about possible molecular targets for drug discovery, 3) toexamine the interactions between genetic and environmental factors to understand the pathogenesis and the progression of diseases, and 4) finally toconstruct an evidence base for the implementation of personalized medicine.
To identify genetic variations related to disease susceptibility and drug responses, the Core for Genomic Medicine first showed a proof of concept of
the genome-wide association study (GWAS) in 2002. To advance this strategy,
the Core for Genomic Medicine has organized laboratories to facilitate comprehensive genomic research on common diseases. To produce comprehensive genomic information, the Laboratory for Genotyping Development ismainly working on large-scale SNP genotyping and genome sequencing forvarious diseases. The resulting huge amount of genomic variation data wasmainly analyzed at the Laboratory for Statistical Analysis to extract significant
genomic variations related to disease susceptibility and drug responses.
These laboratories are in close communication with the research group ofpharmacogenomics (Laboratory for Pharmacogenomics and Laboratory forInternational Alliance on Genomic Research), laboratories for disease-causing mechanisms (Laboratory for Cardiovascular Diseases, Autoimmune Diseases, Digestive Diseases, Bone and Joint Diseases, Endocrinology, Metabolism and Kidney Diseases, and Respiratory and Allergic Diseases) and manyother collaborators worldwide for further analyses. In addition to this strategy, the Laboratory for Genome Sequencing Analysis is mainly working onwhole genome sequencing of cancer genomes to clarify the pathogenesis ofcarcinogenesis.

40

Laboratory for

Lab activities

Genotyping Development
Team Leader: Yukihide Momozawa

Figure: Diagram and association results of haplotypes in the C2-CFB-SKIV2L locus with susceptibility to age-related macular degeneration

Recent Major Publications
Momozawa Y, Akiyama M, Kamatani Y, Arakawa S, Yasuda M, Yoshida S, Oshima Y, Mori R, Tanaka K, Mori K,
Inoue S, Terasaki H, Yasuma T, Honda S, Miki A, Inoue
M, Fujisawa K, Takahashi K, Yasukawa T, Yanagi Y, Kadonosono K, Sonoda KH, Ishibashi T, Takahashi A, Kubo
M. Low-frequency coding variants in CETP and CFB areassociated with susceptibility of exudative age-related
macular degeneration in the Japanese population.
Hum Mol Genet 25, 5027–5034 (2016).
Zhou K, Yee SW, Seiser EL, van Leeuwen N, Tavendale R,
Bennett AJ, Groves CJ, Coleman RL, van der Heijden AA,
Beulens JW, de Keyser CE, Zaharenko L, Rotroff DM, Out
M, Jablonski KA, Chen L, Javorský M, Židzik J, Levin AM,
Williams LK, Dujic T, Semiz S, Kubo M, Chien HC, Maeda
S, Witte JS, Wu L, Tkáč I, Kooy A, van Schaik RH, Stehouwer CD, Logie L; MetGen Investigators.; DPP Investigators.; ACCORD Investigators., Sutherland C, Klovins
J, Pirags V, Hofman A, Stricker BH, Motsinger-Reif AA,
Wagner MJ, Innocenti F, Hart LM, Holman RR, McCarthy
MI, Hedderson MM, Palmer CN, Florez JC, Giacomini
KM, Pearson ER. Variation in the glucose transporter
gene SLC2A2 is associated with glycemic response to
metformin. Nat Genet 48, 1055–1059 (2016)
Sun C, Molineros JE, Looger LL, Zhou XJ, Kim K, Okada
Y, Ma J, Qi YY, Kim-Howard X, Motghare P, Bhattarai
K, Adler A, Bang SY, Lee HS, Kim TH, Kang YM, Suh CH,
Chung WT, Park YB, Choe JY, Shim SC, Kochi Y, Suzuki
A, Kubo M, Sumida T, Yamamoto K, Lee SS, Kim YJ, Han
BG, Dozmorov M, Kaufman KM, Wren JD, Harley JB,
Shen N, Chua KH, Zhang H, Bae SC, Nath SK. High-density genotyping of immune-related loci identifies new
SLE risk variants in individuals with Asian ancestry. Nat
Genet 48, 323–330 (2016)

Invited Presentations
Momozawa Y. “Direction of canine behavior genetics”
Meeting for behavior genetics (Mishima, Japan) October, 2016T

he aims of the Laboratory for Genotyping Development are 1) to produce
precise and large-scale genomic data to identify genetic variants related todisease susceptibility, outcomes, and drug responses in close collaboration with
the Laboratory for Statistical Analysis and 2) to develop methods and databases
useful for personalized medicine. Our laboratory has worked as a research hub oflarge-scale genomic analysis, collaborating with domestic and international universities, research institutes, and pharmaceutical companies.
Our laboratory published 51 papers in 2016. Among them, we newly reported
a multiplex PCR based targeted sequencing method that enables us to sequencespecific genomic regions in tens of thousands of samples. This method showeda high concordance rate (99.97%) of genotypes with other established methods.
We applied this method to investigate the contribution of low frequency variants
in 34 genes to age-related macular degeneration (AMD) susceptibility in 2,886
cases and 9,337 controls. While we showed that low frequency variants in the CFB
gene were protective against AMD (odds ratio = 0.43), disruptive variants in the
CETP gene increased disease risk (odds ratio = 2.48). These findings were considered East Asian-specific. In addition, since dysfunction of CETP is also known
to increase high-density lipoprotein cholesterol (HDL-C), a previously unknown
connection between HDL-C and AMD was proposed. These findings highlight theimportance of targeted sequencing to reveal the impact of rare or low-frequency
coding variants on disease susceptibility in different ethnic populations.
We will continue to work as a research hub for large-scale genomic analysis sothat we can contribute to the implementation of personalized medicine.

41

Laboratory for

Lab activities

Genome Sequencing
Analysis
Team Leader: Hidewaki Nakagawa

Recent Major Publications
Furuta M, Ueno M, Fujimoto A, Hayami S, Yasukawa S,
Kojima F, Arihiro K, Kawakami Y, Wardell CP, Shiraishi Y,
Tanaka H, Nakano K, Maejima K, Sasaki-Oku A, Tokunaga N, Boroevich KA, Abe T, Aikata H, Ohdan H, Gotoh K,
Kubo M, Tsunoda T, Miyano S, Chayama K, Yamaue H,
Nakagawa H. Whole genome sequencing discriminates
hepatocellular carcinoma with intrahepatic metastasis
from multi-centric tumors. J Hepatol 66, 363–373(2017)
Alexandrov LB, Ju Y, Haase K, Van Loo P, Martincorena I,
Nik-Zainal S, Totoki Y, Fujimoto A, Nakagawa H, Shibata
T, Campbell PJ, Vineis P, Phillips DH, Stratton MR. Mutational signatures associated with tobacco smoking inhuman cancer. Science 354, 618–622 (2016)
Fujimoto A, Furuta M, Totoki Y, Tsunoda T, Kato M, Shiraishi Y, Tanaka H, Taniguchi H, Kawakami Y, Ueno M,
Gotoh K, Ariizumi S, Wardell CP, Hayami S, Nakamura
T, Aikata H, Arihiro K, Boroevich KA, Abe T, Nakano
N, Maejima K, Sasaki-Oku A, Ohsawa A, Shibuya T,
Nakamura H, Hama N, Hosoda F, Arai Y, Ohashi S,
Urushidate T, Nagae G, Yamamoto S, Ueda H, Tatsuno
K, Ojima H, Hiraoka N, Okusaka T, Kubo M, Marubashi
S, Yamada T, Hirano S, Yamamoto M, Ohdan H, Shimada K, Ishikawa O, Yamaue H, Chayama K, Miyano S,
Aburatani H, Shibata T, Nakagawa H. Whole genome
mutational landscape and characterization of non-coding and structural mutations in liver cancer. Nature
Genetics 48, 500–509 (2016)

Invited Presentations
Nakagawa H. “Sequencing analysis of plasma and pancreatic juice cell-free DNAs for cancer diagnosis” The75th Annual Meeting of Japanese Cancer Association(Yokohama, Japan) October, 2016
Nakagawa H and Aburatani H. “Molecular analysis forliver cancer progression: step-wise progression and recurrent diagnosis” The 12th Scientific Workshop of ICGC
(Boston, USA) September, 2016
Nakagawa H. “Immunogenomic Analysis in PCAWG(PanCancer Analysis for Whole Genomes) of ICGC/
TCGA” The 25th Korean Genome Organization Annual
Conference (Seoul, Korea) September, 2016
Nakagawa H. “HLA genotyping and immunogenomics
of PanCancer” PCAWG Workshop, ICGC/TCGA (Barcelona, Spain) April, 2016
Nakagawa H. “Whole genome sequencing of liver cancer and forwarding to precision medicine” The 102nd
Annual Meeting of Japanese Society of Gastroenterology (Tokyo, Japan) April, 2016

42

Figure: Molecular discrimination of multi-centric tumors (MC) and intrahepatic
metastasis (IM) of liver cancer by the number of shared somatic SNVs detected bywhole genome sequencing (WGS).
Phylogenetic tree depicts the patterns of clonal evolution of the IND15_1 tumor nodule,
which showed complete remission after sorafenib treatment, and the IND15_2 nodule,
which was diagnosed as a MC after complete remission by sorafenib, but WGS analysis indicated that it was an IM after clonal switching.

C

ancer is essentially a “disease of the genome” that evolves in the backgroundof germline variants with the accumulation of diverse mutations caused byenvironmental exposure. Germline variants are biomarkers predisposing to cancer development and genetic analysis of certain specific genes, such as BRCA1/2,
is commonly performed for cancer risk, while somatic mutations of driver genes
have been targeted for cancer treatment and genotype-based personalized cancer
therapy is now a reality. Furthermore, emerging immune therapies, which modulate immune cell activity or immuno-genes in cancer cells, have been demonstrated to be effective in many types of cancers. Now we have to understand more
about cancer genome-immune interactions and their diversity to develop new andmore effective therapies. Recent explosive advances in next-generation sequencing (NGS) and bioinformatics enable a systematic, genome-wide identificationof all somatic abnormalities and immune activity within cancer tissues by wholegenome sequencing (WGS), whole exome sequencing and RNA sequencing. It is
important to analyze the likely implications of the huge genome datasets fromNGS and to reach a consensus about how to interpret the biological and clinicalaspects of somatic and germline variants. Our research objectives are to understand cancer genome and microenvironmental activities, including immune activity, by utilizing NGS to identify novel genomic biomarkers and to develop analysis
platforms that can be used in clinics for precision medicine.

Laboratory for

Lab activities

Statistical Analysis
Team Leader: Yoichiro Kamatani

Figure: Enrichment of obesity susceptible SNPs
in CD19+ primary B cells
We performed GWAS for obesity using 173,430 subjects and found 85 genome-wide significant loci, including 49 novel obesity variants. We used them andconducted enrichment analysis of these significant variants into cell-type specific regulatory marks obtained
from Roadmap H3K4me3 epigenomics data. We successfully replicated previously reported enrichment ofobesity variants in pancreatic islets and brain tissue,
and found novel enrichment in primary CD19+ B cells.

Recent Major Publications
Shiraishi K, Okada Y, Takahashi A, Kamatani Y, Momozawa Y, Ashikawa K, Kunitoh H, Matsumoto S, Takano
A, Shimizu K, Goto A, Tsuta K, Watanabe S, Ohe Y,
Watanabe Y, Goto Y, Nokihara H, Furuta K, Yoshida A,
Goto K, Hishida T, Tsuboi M, Tsuchihara K, Miyagi Y,
Nakayama H, Yokose T, Tanaka K, Nagashima T, Ohtaki
Y, Maeda D, Imai K, Minamiya Y, Sakamoto H, Saito A,
Shimada Y, Sunami K, Saito M, Inazawa J, Nakamura
Y, Yoshida T, Yokota J, Matsuda F, Matsuo K, Daigo Y,
Kubo M, Kohno T. Association of variations in HLA class
II and other loci with susceptibility to EGFR-mutatedlung adenocarcinoma. Nat Commun 7, 12451 (2016)
Okada Y, Suzuki A, Ikari K, Terao C, Kochi Y, Ohmura K,
Higasa K, Akiyama M, Ashikawa K, Kanai M, Hirata J,
Suita N, Teo YY, Xu H, Bae SC, Takahashi A, Momozawa
Y, Matsuda K, Momohara S, Taniguchi A, Yamada R, Mimori T, Kubo M, Brown MA, Raychaudhuri S, Matsuda
F, Yamanaka H, Kamatani Y, Yamamoto K. Contributionof a Non-classical HLA Gene, HLA-DOA, to the Risk ofRheumatoid Arthritis. Am J Hum Genet 99, 366–374(2016)
Okada Y, Momozawa Y, Ashikawa K, Kanai M, Matsuda
K, Kamatani Y, Takahashi A, Kubo M. Construction of
a population-specific HLA imputation reference panel
and its application to Graves' disease risk in Japanese.
Nat Genet 47, 798–802 (2015)

Invited Presentations
Kamatani Y. “The Biobank Japan” International Stroke
Genetics Consortium 2016 Fall (Milan, Italy) November,
2016
Kamatani Y. “Genetic analysis of non-European population” Neurepiomics (Boston, USA) September, 2016
Kamatani Y. “The largest Asian GWAS of ischemic
stroke” International Stroke Genetics Consortium 2016
Spring (Boston, USA) May, 2016O

ur laboratory aims at identifying susceptibility variants of complex traits by using
genetic association study, as well as connecting the genetic association findings
with biology and medicine. Until now, we have used genome-wide SNP array data andperformed genome wide association studies (GWAS) for dozens of traits. In 2016, wehave completed GWAS for 45 different diseases using samples from ~200,000 BioBank
Japan participants with ~30,000 population controls genotyped for nearly a million
SNPs. Additionally, we have performed GWAS for more than 50 quantitative traits including anthropometric traits and clinical laboratory data, and also for life-style using
the BioBank Japan clinical information database. These are catalogs of the largest
non-European GWAS ever for each trait.
We then moved into addressing three key issues in this field; missing heritability,
functional interpretation of the identified genetic variants, and the genetic architecture of
the diseases in question. Firstly, we have started whole genome sequence (WGS) analysis
for thousands of patients affected by common disorders to evaluate the contribution ofrare variants or structural variations which are not covered by SNP array data. Genotype
imputation of the SNP results of the ~230,000 subjects described above using a WGSbased reference panel is being conducted in parallel. Secondly, we are now focusing on
the interpretation of GWAS signals. We conducted integrative analysis of GWAS signals
with publicly available expression and epigenomic data, and found a novel genetically-associated immune cell, the CD19+ B cell, for obesity, which should be one of the target celltypes to elucidate functional roles of GWAS variants. We are also using in-house eQTL
data of lymphocyte subtypes generated by the Laboratory for Autoimmune Diseases andlong non-coding RNA expression data from several cell types as a collaborative projectwith the RIKEN Center for Life Science Technologies (CLST). Lastly, we are attempting to
use machine learning techniques to reveal non-additive effects of genomic variants thathave not been investigated well in this field. The novel machine learning techniques will
facilitate detecting such effects.

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Laboratory for

Lab activities

Pharmacogenomics
Group Director: Taisei Mushiroda

Figure: Identification of genomic biomarkers
associated with drug responses and implementation into clinical practice for pharmacogenomics-based individualization of drug therapy

Recent Major Publications
Wattanapokayakit S, Mushiroda T, Yanai H, Wichukchinda N, Chuchottawon C, Nedsuwan S, Rojanawiwat
A, Denjanta S, Kantima T, Wongyai J, Suwankesawong
W, Rungapiromnan W, Kidkeukarun R, Bamrungram
W, Chaiwong A, Suvichapanich S, Mahasirimongkol
S, Tokunaga K. NAT2 slow acetylator associated with
anti-tuberculosis drug-induced liver injury in Thai patients. Int J Tuberc Lung Dis 20, 1364–1369 (2016)
Fukunaga K, Nakagawa H, Ishikawa T, Kubo M, Mushiroda T. ABCB1 polymorphism is associated with atorvastatin-induced liver injury in Japanese population.
BMC Genet 17, 79 (2016)Low SK, Fukunaga K, Takahashi A, Matsuda K, Hongo
F, Nakanishi H, Kitamura H, Inoue T, Kato Y, Tomita Y,
Fukasawa S, Tanaka T, Nishimura K, Uemura H, Hara
I, Fujisawa M, Matsuyama H, Hashine K, Tatsugami K,
Enokida H, Kubo M, Miki T, Mushiroda T. Association
Study of a Functional Variant on ABCG2 Gene with
Sunitinib-Induced Severe Adverse Drug Reaction. PLoS
One 11, e0148177 (2016).

Invited Presentations
Mushiroda T. “Current status of pharmacogenomics inJapan and abroad and germline BRCA mutation testing
to determine eligibility for PARP inhibitor maintenancetherapy” The 37th Annual Meeting of The JapaneseSociety of Clinical Pharmacology and Therapeutics (Yonago, Japan) December, 2016
Mushiroda T, “Identification of genomic biomarkers
associated with cutaneous adverse drug reactions andvalidation of clinical utility of genetic testing” The 31st
Annual Meeting of The Japanese Society for the Studyof Xenobiotics (Matsumoto, Japan) October, 2015
Mushiroda T, “Identification of genomic biomarkers
associated with drug responses and pharmacogenomics-based individualization of drug therapy” The 2nd
Annual Meeting of Japanese Society of Drug Safety(Gifu, Japan) July, 2016
Mushiroda T, “Pharmacogenomics-based individualization of drug therapy” The 89th Annual Meeting of The
Japanese Pharmacological Society (Yokohama, Japan)
March, 2016

44

G

enomic analyses, such as genome-wide association study (GWAS) and HLA genotyping, are effective for the identification of pharmacogenomics (PGx) biomarkers.
GWAS presently stands as a standard method in IMS, but can explain only a part of therelationship between genomic variations and drug responses. Thus, we have developed
a new targeted re-sequencing method that can identify all genomic variations in 100
drug-related genes (pharmacogenes) using next generation sequencing. In addition to
genomic analysis, pharmacokinetics and metabolomics analyses using ultra high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) may be necessary for development of accurate predictive systems for risk of adverse drug reactions(ADRs) and for drug efficacy.
Although we have mainly used BioBank Japan, which contains DNA samples andclinical information from about 220,000 Japanese patients, it is difficult for individual
countries acting alone to collect a sufficient number of samples for PGx research. We willcontinue two international PGx collaborative projects with the South East Asian Pharmacogenomics Research Network (SEAPharm) and the US NIH Pharmacogenomics Research Network (PGRN).
Since our mission is implementation of PGx testing, we will conduct prospective clinical trials to test the clinical utility of genetic test using the PGx biomarkers identified byour basic research. If successful, this will lead to use of the PGx biomarkers as in-vitro
diagnostics under the Japan national health insurance system. However, from the viewpoint of a limited budget and human resources, it will not be practical to conduct clinical
trials for all of the identified PGx biomarkers. Thus, we will have to create a plan to investigate whether pre-emptive PGx testing is clinically useful for patients and cost-effective
without actually conducting prospective clinical trials. This strategy is similar to that of
the Ubiquitous Pharmacogenomics (U-PGx) project, which has started prospective collection of pre-emptive PGx testing data and its embedment into the electronic medical
record in European countries.

Laboratory for

Lab activities

Cardiovascular Diseases
Team Leader: Kaoru Ito

Figure: The Landscape of Coronary Artery Diseases revealed by Modified Phenome-wide
Association Study
Significant SNPs are identified by a conventional GWAS
approach. To understand the biological and clinical implications of those SNPs, the modified phenome-wide
association study reveals intermediate phenotypes andthe relationship among them.

Recent Major Publications
Konta A, Ozaki K, Sakata Y, Takahashi A, Morizono T,
Suna S, Onouchi Y, Tsunoda T, Kubo M, Komuro I, Eishi
Y, Tanaka T. A functional SNP in FLT1 increases risk ofcoronary artery disease in a Japanese population. J
Hum Genet 61, 435–441 (2016)
Ozaki K, Tanaka T. Molecular genetics of coronary arterydiseases. J Hum Genet 61, 71–77 (2016)
Onouchi Y, Fukazawa R, Yamamura K, Suzuki H, Kakimoto N, Suenaga T, Takeuchi T, Hamada H, Honda T,
Yasukawa K, Terai M, Ebata R, Higashi K, Saji T, Kemmotsu Y, Takatsuki S, Ouchi K, Kishi F, Yoshikawa T, Nagai T, Hamamoto K, Sato Y, Honda A, Kobayashi H, SatoJ, Shibuta S, Miyawaki M, Oishi K, Yamaga H, Aoyagi N,
Yoshiyama M, Miyashita R, Murata Y, Fujino A, Ozaki
K, Kawasaki T, Abe J, Seki M, Kobayashi T, Arakawa H,
Ogawa S, Hara T, Hata A, Tanaka T. Variations in ORAI1
gene associated with Kawasaki Disease. PLoS One 11,
e0145486 (2016)

Invited Presentations
Kaoru I. “High-throughput Splicing Analysis Revealed
LMNA and MYBPC3 Silent Mutations Affect RNA
Splicing and Cause Pathogenicity” American Heart
Association Scientific Session 2016 (New Orleans, USA)
November, 2016
Kaoru I. “Searching for a hidden variant for laminopathy
using machine learning and NGS” The 20th Annual
Scientific Meeting of the Japanese Heart Failure Society(Sapporo, Japan) October, 2016
Kaoru I. “Heart Failure from the viewpoint of Genetics”
The 20th Annual Scientific Meeting of the JapaneseHeart Failure Society (Sapporo, Japan) October, 2016
Kaoru I. “Heart Failure and Atherosclerosis in Genetics”
Science Harmony vol.3 (Tokyo, Japan) October, 2016
Kaoru I. “High-throughput Multiplexed Splicing Analysis
Pipeline Revealed that Lamin A/C Synonymous andMissense Mutations Affect RNA Splicing and Cause
Pathogenicity.” The 22nd Congress of TMFC (Osaka,
Japan) July, 2016

S

ince cardiovascular diseases cause more than 15% of the deaths in the Japanese population and represent more than 20% of the total medical expenses inJapan, it is important for our society to understand the mechanisms underlying
these disorders and to uncover new therapeutic targets for their treatment. To
achieve these goals, we combine “dry” (NGS technologies, OMICS strategies andcomputer science) and “wet” (so-called conventional molecular biology) technologies, in an attempt to achieve a comprehensive and precise understanding ofthese diseases. In other words, there is no border between “dry” and “wet” research in our lab, since we postulate that both will be required in order to expand
the horizon of our knowledge.
Diseases of our interest to date are coronary artery diseases (CAD), atrial fibrillation (AF), Kawasaki disease (KD), peripheral artery disease (PAD) and cardiomyopathy (CM). We are currently seeking to 1) understand the interactive
effect between Japanese top SNPs (e.g. SNPs in BRAP, ALDH2, etc.) and othergenes, 2) provide a reasoned interpretation of significant SNPs from the viewpoint of clinical phenotypes (Modified Phenome-wide Association Study, see Figure), and 3) develop a more sophisticated genetic risk scoring system by machinelearning algorithms in the CAD project. Additionally, in the CM project, we aredeveloping an in silico splicing variant prediction algorithm, a high-throughput
cell-based splicing assay and a downstream in silico pipeline to uncover cryptic
splicing variants, which have been overlooked in the current established pipeline.
We are conducting our research with not only a scientific mind but also a clinical eye, because our ultimate goal is to provide improved diagnostic / management / therapeutic approaches for patients suffering from those diseases.

45

Laboratory for

Lab activities

Autoimmune Diseases
Team Leader: Kazuhiko Yamamoto

Figure: eQTL study of peripheral-blood (PB) subsets in Japanese
1. We collected peripheral blood samples from Japanese healthy volunteers and sorted the cells into fivesubsets. We extracted mRNAs and quantified gene
expression by RNA-seq.
2. By examining the association between SNP genotype
and gene expression, we identified all eQTLs to establish an eQTL catalog for each subset.
3. By combining disease GWAS data, eQTL data, andepigenome data using bioinformatics approaches, we
identified disease causal genes to understand diseasemechanisms.

Recent Major Publications
Sun C, Molineros EJ, Looger LL, Zhou X, Kim K, Okada
Y, Ma J, Qi Y, Kim-H X, Motghare P, Bhattarai K, Adler
A, Bang S-Y, Lee HS, Kim TH, Kang YM, Suh CH, Chung
WT, Park YB, Choe JY, Shim SC, Kochi Y, Suzuki A, Kubo
M, Sumida T, Yamamoto K, Lee SS, Kim YJ, Han BG,
Dozmorov M, Kaufman MK, Wren DJ, Harley BJ, Shen N,
Chua KH, Zhang H, Bae SC, Nath KS. High-density genotyping of immune-related loci identifies new SLE risk
variants in individuals with Asian ancestry. Nat Genet
48, 323–330 (2016)
Kochi Y. Genetics of autoimmune diseases: perspectives
from genome-wide association studies. Int Immunol
28, 155–161 (2016)
Yamamoto K, Okada Y, Suzuki A, Kochi Y. Genetics ofrheumatoid arthritis in Asia--present and future. Nat
Rev Rheumatol 11, 375–379 (2015)

Invited Presentations
Yamamoto K. “Regulation of B cell functions and cellular metabolism in autoimmunity” Cold Spring Harbor
Asia (Awaji, Japan) October, 2016
Yamamoto K. “Genomics and functional genomics ofrheumatoid arthritis” 20th Asia Pacific League of Associations for Rheumatology Congress (Shanghai, China)
September, 2016
Yamamoto K. “TGF-β3 as a novel target of immunotherapy” 10th International Congress on Autoimmunity(Leipzig Germany) April, 2016
Yamamoto K. “Genetics of rheumatoid arthritis: From
genetics to functional genetics” 18th Conference on
Advances in Targeted Therapies (ATT) (Palma, Spain)
March, 2016
Yamamoto K. “Genetics of rheumatic diseases: Functional genetics of rheumatoid arthritis.” IRA - APLAR
COURSE ON RHEUMATOLOGY (Jakarta, Indonesia),
February, 2016

46

M

ost autoimmune diseases, such as rheumatoid arthritis (RA), systemic
lupus erythematosus (SLE) and Graves’ disease (GD), are multifactorialdiseases involving both genetic and environmental factors. The aim of our laboratory is to elucidate the etiology of these autoimmune diseases by dissecting their
genetic contributions. Furthermore, through functional analyses of disease-associated variants and genes together with epigenetic studies, we will also clarify the mechanisms of disease pathogenesis. The majority of disease risk variantsof complex traits and diseases are expression quantitative trait loci (eQTLs) andthese eQTLs mostly behave functionally in a cell type-specific manner. Therefore,
subpopulations of immune related cells, such as T cells, B cells, dendritic cells,
macrophages, and others should be isolated and investigated. One advantage of
research on immune mediated diseases is that we can relatively easily obtain such
immune-related cells from human subjects, e.g., from peripheral blood (PB).
Through genetic as well as informatics analyses of these samples, we are investigating the gene expression profiles and functions of each subset of immune-related
cells and will dissect their roles in the global immune systems as well as in several
autoimmune diseases.

Laboratory for

Lab activities

Digestive Diseases
Team Leader: Kazuaki Chayama

Recent Major Publications
Furuta M, Ueno M, Fujimoto A, Hayami S, Yasukawa S,
Kojima F, Arihiro K, Kawakami Y, Wardell CP, Shiraishi
Y, Tanaka H, Nakano K, Maejima K, Sasaki-Oku A,
Tokunaga N, Boroevich KA, Abe T, Aikata H, Ohdan H,
Gotoh K, Kubo M, Tsunoda T, Miyano S, Chayama K,
Yamaue H, Nakagawa H. Whole genome sequencing
discriminates hepatocellular carcinoma with intrahepatic metastasis from multi-centric tumors. J Hepatol
66, 363-373 (2017)
Fujimoto A, Furuta M, Totoki Y, Tsunoda T, Kato M, Shiraishi Y, Tanaka H, Taniguchi H, Kawakami Y, Ueno M,
Gotoh K, Ariizumi S, Wardell CP, Hayami S, Nakamura
T, Aikata H, Arihiro K, Boroevich KA, Abe T, Nakano
K, Maejima K, Sasaki-Oku A, Ohsawa A, Shibuya T,
Nakamura H, Hama N, Hosoda F, Arai Y, Ohashi S,
Urushidate T, Nagae G, Yamamoto S, Ueda H, Tatsuno
K, Ojima H, Hiraoka N, Okusaka T, Kubo M, Marubashi
S, Yamada T, Hirano S, Yamamoto M, Ohdan H, Shimada K, Ishikawa O, Yamaue H, Chayama K, Miyano S,
Aburatani H, Shibata T, Nakagawa H. Whole-genome
mutational landscape and characterization of noncoding and structural mutations in liver cancer. Nat Genet
48, 500–509 (2016)
Fujimoto A, Furuta M, Shiraishi Y, Gotoh K, Kawakami Y,
Arihiro K, Nakamura T, Ueno M, Ariizumi S, Nguyen HH,
Shigemizu D, Abe T, Boroevich KA, Nakano K, Sasaki A,
Kitada R, Maejima K, Yamamoto Y, Tanaka H, Shibuya
T, Shibata T, Ojima H, Shimada K, Hayami S, Shigekawa
Y, Aikata H, Ohdan H, Marubashi S, Yamada T, Kubo M,
Hirano S, Ishikawa O, Yamamoto M, Yamaue H, Chayama K, Miyano S, Tsunoda T, Nakagawa H. Whole-genome mutational landscape of liver cancers displaying
biliary phenotype reveals hepatitis impact and molecular diversity. Nat Commun 6, 6120 (2015)

Invited Presentations
Chayama K. “Best Strategy for Curing HCV Genotype 1
and 4: East vs. West” 5th The Asian Pacific Association
for the Study of the Liver (APASL) Single Topic Conference on Hepatitis C (Kaohsiung, Taiwan) June, 2016
Chayama K. “HCV Treatment Update-a New Era of Alloral HCV Treatment” The 46th Annual Meeting of GEST
and The 25th Annual Meeting of DEST (Taipei, Taiwan)
March, 2016

Figure: The effects of IFNL4 and ITPA polymorphisms on novel regimes forthe treatment of HCV.
a) Relationship between treatment response and IFNL4 genotype in older Japanesepatients with genotype 1 chronic hepatitis C. Sustained virological response rates for
simeprevir triple therapy grouped by response to prior interferon treatment and IFNL4
genotype. b) Reduction of hemoglobin levels correlated with an ITPA polymorphism
during sofosbuvir and ribavirin combination treatment. Patients were grouped by ITPA
rs1127354 genotype.

Using a GWAS approach, we have focused on investigating host genetic factors
involved in various liver diseases, such as chronic HBV and HCV infection,
HCV-induced liver cirrhosis and cancer, and responsiveness to therapy. We have now
intensively investigated and verified how to apply such genetic information to clinicalpractice. We demonstrated that a polymorphism in the interferon lambda-4 (IFNL4) geneaffects the outcome of simeprevir, peginterferon and ribavirin therapy for older patientswith genotype 1 chronic hepatitis C (Hepatol Res in press). Analysis of IFNL4 polymorphisms is a valuable predictor in both younger and older patients. We also found that an
inosine triphosphatase (ITPA) polymorphism influences hemoglobin levels and incidenceof ribavirin dose reduction during simeprevir triple therapy, indicating the importance ofmonitoring anemia during treatment, particularly for ITPA genotype CC patients (Hepatol Res 2016). In addition, we investigated 244 patients with genotype 2 chronic hepatitis
C, who were treated with the interferon-free regimen of sofosbuvir plus ribavirin, andreported ITPA polymorphism effects on the decrease of hemoglobin during combination
therapy (J Gastroenterol in press).
We have also participated in whole-genome sequencing (WGS) analyses that haveidentified several recurrently mutated genes/pathways, HBV integration events, mutations in non-coding regions, and structural variations associated with liver cancer, incollaboration with Laboratory for Genome Sequencing Analysis (Nat Genet 2016, Nat
Commun 2015). We recently showed that WGS of multiple liver tumors enabled the accurate diagnosis of multi-centric occurrence and intrahepatic metastasis prior to selectinga therapeutic strategy for multiple tumors in the liver (J Hepatol in press). By using WGS
data, we analyzed circulating tumor DNA and found that it reflects tumor progression,
microscopic vascular invasion of the portal vein and cancer recurrence (Cell Mol Gastroenterol Hepatol 2015).
In collaboration with other IMS teams, we are currently investigating the host immune response to further elucidate the mechanism of hepatitis and carcinogenesis.

47

Laboratory for

Lab activities

Bone and Joint Diseases
Team Leader: Shiro Ikegawa

Figure: LRRK1 mutant does not rescue the bone
resorption defect of Lrrk1-deficient osteoclasts.
(A) Expression of Flag-tagged wild type and mutant(p.E1980Afs*66) human LRRK1 proteins. Western blot
using anti-Flag antibody. (B) Efficient transduction oflentiviral particle in osteoclast precursors. The transduction efficiency was monitored by GFP expression. (C)TRAP positive osteoclasts and resorptive pit formation
on bone slices (100x). Left panel: lentivirus transduced
monocytes were differentiated on bone slices for 6days, followed by TRAP staining. Right panel: transduced monocytes were differentiated on bone slices for 10
days. (D) Quantitative data of TRAP-positive multinuclear cells (left) and pit formation (right) (N = 9). Data
are mean ± SEM. *: P< 0.01.

Recent Major Publications
Guo L, Girisha KM, Iida A, Hebbar M, Shukla A, ShahH, Nishimura G, Matsumoto N, Nismath S, Miyake N,
Ikegawa S. Identification of a novel LRRK1 mutation ina family with osteosclerotic metaphyseal dysplasia. J
Hum Genet 62, 437–441 (2017)
Nishimura G, Nakajima M, Takikawa K, Haga N, Ikegawa S. Distinctive skeletal phenotype in high bone massosteogenesis imperfecta due to a COL1A2 cleavage site
mutation. Am J Med Genet A 170, 2212–2214 (2016)
Guo L, Yamashita H, Kou I, Takimoto A, Meguro-Horike
M, Horike S, Sakuma T, Miura S, Adachi T, Yamamoto T,
Ikegawa S, Hiraki Y, Shukunami C. Functional investigation of a non-coding variant associated with adolescentidiopathic scoliosis in zebrafish: elevated expression of
the ladybird homeobox gene causes body axis deformation. PLoS Genet 12, e1005802 (2016)

Invited Presentations
Ikegawa S. “How to Study Genetic Diseases.” Taipei
Medical University Invited Seminar (Taipei, Taiwan)
January 2017
Ikegawa S. “Identification of the disease gene by wholeexome sequencing” 10th Annual Introductory Course on
Skeletal Dysplasia (Lausanne, Switzerland) July, 2016
Ikegawa S. “Why study scoliosis genetics? A geneticalperspective” (Singapore, Singapore) May, 2016.
Ikegawa S. “Genomic study of common spinal diseases.” 6th Jishuitan Orthopedics Forum (Beijing, China)
April, 2016
Ikegawa S. “How to identify disease genes in skeletal
dysplasia” Third Scandinavian Skeletal Dysplasia Workshop (Oslo, Norway) March, 201648a) Genomic Study of Common Diseases
Common bone and joint diseases have become serious concerns for the world health andeconomy, as exemplified by the WHO initiative “Bone and Joint Decade” (2000–2010) andthe “Locomo” campaign in Japan. We are searching for susceptibility genes for common(polygenic) bone and joint diseases including osteoarthritis (OA), lumbar disc disease(LDD)/herniation (LDH), osteoporosis, avascular necrosis of the femoral head (ANF),
scoliosis, and ossification of the posterior longitudinal ligament of the spine (OPLL).
Through genome-wide association studies (GWASs) and next-generation sequencing
approaches, we identify and characterize susceptibility genes, and then clarify their disease-causing mechanisms at the molecular level. Using the genome information obtainedby these studies, we will realize our final goal, “precision medicine” for these diseases.
GWASs for OA, LDD/LDH, adolescent idiopathic scoliosis (AIS), OPLL, and ANF are in
progress, and some of them have already succeeded in identification of susceptibility
genes. Functional studies of the genes in vitro and using animal models are underway.
b) Genomic Study of Skeletal Dysplasia
Skeletal dysplasia is a group of heritable (monogenic) disorders affecting the skeleton,with more than 450 diseases belonging to this category. Skeletal dysplasia is an intractable disease and thus many patients are waiting for treatment. We are engaging in clinicaland basic studies of these difficult diseases. By large-scale mutation screening, includingexome sequencing, we are identifying the disease genes. So far, we identified new genes
for 24 diseases. One of them which we recently identified is LRRK1 in osteosclerotic metaphyseal dysplasia. We found an elongation mutation of the gene that resulted in the lossof the bone-resorbing capacity of osteoclasts (Figure).
Though analysis of their phenotypes and diseases genes, we approach molecularmechanisms of bone and joint formation, pathogenesis of common bone and joint diseases, as well as the diagnosis and treatment of these crippling intractable diseases (‘nanbyo’). Using the disease genes for skeletal dysplasia as candidate genes, we are performing
association studies for the corresponding common bone and joint diseases.

Laboratory for

Lab activities

Diabetes, Metabolism
and Renal diseases
Team Leader: Momoko Horikoshi

Figure: Results of GWAS meta-analysis for type 2
diabetes in the Japanese population
Names of the previously identified loci are in black andthe seven novel loci identified in this study are shownin red.

Recent Major Publications
Horikoshi M, Beaumont RN, Day FR, Warrington NM,
Kooijman MN, Fernandez-Tajes J, Feenstra B, vanZuydam NR, Gaulton KJ, Grarup N, Bradfield JP, Strachan DP, Li-Gao R, Ahluwalia TS, Kreiner E, Rueedi R,
Lyytikäinen LP, Cousminer DL, Wu Y, Thiering E, Wang
CA, Have CT, Hottenga JJ, Vilor-Tejedor N, Joshi PK, Boh
ET, Ntalla I, Pitkänen N, Mahajan A, van Leeuwen EM,
Joro R, Lagou V, Nodzenski M, Diver LA, Zondervan
KT, Bustamante M, Marques-Vidal P, Mercader JM,
Bennett AJ, Rahmioglu N, Nyholt DR, Ma RC, Tam CH,
Tam WH; CHARGE Consortium Hematology Working
Group., Ganesh SK, van Rooij FJ, Jones SE, Loh PR, Ruth
KS, Tuke MA, et al. Genome-wide associations for birthweight and correlations with adult disease. Nature
538, 248–252 (2016)
Fuchsberger C, Flannick J, Teslovich TM, Mahajan A,
Agarwala V, Gaulton KJ, Ma C, Fontanillas P, Moutsianas L, McCarthy DJ, Rivas MA, Perry JR, Sim X, Blackwell TW, Robertson NR, Rayner NW, Cingolani P, Locke
AE, Fernandez Tajes J, Highland HM, Dupuis J, Chines
PS, Lindgren CM, Hartl C, Jackson AU, Chen H, HuygheJR, van de Bunt M, Pearson RD, Kumar A, Müller-Nurasyid M, Grarup N, Stringham HM, Gamazon ER, Lee J,
Chen Y, Scott RA, Below JE, Chen P, Huang J, Go MJ,
Stitzel ML, Pasko D, Parker SC, Varga TV, Green T, Beer
NL, Day-Williams AG, Ferreira T, Fingerlin T, Horikoshi
M, et al. The genetic architecture of type 2 diabetes.
Nature 536, 41–47 (2016)O

ur primary focus is on establishing the genetic contribution to type 2 diabetes (T2D) susceptibility in the Japanese population. To that end, we areworking directly with the state-of-the-art genetic resources generated by Biobank
Japan, which includes GWAS data for more than 35,000 T2D subjects. Our recent analyses of a subset of these GWAS data identified seven novel regions of the
genome associated with T2D (Fig). Of the more than 90 T2D loci reported as of2016, we have increased the number of loci reported from the Japanese population
to 14 in total. We are currently expanding this effort to the full set of Biobank
Japan. We have also investigated genomic regions associated with microvascularcomplications of T2D, namely, diabetic retinopathy and nephropathy. GWAS for
these complications have been performed by several groups, but worldwide efforts
to identify susceptibility to these diabetic complications have not met with clear
success. We are strengthening our ties with the neighboring collaborators by contributing our T2D association data to the Asian Genetic Epidemiology Network(AGEN) Consortium as well as to the world-wide DIAMANTE Consortium.

Imamura M, Takahashi A, Yamauchi T, Hara K, Yasuda
K, Grarup N, Zhao W, Wang X, Huerta-Chagoya A, Hu
C, Moon S, Long J, Kwak SH, Rasheed A, Saxena R, Ma
RC, Okada Y, Iwata M, Hosoe J, Shojima N, Iwasaki M,
Fujita H, Suzuki K, Danesh J, Jørgensen T, Jørgensen
ME, Witte DR, Brandslund I, Christensen C, Hansen T,
Mercader JM, Flannick J, Moreno-Macías H, Burtt NP,
Zhang R, Kim YJ, Zheng W, Singh JR, Tam CH, Hirose
H, Maegawa H, Ito C, Kaku K, Watada H, Tanaka Y,
Tobe K, Kawamori R, Kubo M, Cho YS, Chan JC, et al.
Genome-wide association studies in the Japanese
population identify seven novel loci for type 2 diabetes.
Nat Commun 7, 10531 (2016)

49

Laboratory for

Lab activities

Respiratory andAllergic Diseases
Team Leader: Mayumi Tamari

Figure: Manhattan plot of the wheat-dependent
exercise-induced anaphylaxis (WDEIA) due to
HWP
GWAS of WDEIA induced by HWP-containing facial
soap “Cha-no-shizuku”. Manhattan plot showing the–log10 P values in the GWAS for cases and controls
plotted against their respective positions on autosomes. The red line shows the genome-wide significance threshold for this study (P = 5 × 10–8).

Recent Major Publications
Yatagai Y, Hirota T, Yamada H, Masuko H, Kaneko Y, Iijima H, Naito T, Noguchi E, Tamari M, Kubo M, Takahashi
A, Konno S, Makita H, Nishimura M, Hijikata M, Keicho
N, Homma S, Taguchi Y, Azuma A, Kudoh S, Hizawa
N. Variants near the HLA complex group 22 geneconfer increased susceptibility to late-onset asthma inJapanese populations. J Allergy Clin Immunol 138,
281–283 (2016)
EArly Genetics and Lifecourse Epidemiology (EAGLE)
Eczema Consortium. Multi-ancestry genome-wide
association study of 21,000 cases and 95,000 controls
identifies new risk loci for atopic dermatitis. Nat Genet
47, 1449–1456 (2015)
Tokunaga T, Sakashita M, Haruna T, Asaka D, Takeno S,
Ikeda H, Nakayama T, Seki N, Ito S, Murata J, Sakuma
Y, Yoshida N, Terada T, Morikura I, Sakaida H, Kondo K,
Teraguchi K, Okano M, Otori N, Yoshikawa M, Hirakawa
K, Haruna S, Himi T, Ikeda K, Ishitoya J, Iino Y, Kawata
R, Kawauchi H, Kobayashi M, Yamasoba T, Miwa T,
Urashima M, Tamari M, Noguchi E, Ninomiya T, Imoto Y,
Morikawa T, Tomita K, Takabayashi T, Fujieda S. Novelscoring system and algorithm for classifying chronicrhinosinusitis: the JESREC Study. Allergy 70:995–1003(2015)

Invited Presentations
TTamari M. “Genetic Study of Allergic Diseases” Graduate School Seminar in Fukui University (Fukui, Taiwan)
December, 2016
Tamari M. “Genomics in Allergic Disease” Symposium,
The 44th meeting of society for inhalation therapy. (Tokyo, Japan) November, 2016
Tamari M. “Gene to Environmental Interactions in Asthma” Joint Congress of APAAACI AND APAPARI 2016.
(Kuala Lumpur, Malaysia) October, 2016
Tamari M. “Genomics in Allergic Disease” Genomic
Medicine Promotion Forum in Nippon Medical School.
(Tokyo, Japan) July, 2016
Tamari M. “Genetic Study of Allergic Diseases”
PGRN-RIKEN IMS Strategic Alliance Meeting. (San Francisco, USA) April, 201650

T

he aim of our project is to explore genetic components of respiratory andallergic diseases. Genome-wide association study (GWAS) is a method tocomprehensively assess genes underlying susceptibility to human disorders. Foodallergy (FA) is a problem throughout the world, but the genetic factors affecting
FA susceptibility in children remain largely unexplored. We recruited patientswith FA who were diagnosed by pediatric specialists based on positive oral foodchallenge or a definitive clinical history after food intake. We performed GWAS
for childhood FA in Japanese populations, and identified a total of three loci withgenome-wide significance. One of the loci contained a gene that plays an essential
role in Th2 cytokine expression. Wheat-dependent exercise-induced anaphylaxis (WDEIA) is severe food allergy that usually develops after ingestion of wheat
products followed by physical exercise. Hydrolysed wheat gluten protein (HWP)
is used as an additive for facial soap. In Japan, a total of 2026 cases of immediate wheat allergy and WDEIA due to HWP have been reported. Most patients
seemed to be sensitized to HWP (Glupearl 19S®) through the use of the facial soap
“Cha-no-shizuku”. Glupearl 19S® is a degraded gluten made from the direct resolution of wheat by hydrochloric acid. We conducted GWAS of WDEIA induced by
HWP-containing facial soap of 464 cases and 3,099 controls. SNPs at a region on
chromosome 6 were associated with WDEIA induced by HWP-containing facial
soap (Figure). We are conducting a replication study in an independent sampleset.
We participate in the Global Alliance between the U.S. NIH Pharmacogenetics
Research Network (PGRN) and RIKEN IMS. A pharmacogenetic study of asthma
is ongoing now in this international collaboration.

Program for
Medical Innovations
Six original projects for clinical applications have been performed in Program for Medical Innovations. 1) A biochemical drug using a PEGylated
Cryj-1/2 fusion recombinant protein for Cedar Pollinosis has been developedby the Torii pharmaceutical company and IMS and introduced to the Japanese market. 2) A chemical compound recently developed using the α-galactosylceramide (α-GalCer) analog, RCAI-X, selectively induces apoptosis of IgE
but not IgG B cells and preferentially suppresses IgE production. It therefore
potentially could be applied to any type of allergic disorder, such as pollinosis,
food allergy, as well as allergic asthma. 3) NKT cell-targeted therapy for headand neck tumors has been done in collaboration with Chiba University, andwas authorized by the Japanese government as Advanced Medical Treatment
B. 4) The artificial adjuvant vector cell as an anti-tumor vaccine project has
been accepted by the translational research network program and developed. This vaccine can be dosed with tumor antigen mRNA together with
α-GalCer, so that it activates both innate and acquired protective immunity
and also induces long-term memory. 5) The human iPS project for clinical useof in vitro-generated NKT cells has been accepted as a Center for Clinical Application Research (Type B) in the Research Center Network for Realization ofRegenerative Medicine, Japan. 6) A humanized mouse model for MLL
gene-rearranged leukemia was established. Also, certain genes were identified that are differentially expressed between normal stem cells and leukemicstem cells.

51

Laboratory for

Lab activities

Immune Regulation
Group Director: Masaru Taniguchi

Figure: Identification of an alternative pathway
for the development of the Vα14 invariant NKT
cell lineage directly from DN thymocytes thatbypasses the DP stage(A) Demonstration of the DN-stage-thymocyte origin
of NKT cells by a novel fate-mapping approach. Flowcytometry of cells from E8III-Cre+Rosa26-YFP mice.
(B) Presence of NKT cells developed directly from
DN-stage-thymocytes in the E8III-Cre+Rag2fl/fl mousethat lacks the rearranged Trav11Traj18 mRNA at the
DP stage but not at the DN4 stage thymocytes. (C)DN-stage-thymocyte origin NKT cells possess TH1type-biased cytokine secretion, and (D) show superior cytotoxicity against YAC-1 targets compared to
DP-stage-thymocyte origin cells. (E) Proposed modelof an alternative developmental pathway of NKT celllineage cells. Our data demonstrate a presence of apreviously unknown NKT cell developmental pathwaythat originates from DN stage thymocytes, i.e., before
the DP stage of thymopoiesis. This novel developmental pathway gives rise to a subset of DN-NKT cellswith highly potent TH1-type effector functions that are
mainly found in the liver. Our data also suggest that theacquisition of diverse functional characteristics by NKT
cells might be dependent on the timing of TCR expression in precursor cells undergoing positive selection.

Recent Major Publications
Dashtsoodol N, Shigeura T, Aihara M, Ozawa R, Kojo S,
Harada M, Endo T, Watanabe T, Ohara O, Taniguchi M.
Alternative pathway for the development of Vα14+ NKT
cells directly from CD4–CD8– thymocytes that bypasses
the CD4+CD8+ stage. Nat Immunol 18, 274–282(2017)
Yamada D, Iyoda T, Vizcardo R, Shimizu K, Sato Y, Endo
T, Kitahara G, Okoshi M, Kobayashi M, Sakurai M,
Ohara O, Taniguchi M, Koseki H, Fujii S. Efficient regeneration of human V alpha 24+ invariant natural killer T
cells and their anti-tumor activity in vivo. Stem Cells
34, 2852–2860 (2016)
Dashtsoodol N, Shigeura T, Ozawa R, Harada M, Kojo
S, Watanabe T, Koseki H, Nakayama M, Ohara O, Taniguchi M. Generation of Novel Traj18-Deficient Mice
Lacking Vα14 Natural Killer T Cells with an Undisturbed
T Cell Receptor α-Chain Repertoire. PLOS ONE 11,
e0153347 (2016)

Invited Presentations
Dashtsoodol N. “Identification of novel alternative
pathway of NKT cell development” 5th International
Conference “Current Advances in Microbiology and Immunology 2016” (Ulaanbaatar, Mongolia) September,
2016
Taniguchi M. “Overview on development and functionof NKT cells” Immunology Summer School 2016 (Onuma, Japan) July, 2016

52

N

KT cells represent a unique subset of innate-like T lymphocytes expressing
an invariant T cell receptor (TCR) α chain encoded by rearranged Trav11Traj18 gene segments in mice, and TRAV10-TRAJ18 in humans. This receptor isused preferentially by NKT cells but not by conventional T cells, defining NKT
cells as a distinct lineage from conventional T cells. NKT cells recognize glycolipid ligands in conjunction with the monomorphic MHC-like molecule CD1d,and mediate intermediary functions that link the innate and acquired immune
systems by their rapid secretion of large amounts of cytokines such as IL-4, IL17A and IFN-γ after activation. Since the discovery of this unique cell lineage,
the developmental aspect of NKT cells has been one of the most intriguing topics.
Although it is widely believed that NKT cells are generated from CD4+CD8+ double-positive thymocytes, a process termed the DP pathway, it was still not fully
understood whether NKT cells develop exclusively by the DP pathway in a manner closely resembling that of conventional T cells, or whether alternatives to thispathway exist. We have recently tested this long-standing question by generating
mice with DP stage-specific ablation of Rag2 gene expression and by a fate-mapping approach with the use of novel E8III-CreTg mice. Our results demonstrated definitive genetic evidence for the existence of an alternative developmental pathway
through which a fraction of DN NKT cells with strong TH1-biased and cytotoxic
characteristics develops directly from late DN4 stage thymocytes that bypasses the
DP stage. These finding provide new insights into our understanding of NKT cell
development.
Also, our group focuses on translational research, The Japan Agency for Medical Research and Development (AMED), on the establishment of an immunotherapy method using a novel NKT cell glycolipid ligand RK-X that shows superior antitumor and adjuvant effects compared to the widely used glycolipid ligand
αGalCer.

Laboratory for

Lab activities

Immunotherapy
Team Leader: Shin-ichiro Fujii

Figure: Trafficking of CTL to tumor site by aAVC
therapy
C57BL/6 mice were inoculated with MO4 (OVA-expressing B16 melanoma) subcutaneously. A week
later, the mice were treated with aAVC-OVA (α-GalCer
loaded, CD1d-expressing NIH3T3 cells transfected with
OVA mRNA). Vaccinating the animals with aAVC-OVA
caused shrinkage of the otherwise aggressive tumors.
CD8+ T cells (red) were accumulated in the tumor. CD31
(green) identifies blood vessel endothelial cells and
DAPI (blue) stains DNA. Most of the CD8 T cells wereverified as tetramer+ antigen-specific T cells by FACS
analysis.

Recent Major Publications
Yamada D, Iyoda T, Vizcardo R, Shimizu K, Sato Y, Endo
TA, Kitahara G, Okoshi M, Kobayashi M, Sakurai M,
Ohara O, Taniguchi M, Koseki H, Fujii S. Efficient regeneration of Human Vα24+ invariant NKT cells and theiranti-tumor activity in vivo. Stem Cells 34, 2852–2860(2016)
Yamasaki S, Shimizu K, Kometani K, Sakurai M, Kawamura M, Fujii S. In vivo dendritic cell targeting cellular
vaccine induces CD4+ Tfh cell-dependent antibody
against influenza virus. Sci Rep 6, 35173 (2016)
Shimizu K, Yamasaki S, Shinga J, Sato Y, Watanabe T,
Ohara O, Kuzushima K, Yagita H, Komuro Y, Asakura
M, and Fujii S. Systemic DC activation modulates thetumor microenvironment and shapes the long-lived tumor-specific memory mediated by CD8+ T cells. Cancer
Res 76, 3756–3766 (2016)

Invited Presentations
Fujii S. “Cancer Immunotherapy using tumor antigen-derived mRNA and natural killer T cell ligand” The
31st Transfusion Medical Conference (Hayama, Japan)
January, 2017
Fujii S. “Development of multifunctional cancer vaccine,
artificial adjuvant vector cells (aAVC)” The 2nd RIKEN
symposium “Engineering Research at RIKEN” (Wako,
Japan) November, 2016
Fujii S. “Development of cancer vaccine using innate
immunity” Seminar in Graduate School of Medicine
School of Medicine, Chiba University. (Chiba, Japan)
August, 2016
Fujii S. “Development of multifunctional cancer vaccine,
artificial adjuvant vector cells (aAVC)” The 20th Annual
Meeting of Japanese Association of Cancer Immunology (Osaka, Japan) July, 2016
Fujii S. “Systemic DC activation modulates the tumor
microenvironment and shapes the long-lived tumor
specific memory mediated by CD8+ T cells” RIKEN IMSJSI International Symposium on Immunology 2016
(Yokohama, Japan) June, 2016T

he aims of the laboratory are to extend basic studies for advancing immunotherapy
and translational research, from basic studies back and forth to the bedside in thefield of cancer and the control of other diseases. For this purpose, we have been focusing
on the following 5 projects related to NKT cells. The synthetic glycolipid, α-galactosylceramide (α-GalCer) as an NKT cell ligand is presented by CD1d molecules to invariant NKT
lymphocytes. When NKT cells are activated in this way, they show unique immunostimulatory features for innate immunity. First, as a basic study, we have recently identified
memory-like KLRG1+NKT cells and have been characterizing them in terms of intrinsic
and extrinsic factors (project 1). Additionally, we have been developing translational research projects to establish therapeutic strategies to generate strong antitumor immunity.
We previously reported the full maturation of DCs soon after the activation of NKT cells
in vivo. Based on this observation, we have developed and established artificial adjuvant
vector cells (aAVC) as a new type of cell-based drug delivery system composed of NKT
cell ligand and tumor-associated antigen. In this year, we found that anti-tumor CTL wererecruited to tumor sites (Figure) (project 2). Since this project has also been accepted by
the RIKEN translational program and Translational Research center program at Tokyo
University, we are making efforts in translational research toward eventual clinical trials(project 3). As project 4, we work toward the establishment of iPS-NKT cells to develop
an iPS-NKT cell transfer strategy, in a collaborative study with the RIKEN iPS-group.
In this project, our group plays a role in the preparation of primary NKT cells as thestarting material for generating NKT cell-derived iPS cells, and in analyzing the functionof the iPS-derived NKT cells. As a final project (project 5), we have been working on a
joint clinical phase I /IIa study of NKT cell therapy for early stage post-operative lungcancer patients with the National Hospital Organization (NHO). In this study, our roleis the analyses of immune cells in the trial and also, in collaboration with RIKEN IMSGenomic Medicine, SNP analysis of responder and poor-responder groups to find suitable biomarkers.

53

Lab activities

Drug Discovery
Antibody Platform Unit
Unit Leader: Toshitada Takemori

Figure: Effects of liposomal RK-141 (RK-569)
and Figure: Toward establishment of HBV entry
inhibitors in host cells
A. Binding of the pre-S1 domain to NTCP interfereswith bile acid transport by NTCP
Flag-NTCP-expressing HepG2 cells were incubated with
preS1 or preS1 isomer peptides (0-1000nM) for 60 min
at 37°C, followed by assays for bile acid uptake.
Bile acid uptake was carried out by incubating cellswith 1000nM [3H]taurocholate dissolved in sodium ion+for 15 min at 37°C. After washing, cell lysates wereprepared using 1% Triton X-100 in H2O and transferred
into liquid scintillation tubes and mixed with liquid
scintillation cocktail.
B. NTCP recognition by the pre-S1 is mediated or
modulated by two regions of NTCP, aa 84 to 87 and aa
157 to 165. Amino acids 157 to 165 may be critical for
maintaining the correct conformation of NTCP. The substitutions corresponding to murine (m) NTCP regions
abrogate HBV infection, but not bile acid uptake. NTCP
structure or its amino acid residues that are susceptible
to viral infection, but not taurocholate uptake activity,(Yan et al. 2013) could be a useful target(s) for mAbs
that selectively block viral infection with no or minimal
interference in bile salt transport.

Recent Major Publications
Miyauchi K, Sugimoto-Ishige A, Harada Y, Adachi Y,
Usami Y, Kaji T, , Hasegawa H, Watanabe T, Hijikata A,
Fukuyama S, Maemura T, Ohara O, Kawaoka Y, Takahashi Y, Takemori T, Kubo M. Protective neutralizing influenza antibody response in the absence of T follicularhelper cells. Nat Immunol 17, 1447–1458 (2016)
Kaji T, Hijikata A, Ishige A, Kitami T, Watanabe T, Ohara
O, Yanaka N, Okada M, Shimoda M, Taniguchi M,
Takemori T. CD4 memory T cells develop and acquire
functional competence by sequential cognate interactions and stepwise gene regulation. Int Immunol 28,
267–282 (2016)
Nance JP, Belanger S, Johnston RJ, Hu JK, Takemori
T, Crotty S. Bcl6 middle domain repressor function isrequired for T follicular helper cell differentiation andutilizes the coreceptor MTA3. Proc Natl Acad Sci U S A112, 13324–13329 (2015)

Invited Presentations
Takemori T. “How T and B cells cooperate to establish
immunological memory” (Overview talk) The 45th
annual meeting of Japanese Society for Immunology.
(Okinawa, Japan) December, 201654

W

e develop antibodies attractive for clinical cancer therapy and prevention of infectious diseases. We are now developing mAbs for AML stem cells and novel
entry inhibitors for the treatment of Hepatitis B virus infections. In addition, we are analyzing a mechanism of killing B lymphoma cells upon HLA-DR engagement by mAb.
Chronic Hepatitis B virus (CHBV) chronically infects approximately 250 million
people in the world. CHBV infection can lead to the development of cirrhosis and hepatocellular carcinoma (HCC). Several antiviral agents have been recommended as firstline anti-HBV drugs for excellent viral suppression with a low risk of antiviral resistance.
However, their cost and the need for essentially life-long treatment are considerable problems. Furthermore, none of these current treatments can eradicate the intracellular virus.
Therefore, there is still a medical need for an effective HBV cure.
As HBV binds via the preS1-domain of the viral L protein to the apical sodium-aciddependent bile acid transporter (NTCP) (Yan et al. J Virol 2013), NTCP is an ideal target
for HBV entry inhibition. Entry inhibition both during the acute and persistent phase of
the infection could protect naive hepatocytes from uptake, replication, and disseminationof the virus. Long-term treatment during persistent infections may lead to eventual viralclearance due to the natural or immune-mediated turnover of the infected hepatocytes.
As an entry inhibitor, a chemically synthesized lipopeptide derived from the pre-S1domain was developed and is now in a Phase2a clinical trial in Russia. However, bindingof the pre-S1 domain to NTCP interferes with bile acid transport by NTCP ( Figure A).
It has been recently suggested that human NTCP regions, aa 84 to 87 and aa 157 to
165 may be useful targets for selectively blocking viral infection with no or minimal interference with bile salt transport (Yan et al. J Virol 2014). According to this possibility,
we are currently attempting to establish mAbs against human NTCP that block viral interaction, but not bile acid uptake (Figure B). Successful development of such mAbs may
contribute to the development of novel entry inhibitors for the treatment of acute andchronic HBV infections.

YCI Laboratory for

Lab activities

Immune Regeneration
Young Chief Investigator: Tomokatsu Ikawa

Figure: Transcriptional networks during B cell
commitment(A) Gene expression profiles of 115 Transcription
factors (TFs) (33 constant, 27 early, 23 mid and 32late) which comprise the transcriptional network
during B cell commitment. Each column represents theexpression pattern of the factor shown in the node of
network (B).
(B) Inter-regulation between time phase-specific TFs.
Frequently associated interactions (detected in over
70% of ChIP-seq datasets) are represented as an edge.
Representative genes are shown by arrows.

Recent Major Publications
Ikawa T*, Masuda K, Endo TA, Endo M, Isono K, Koseki
Y, Nakagawa R, Kometani K, Takano J, Agata Y, Katsura
Y, Kurosaki T, Vidal M, Koseki H and Kawamoto H*.
Conversion of T cells to B cells by inactivation of polycomb-mediated epigenetic suppression of the B-lineage program. Genes Dev 30, 2475–2485 (2016)
Ikawa T*, Masuda K, Huijskens MJAJ, Satoh R, Kakugawa K, Agata Y, Miyai T, Germeraad WTV, Katsura Y,
Kawamoto H*. Induced developmental arrest of early
hematopoietic progenitors leads to the generation ofleukocyte stem cells. Stem Cell Reports 5, 716–727(2015)
Inoue T, Morita M, Hijikata A, Fukuda-Yuzawa Y, Adachi
S, Isono K, Ikawa T, Kawamoto H, Koseki H, Natsume
T, Fukao T, Ohara O, Yamamoto T, Kurosaki T*. CNOT3
contributes to early B cell development by controlling
Igh rearrangement and p53 mRNA stability. J Exp Med
212, 1465–1479 (2015)

Invited Presentations
Ikawa T. “Gene regulatory networks that orchestrate
T and B lymphocyte development.” Liaison Laboratory
Seminar (Kumamoto, Japan) February, 2016
Ikawa T. “Transcriptional networks that control B cellfate determination.” BMB2015 (Kobe, Japan) December, 2015

B

lymphocytes are generated from pluripotent hematopoietic stem cells (HSCs)
through a successive series of lineage restriction processes. Although many
essential transcription factors (TFs), such as Ikaros, PU.1, E2A, EBF1 and Pax5
have been implicated in regulating B lineage cell fate choice, molecular mechanisms underlying the generation of these patterns during cell fate determination
remain unexplored because of the absence of suitable experimental systems.
We have recently established an ideal system that can be used to examine gene
regulatory networks during lymphoid lineage specification from HSCs. We overexpressed Id3 protein fused with ERT2 (Estrogen receptor) protein, whose nuclear
translocation is induced by 4-hydroxytamoxifen (4-OHT), in hematopoietic progenitors and cultured them in B cell differentiation conditions. B cell differentiation of Id3-transduced cells was blocked at an early developmental stage, but thecells grew enormously and maintained multipotency in the presence of 4-OHT(Ikawa et al. Stem Cell Reports, 2015). We named these multipotent progenitors
induced leukocyte stem (iLS) cells.
This novel system enabled the analysis of a large set of regulatory moleculesthat control the generation of T and B lymphocytes. We have recently discovered
the transcriptional network operative during B lineage commitment (Figure). Thissystem can also be applied for ex vivo expansion of human hematopoietic stem/
progenitors, which will be required for immune cell therapy or transplantationof HSCs. Thus, the aims of our study are 1) from a basic science perspective, toelucidate the mechanisms that orchestrate cell fate specification, commitment anddifferentiation during lymphocyte development and 2) from a clinical medicine
perspective, to establish a novel method to expand human hematopoietic stem/
progenitors for the development of HSC transplantation as a clinical strategy.

55

YCI Laboratory for

Lab activities

Cellular Bioenergetic
Network
Young Chief Investigator: Toshimori Kitami

Figure: Searching for chemicals that modulate
mitochondrial function using the NLRP3 inflammasome as a disease pathway model
In order to identify chemicals that could activate as wellas suppress the NLRP3 inflammasome pathway, we
performed multiple layers of screens using IL-1β as areadout of the pathway activity.

Recent Major Publications
Kaji T, Hijikata A, Ishige A, Kitami T, Watanabe T, Ohara
O, Yanaka N, Okada M, Shimoda M, Taniguchi M,
Takemori T. CD4 memory T cells develop and acquire
functional competence by sequential cognate interactions and stepwise gene regulation. Int Immunol 28,
267–282 (2016)56

T

he overarching goal of our laboratory is to understand the role of cellular
metabolism in the pathogenesis of complex diseases. Research over the past
decades has shown that monogenic mutations in metabolic pathways cause a wide
variety of human diseases. However, more recent studies have highlighted the roleof cellular metabolism in the development of a wide variety of complex humandiseases. Our laboratory in particular has been studying the function of mitochondrial energy metabolism, which is associated with neurodegeneration, cardiovascular disease, type 2 diabetes, and aging. We hope to identify novel pathways
that restore or improve mitochondrial function through genetic and chemical
screens and to examine their potential therapeutic value using genetically engineered mouse models and unique chemical probes.
Towards our goal, we have begun to explore the role of mitochondria in an
innate immune pathway called the NLRP3 inflammasome, which is involved invariety of age-associated diseases. The NLRP3 inflammasome is activated by cell
physiological changes, including mitochondrial damage, although the molecular players involved have not been fully elucidated. To systematically identify key
players in this complex pathway, we performed a series of chemical screens andhave successfully identified chemicals that can activate as well as suppress the
NLRP3 inflammasome via mitochondria. Our analysis of these unique chemicals
highlighted an important role of mitochondrial energy metabolism in maintaining intracellular ion homeostasis and preventing NLRP3 activation. Moreover,
two outputs of the mitochondria, ATP and reactive oxygen species, both compete
for activation and suppression of the NLRP3 inflammasome.
In the coming year, we hope to gain more in-depth quantitative understandingof the interplay between mitochondria and the key interacting molecular players
we identified from the screen. In addition, we hope to understand the mechanism
by which suppressors of the NLRP3 inflammasome pathway alter or restore thefunction of mitochondria.

Central Facilities

Central Facilities
C

entral Facilities in IMS provide all researchers in the Centerwith access to the most advanced equipment and technologies. Central Facilities consist of four sections; the FACS Laboratory
managed by Dr. Takashi Saito, the Confocal Laboratory managed

by Dr. Takaharu Okada, the Genomics Laboratory managed by Dr.
Osamu Ohara, and the Animal Facility managed by Dr. Haruhiko
Koseki.

FACS Laboratory

The FACS Laboratory provides a range of support for flow cytometry
and cell sorting, techniques that are essential for nearly all immunological experiments. The FACS Lab has upgraded all FACS Arias andadded an Aria Fusion. In addition to FACS machines, the lab installed ImageStreamX, a device that combines flow cytometry with the visual detailof microscopy in a single platform, and upgraded CyTOF2, a mass-spectrometry-based cytometer that has the potential for analyzing more than
30 markers simultaneously with metal-labeled antibodies.
In 2016, 1611 analytical and 1963 sorting experiments were performed in the lab. For the users of the FACS machines (cell analyzers
and cell sorters), two staff members offer various services: (1) Technicalsupport and training: In 2016, the facility offered 11 technical courses (7for cell sorting and 4 for cell analysis). Courses were held at 3 different
levels, Calibur basic, Canto II and Aria basic. A total of 60 researchers
took the courses in 2016. (2) Cell sorting operation service: The FACS Lab
provides a cell sorting operation service, in which researchers can ask
an experienced operator to conduct the sorting experiment. In 2016, thelab provided 205 such operation services. Special cell sorting techniques,
such as single cell sorting, have also been performed. (3) Management/

maintenance of FACS machines: FACS machines are available for registered users 24 hours a day and reservations are accepted up to one monthin advance through an internal website. In addition to the in-house FACS
Lab staff, engineers from Becton Dickinson visit once a week to provide
maintenance and technical support.
Table: Instruments in the FACS Lab
Machine types

Machines# of machines

FACS cell analyzer

Calibur
Canto II

4
2

FACS cell sorter

Aria II
Aria III
Aria Fusion

3
2
1

Mass-cytometer

CyTOF2

1

Imaging flow cytometer

ImageStreamX

1

Confocal LaboratoryThe Confocal Laboratory provides equipment for cell and tissue imaging, and coordinates technical support. There areeight fluorescence microscopes available to researchers at IMS.
1. Inverted Leica SP5 system with visible lasers for single-photon
excitation including a 405 violet laser.
2. Inverted Leica SP2 system with visible lasers for single-photon excitation including a 405 violet laser. This microscope isequipped with a chamber system that controls CO2 concentration, temperature and humidity for live cell imaging.
3. Inverted Leica SP8 system with two femtosecond Ti:Sa lasers for
multiphoton excitation. This system is equipped with two typesof scanners (resonant and galvano) and hybrid detectors withhigh sensitivity and low background noise. One of the two Ti:Sa lasers is connected to an optical parametric oscillator (OPO)
that enables two-photon imaging by long wavelength excitation.
4. Upright Leica SP5 system with two femtosecond Ti:Sa lasers for
multiphoton excitation. This system utilizes resonant scanners
that enable high-speed acquisition of large z-stacks for live tissue
imaging.

Photo: Inverted Leica SP5 confocal (1), inverted Leica SP8
multiphoton (3), and Keyence BZ-X700 (7) microscopes

5. Inverted Olympus FV1200 system with visible lasers for single-photon excitation.
6. Inverted Nikon N-SIM/N-STORM super-resolution microscopefor dual color imaging.
7. Keyence BZ-X700 all-in-one fluorescence microscope.
8. GE Healthcare DeltaVision Elite system.

57

Central Facilities

Genomics LaboratoryWe are a technical support service lab that provides genomeand proteome-wide analysis for research groups in theCenter for Integrative Medical Sciences (IMS). We offer a variety ofservices to suit the needs of different labs. These include DNA sequencing, proteomics analysis, multiplex suspension array, cDNA/
Genomic clone distribution, and Primer/labeled probe distribution
for qRT-PCR analysis of immune cells (TABLE attached). Supplying advanced technologies on demand, we provide comprehensive
interrogation of the nucleic-acid based information in a cell at single-base resolution with the Illumina HiSeq1500 and as well as proteomic approaches using the AB SCIEX TripleTOF 5600. Using thisunbiased sequencing approach, we have interrogated: transcription
units, mapping/genome annotation, alternative splice sites, andtranscription factor binding sites. Our mass spectrometry system
will make it possible to use quantitative proteomic approaches invarious immunological studies. These technologies will help to reveal additional hidden features of the dynamic genomic and proteomic landscape that are regulated by both genetic and epigenetic
pathways in all organisms.

Table: Central services provided by the Genomics Lab in 2016
Next-generation DNA sequencing
RNA-sequencing
Chip-sequencing
Others (Exome etc)
Proteomics
Mass Spectrometry Analysis
Multiplex suspension array
Sanger DNA sequencing# of samples# of teams

1,525
444
148

29
9
7

# of samples# of teams
1
7

1
1,451
# of samples# of teams

36cm capillary
50cm capillary

7,728
4,848

19
15

cDNA clone delivery# of samples# of teams
2

14
Primer/labeled probe delivery# of samples# of teams

17

Animal FacilityW

e continue to maintain over 50,000 mice in the SPF area,and 1,500 mice in an isolated area. The SPF area also contains 550 germ-free or gnotobiotic mice in Vinyl Isolator rooms,
and in Vinyl Isolator bio-bubble rooms. The former is used byseveral IMS research groups, in particular the mucosal immunologists, and the latter is for “humanized mice”. We introduce mouselines into the SPF area via a combination of in vitro fertilization(IVF) and embryo transfer methods, and have also generated cryostocks of genetic resources for 711 lines. We also maintain relatively
large colonies of several commonly used strains such as NOD.CgPrkdcscid Il2rgtm1Wjl/SzJ (NSG) mice, Rag1 KO and Cre deleters, andprovide them to users on demand. We have also provided technicalassistance to generate knockout and transgenic mice (62 lines). In
addition, we made KO and KI mice (83 lines) using the CRISPR/
Cas system, and have created 13 lines of germ-free mice.
We generated mice to improve the efficacy of transplantationof human hematopoietic stem cells into NSG mice by better “humanizing” the host strain. For this purpose, we have introduced
large genomic fragments containing human genes encoding MHC,
cytokines, adhesion molecules, virus receptors and others into the
NSG mice. We maintain transgenic mice and knock-in mice withconfirmed expression of human genes on a C57BL/6 backgroundand have begun backcrossing these mice onto the NSG backgroundusing the speed-congenic method.

58

Photo: Breeding equipment for germ-free mice and gnotobiotic mice
Lower Left: stainless-isolator
Lower right: vinyl-isolator standard size
Upper right: vinyl-isolator half size

1

Award winners 2016

Central Facilities

Name of the awardee

Name of the award

Date of theannouncement

Masaru Taniguchi,Group Leader, Laboratory for Immune Regulation

The Order of the Sacred Treasure

May, 2016

Masayuki Amagai,
Team Leader, Laboratory for Skin Homeostasis

International Member of the National Academy of Medicine

Oct, 2016

Masayuki Amagai,
Team Leader, Laboratory for Skin Homeostasis

International Honorary Member of American Dermatological Association

Oct, 2016

Kenya Honda,
Team Leader, Laboratory for Gut Homeostasis

The 53rd Baelz Prize (1st prize)

Nov, 2016

Kenya Honda,
Team Leader, Laboratory for Gut Homeostasis

Academic Award of the Mochida Memorial Foundation(Mochida Memorial Foundation for Medical and Pharmaceutical Research)

Nov, 2016

Hiroshi Ohno,Group Director, Laborator for Intestinal Ecosystem

The 53rd Bälz Award (2nd prize)

Nov, 2016

Hiroshi Ohno,Group Director, Laborator for Intestinal Ecosystem

Hot Topics Award (Annual Meeting of Japan Society for Bioscience, Biotechnology,
and Agrochemistry)

Apr, 2016

Hiroshi Ohno,Group Director, Laborator for Intestinal Ecosystem

Momofuku Ando Award (Grand Prize)

Mar, 2016

Hidewaki Nakagawa,
Team Leader, Laboratory for Genome Sequencing Analysis

JCA (Japanese Cancer Association) -Mauvernay Award

Sep, 2016

Makoto Arita,
Team Leader, Laboratory for Metabolomics

Lands Award (Japan Society for Lipid Nutrition)

Sep, 2016

Hilde Cheroutre,
Team Leader, Laboratory for ImmuneCrosstalk

National Institutes of Health (NIH) Mucosal Immunology Studies Team (MIST)(U01) Award

Jun, 2016

Artem Lysenko, Postdoctoral Reasearcher, Piotr J. Kamola, Visiting
Researcher, Keith A. Boroevich, Technical Staff and Tatsuhiko
Tsunoda, Group Director, Laboratory for Medical Science Mathematics

Dream Challenges Certificate of Best Performance - Disease Module Identification
DREAM Challenge Sub-Challenge 2

Nov, 2016

Tetsuya Kubota,
Senior researcher, Laboratory for Metabolic homeostasis

Best Presentation Award (Japan Society for Adaptation Medicine)Dec, 2016

Yasutaka Motomura,Special Postdoctoral Researcher, Laboratory for Innate immune
systems

Best Presentation Award (The 45th Annual Meeting of the Japanese Society for
Immunology)

Dec, 2016

Eiji Miyauchi,
Visiting Scientist, Laborator for Intestinal Ecosystem

Best Presentation Award (The 44th Annual Meeting of the Japanese Society for
Immunology)

Feb, 2016

Shunichi Kosugi,
Research Scientist, Laboratory for Statistical Analysis

KAKENHI Reviewers Award

Sep, 2016

Alok Sharma,
Research Scientist, Laboratory for Medical Science Mathematics

VC's Research Award (University of the South Pacific)

Oct, 2016

Masato Akiyama,
Research Associate, Laboratory for Statistical Analysis

Charles J. Epstein Trainee Awards for Excellence in Human Genetics Research(semifinalist) (2016 American Society of Human Genetics)

July, 2016

Masahiro Nakajimo,
Research Associate, Laboratory for Bone and Joint Diseases

Young Investigator Award (Japan Society of Human Genetics)

Jun, 2016

Yuki Aoki,
Student Trainee, Laboratory for Human Disease Models

Young Investigator Award (The Japanese Society of Pediatric Hematology/
Oncology)

Dec, 2016

Yuki Aoki,
Student Trainee, Laboratory for Human Disease Models

Young Investigator Award (Japan Pediatric Society)

May, 2016

Toshimori Kitami,
Senior Scientist, YCI Laboratory for Cellular Bioenergetic Network

Poster Award (The 13th Conference of Asian Society for Mitochondrial Researchand Medicine and the 16th Conference of Japanese Society of Mitochondrial
Research and Medicine)Oct, 2016

Shinnosuke Matsueda,
Student Trainee, Laboratory for Metabolomics

Best Poster Award (Lipoquality, JSPS Scientific Research on Innovative Area)

Jul, 2016

Ryohei Aoyagi,
Junior Research Associate, Laboratory for Metabolomics

Poster Award (Lipoquality, JSPS Scientific Research on Innovative Area)

Jul, 2016

Taiga Kato,
Student Trainee, Laboratory for Metabolomics

Poster Award (Lipoquality, JSPS Scientific Research on Innovative Area)

Jul, 2016

Tomomitsu Hirota,
Research Scientist, Laboratory for Respiratory and Allergic Diseases

Kenji Mano Travel Grant (Joint Congress of APAAACI and APAPARI 2016)

Oct, 2016

Piotr J. Kamola,
Visiting Researcher, Laboratory for Medical Science Mathematics

ISCB Travel Award

Oct, 2016

59

RIKEN International Program Associate
(IPA)

I

MS accepted four international students as RIKEN International Program Associates (IPA). Under this IPA program, IMS
lab heads host international students from collaborating graduate
schools and supervise their Ph.D. program as Joint Supervisors. The
students receive a daily living allowance and housing costs for up toa maximum of three years.
The IPA students who studied at IMS in 2016 were
Krutula Nair (Graduate School of Frontier Biosciences, Osaka Uni-

Other programs

versity) from India studied in the Laboratory for Transcriptional
Regulation.
Mei Suen Kong (University of Science, Malaysia) studied in the
Laboratory for Cell Signaling.
Chanyoung Shin (Tokyo Institute of Technology) from Korea studied in the Laboratory for Inflammatory Regulation.
Sufeng Chiang (Genome and Systems Biology Degree Program,
National Taiwan University) from Taiwan studied in the Laboratory
for Integrated Cellular Systems

RIKEN Foreign Postdoctoral Researcher (FPR) ProgramT

he RIKEN Foreign Postdoctoral Researcher (FPR) program offers aspiring young foreign researchers with creative ideas andwho show promise of becoming internationally active in the future
the opportunity to pursue innovative research at RIKEN under the
direction of a RIKEN laboratory head. The FPR Program is one of
RIKEN’s initiatives to open up its facilities and resources to the forefront of global science and technology.

In 2016, two young researchers studied at IMS as RIKEN FPRs.
Michelle Kendle Maslowski studied in the Laboratory for Intestinal Ecosystem.
Ealey Nequan Kafi studied in the Laboratory for Innate Immune
Systems.

RIKEN Junior Research Associate (JRA) ProgramT

he Junior Research Associate Program was launched in 1996
to encourage young scientists with fresh ideas and youthful
enthusiasm to collaborate with, and learn from, senior scientists
with years of experience. This program provides part-time positions
at RIKEN for young researchers enrolled in university Ph.D. programs. The JRA program serves the dual purpose of fostering thedevelopment of these young scientists while also energizing RIKEN
with their innovative thinking.
This year, 24 JRA students studied in IMS.
Junichiro Takano (Laboratory for Developmental Genetics)
Yoshihiro Ito (Laboratory for Skin Homeostasis)
Eiichiro Watanabe (Laboratory for Gut Homeostasis)
Satoko Yokoyama (Laboratory for Intestinal Ecosystem)
Ryohei Aoyagi (Laboratory for Metabolomics)
Yuki Furuichi (Laboratory for Skin Homeostasis)
Tadashi Takeuchi (Laboratory for Intestinal Ecosystem)

Rintaro Ono (Laboratory for Human Disease Models)
Yujiro Yamamoto (Laboratory for Genotyping Development)
Yurina Miyajima (Laboratory for Innate Immune Systems)
Takato Kobayashi (Laboratory for Innate Immune Systems)
Takaaki Kawaguchi (Laboratory for Gut Homeostasis)
Kensuke Yamaguchi (Laboratory for Autoimmune Diseases)
Yuma Sakamoto (Laboratory for Bone and Joint Diseases)
Keiko Usui (Laboratory for Skin Homeostasis)
Natsuko Otaki (Laboratory for Innate Immune Systems)
Mamoru Ogawa (Laboratory for Metabolomics)
Hiroki Sugishita (Laboratory for Developmental Genetics)
Hiroe Tetsu (Laboratory for Innate Immune Systems)
Manabu Nagayama (Laboratory for Gut Homeostasis)
Daisuke Hisamatsu (Laboratory for Developmental Genetics)
Yoshiki Momiuchi (Laboratory for Innate Immune Systems)
Ari Morimoto (Laboratory for Skin Homeostasis)
Tomoko Yoshihama (Laboratory for Pharmacogenomics)

RIKEN Special Postdoctoral Researcher (SPDR) Program

R

IKEN’s Special Postdoctoral Researcher Program was instituted to provide young and creative scientists the opportunity to
be involved in autonomous and independent research in line withRIKEN objectives and research fields. The positions are competitive, but if selected, researchers receive salaries and research budgets (1 million yen) from RIKEN and they are able to conduct theirresearch at one of its laboratories.
This year, seven postdocs conducted their research at IMS
through the SPDR program.

60

Juan Guillermo Betancur Medina (Laboratory for Developmental
Genetics)
Takeshi Tanoue (Laboratory for Gut Homeostasis)
Yosuke Isobe (Laboratory for Metabolomics)
Yasutaka Motomura (Laboratory for Innate Immune Systems)
Eiji Miyauchi (Laboratory for Intestinal Ecosystem)
Tatsuro Naganuma (Laboratory for Metabolomics)
Alexis Vogelzang (Laboratory for Mucosal Immunity)

Part 3
Research
Projects

iPS projectI

nduced pluripotent stem (iPS) cells possess tremendous therapeutic potential in many areas, including regenerative medicine and immune therapy. We have begun an activity to apply iPS
technology to both mouse and human immunology research andto develop therapeutics. On a collaborative basis with individual
RCAI-IMS research laboratories, the core facility for iPS researchis engaged in developing efficient protocols to reprogram various
types of lymphocytes into iPS cells as well as to induce differentiation of iPS cells into a variety of lymphoid lineage cells. This activity
is partly supported by the Research Center Network for Realizationof Regenerative Medicine from the Japan Agency for Medical Research and Development (AMED) and CREST, Japan Science andTechnology Agency.

Project

This year, the facility investigated whether iPS-Vα24+iNKT cells
could exert anti-tumor activity in vivo. They demonstrated tumor-directed cytotoxicity by iPS-Vα24+iNKT cells in tumor-bearing mice and adjuvant activity to activate autologous NK cells inhumanized mice. The anti-tumor activity of iPS-Vα24+iNKT cells
in vivo validates their clinical potential in cancer immunotherapy.
This study was published in Stem Cells (34, 2852–2860 (2016)).
They also have operated an IMS Cell Manufacturing Unit (CMU) to
prepare iPS-Vα24+iNKT cells under the GMP (Good Manufacturing Practice)/GCTP (Good Gene, Cellular, and Tissue-based Products Manufacturing Practice) guidelines and have started preclinical studies on the safety of these iPS-Vα24+iNKT cells.

Reference
Yamada D, Iyoda T, Vizcardo R, et al. Efficient Regeneration of Human Vα24+
Invariant Natural Killer T Cells and Their Anti-Tumor Activity In Vivo. Stem Cells 34,
2852–2860 (2016)

Figure: Antitumor activity of iPS-Vα24+iNKT cells in a NOG mouse model(A) Imaging of NOG mice that were inoculated with K562-Luc cells. Starting five days
later, the mice were treated with 3 x 106 human iPS-Vα24+iNKT cells or PBS as control,a procedure repeated 5 times at 2 day intervals. Tumor growth was assessed by IVIS at
the indicated time points. (B) Total body flux (photons/s) for each mouse. PBS-treatedmice are indicated in black and iPS-Vα24+iNKT cell-treated mice are indicated in red.

Modeling skin diseases

A

topic dermatitis (AD) is regarded as a multifactorial, heterogeneous disease. A number of animal model studies of AD have beendeveloped to improve our understanding of the clinical implications, but
as yet there has been limited success in better treatment for AD patients.
AD develops and progresses through highly complex processes and theseunderlying pathological mechanisms are different in each patient. To
overcome the pathological complexity, we have constructed center-wide
projects and integrated analysis for multimodal data from several dermatitis model mice. Furthermore, we aim to provide “reverse translation”
research, applying patient-based findings to animal models.
We take advantage of several AD/chronic dermatitis model mice,
including Spade, SOCS3 cKO, STAT3 cKO, Tmem79 KO, and FLG KO
and analyze RNA-seq-based gene expression profiles to understandpre-symptomatic events occurring in the skin, beginning from birth tothe onset of dermatitis. Using such samples, Dr. Ohara’s group has conducted transcriptomic analysis, and Dr. Kawakami performed comparative analysis for each animal model by using these transcriptomic data.
Dr. T. Okada examines peripheral nerve activation in inflamed skin byconfocal and two-photon excitation microscopy.
In addition to animal model data analysis, we are establishing a

Figure: Each of the dermatitis model mice presented a clearly different immune status.
We performed sequential transcriptomic analysis in the skin of each dermatitis model
mouse [Spade, SOCS3 cKO, STAT3 cKO (early developed, slowly developed, healthy),
FOXP3 mut] and wild-type (WT) mice. Although all AD model mice showed visible
inflammation, scratching behavior and epidermal barrier dysfunction, parametersreflecting their immune status were very distinctive.

62

system for sharing human clinical data and AD animal model data, incollaboration with the medical innovation hub project team (MIH) andthe Dermatology Department of Keio University School of Medicine.
Dr. Amagai and Dr. Kubo have been working to compare the findings inmouse and human AD in order to understand which processes are common in AD pathogenesis in the two species. We then plan to select a suitable animal model for each variety of human AD.
There is cooperation in these studies between the experimental labs
led by Dr. Kubo, T. Okada and Amagai, and the computational analysis/modeling labs led by Dr. Ohara and MIH members.

Project

Integrated omics approach toelucidate the impact of the gut microbiomeon the pathogenesis of type 2 diabetesR

ecent findings in gut microbiome studies indicate that the lossof microbial diversity, or dysbiosis, is not the consequence
but rather the cause of various diseases including type 2 diabetes(T2D). As an IMS Center project, we are applying a comprehensive
multiple omics approach to elucidate the impact of the gut microbiome on the pathogenesis of T2D. This is a collaborative effort
with Professors Takashi Kadowaki and Tsutomu Yamazaki fromthe University of Tokyo Hospital. From RIKEN IMS, the Laboratories for Metabolic Homeostasis, Intestinal Ecosystem, Microbiome
Sciences, Metabolomics, Integrative Genomics, and Integrated Bioinformatics are mainly involved in this project. Fecal metagenomic, metatranscriptomic and metabolomic data, as well as plasma and urine metabolomic data are obtained from three groups ofvolunteers undergoing a complete medical checkup at the University of Tokyo Hospital (n=100 each): 1) no abnormal examination
outcome, 2) obesity (BMI 25), and 3) glucose intolerance (fastingblood glucose ≥ 110 mg/dl and HbA1c 6.0%). In addition, exomes

and SNPs of T2D susceptibility genes are being analyzed. The goalof the project is to identify T2D risk factors, such as certain bacteria
and/or their metabolites, by analyzing the metadata from the comprehensive multiple omics analyses, combined with clinical and genetic datasets. One hundred volunteers have already been recruited
for all groups, and data acquisition is underway.

Figure: Analytical scheme to identify microbial biomarker(s) for type 2
diabetes
Metadata sets shown in the figure are collected from the volunteers (three groups:
healthy, obese without metabolic abnormality, and those with impaired glucose tolerance; 100 each) and comprehensively analyzed to identify differential microbial and/or
microbial metabolic factors among the groups, which should be candidate biomarkers
for the pathogenesis of type 2 diabetes.

Humanized mouseTo help overcome the problem of limited availability of human samples and to understand kinetics and behavior of thehuman immune system in vivo, we have developed humanized
mice by injecting cord blood-derived hematopoietic stem/progenitor cells into immune-deficient NOD.Cg-Prkdcscid Il2rgtm1Wjl/
SzJ (NSG) newborn mice. Their immune-compromised status affects the engraftment levels of human hematopoietic and immune
cells. Through a collaborative effort between the Jackson Laborato-ry and RIKEN, we have generated several new strains of NSG micelacking mouse immunity and expressing human genes encoding,e.g, cytokines, adhesion molecules, and MHC molecules. By using
these new humanized mouse models, we can determine the differentiation capacity of human cells of interest. We are trying to correlate gene expression signatures and the in vivo fate of human cells.

Figure: Using humanized mice to identify the differentiation capacity of human cells
Human CD8+ T cells were purified from a humanized mouse.
By injecting purified human cord blood cells into immune-deficient newborn mice, we can analyze to what extent thecells engraft in multiple organs and which immune subsets
are generated. By assessing gene expression of the donor
cells together with functional characteristics of the cells theygenerate in vivo, we can identify gene expression signatures
defining the developmental capacity of the human cells.

63

NKT Cell Projects

N

KT cells have the capacity to enhance subsequent immune
responses. When an NKT cell ligand is loaded onto CD1d+cells, such as dendritic cells (DCs) or CD1d transfectants, NKT cells
can respond and produce IFN-γ. The medical innovation groups in
IMS have launched projects aimed at application of NKT cell therapy to cancer as translational research. Here we introduce four NKT
cell-related projects aimed at cancer treatment.
In the first project, we have been collaborating with 15 National Hospital Organization (NHO) hospitals on clinical therapeuticstudies using the NKT glycolipid, α-galactosylceramide (α-GalCer)
pulsed autologous DCs (DC/Gal) in a randomized phase IIa trial inearly stage lung cancer. At present, 39 patients are entered in thistrial. In our part of this study, we perform immunological analysesof the DC/Gal-treated and control cancer patients, including NKT
cell functional analyses. As a second project, IMS was selected to
become part of the research center network for realization of regenerative medicine. We have successfully established human iPS-NKT
cells and published a paper in which iPS-NKT cells were shown tohave the potential to produce more IFN-γ and to have strong antitumor effects in vivo (Figure 1). In addition, we verified that iPSNKT cells behave as an adjuvant for NK cells. This iPS-NKT cellproject has been planned as a clinical application research project.
Third, as new type of cancer vaccine, we established artificial adju-

Project

vant vector cells, which contain tumor antigen mRNA and α-GalCer, leading to activation of both innate and adaptive immunity. We
have almost completed the preclinical studies through discussions
with the Pharmaceuticals and Medical Devices Agency (PMDA).
Fourth we developed several new NKT cell ligand candidates andhave been investigating the function of one of them. We have been
examining its efficacy in preclinical studies for next generation
NKT cell ligand cancer immunotherapy. These three projects (iPSNKT, new NKT ligand and aAVC therapy) have been supported by
the Japan Agency for Medical Research and Development (AMED)
and are also supported by the RIKEN Drug Discovery and Medical
Technology Platforms (DMP).

Figure 1: iPS-Vα24+iNKT cells elicit antitumor activity in an NOG mouse
model
NOD/Shi-scid IL-2rγnull (NOG) immuno-deficient mice were inoculated with K562-Luc
cells and five days later, 3x106 iPS-Vα24+iNKT cells or PBS were given 5 times at 2 day
intervals. Tumor growth was assessed by IVIS at the indicated time points.

Linkage to RIKEN Program for Drug Discovery
and Medical Technology Platforms (DMP)I

MS collaborates with DMP to develop innovative new pharmaceuticals and medical technologies by facilitating the transfer ofbasic research within the institute. The DMP was founded in RIKEN in 2010 in order to support all phases of development of new
therapeutics, from the discovery of promising targets to the identification of potential lead compounds, such as small molecules
and antibodies, and the acquisition of intellectual property rights
to drugs and technologies that can then be brought to the development phase.
To achieve effective progress in this area, DMP established 8
Drug Discovery Basic Units, in which the types of studies being
performed are organized according to the expertise of each PI. IMS
contributes to this effort in several ways, including by setting up a

Figure: Collaboration with IMS and DMP for thedevelopment of innovative new pharmaceuticals
and medical technologies

64

facility for the development of antibody drugs, as the Drug Discovery Antibody Platform Unit. In addition, IMS now has seven collaborative programs with DMP, including Artificial adjuvant vector
cells (Shin-ichiro Fujii), Cancer therapy with iPS-derived NKT cells(Haruhiko Koseki), Cancer therapy with NKT cells (Masaru Taniguchi), Drugs for allergic diseases (Yasushi Ishii), Leukemia treatment drugs targeting leukemic stem cells (Fumihiko Ishikawa),
mAb therapy for lymphomas (Yasushi Ishii), and a novel inhibitorfor HBV infection (Daiki Miki). The preclinical study in the Artificial adjuvant vector cell project for cancer therapy has been completed and, therefore, Fujii et al. are preparing for an investigatorinitiated-clinical trial.

PGRN-RIKEN ProjectT

he U.S. NIH Pharmacogenomics Research Network (PGRN) is
a consortium of research groups funded as individual cooperative agreements with the NIH. PGRN investigators are top researchers from U.S. academic institutions and conduct studies of variationin human genes relevant to drug metabolism, pharmacokinetics
and pharmacodynamics, and the relationship of these genetic variation to drug responses. Principal investigators of the PGRN and
RIKEN Center for Genomic Medicine (now RIKEN IMS Core for
Genomic Medicine: CGM) held a series of discussions on the needto accelerate discoveries in pharmacogenomics (PGx) and launched
the Global Alliance of Pharmacogenetics (GAP) in 2008. Currently, the PGRN-RIKEN Project is conducted under a PGRN-Hub, anewly-formed resource established to enhance scientific exchange
between the PGRN and the scientific community at large.
In this international collaboration, the PGRN has been successfully assembling a very large collection of DNA samples from

Project

well-phenotyped patients receiving specific drugs and drug combinations in clinical trials conducted in the U.S. The CGM focuses onhigh-throughput genome-wide SNP scans and targeted sequencing of selected genes or regions using next generation sequencing(NGS). We also provide technological and methodological expertise to identify genetic factors associated with drug responses, riskof severe adverse drug reactions and non-response to medications.
Together, the PGRN-RIKEN Project capitalizes on these strengths
to advance discoveries in PGx research. To date, we have initiated46 collaborative projects and have 54 publications and over 52,000
DNA samples genotyped through this collaboration, which willlead to development of better and safer medications and realize the
dream of global precision medicine.

Figure: The Pharmacogenomics Research Network (PGRN)-RIKEN IMS
strategic alliance
Please visit http://www.pgrn.org/pgrn-riken.html

Collaboration with Asian institutes and SEAPharmI

t has been noticed that severe cutaneous adverse drug reactions(ADRs), including Stevens-Johnson syndrome (SJS) and toxic
epidermal necrolysis (TEN), occur at a much higher frequency inEast Asian and Southeast Asian populations, and several genetic
factors have been reported to be associated with the risk of cutaneous ADRs. In the case of the anti-epileptic drug carbamazepine,
the US FDA now recommends HLA-B*15:02 screening prior to theadministration of this drug for Han Chinese and other Asian populations with a high prevalence of HLA-B*15:02, due to the risk of
ADR associated with this allele. To tackle this problem regionally, in2012 we established the South East Asian Pharmacogenomics Research Network (SEAPharm) together with five other Asian countries (Korea, Indonesia, Malaysia, Taiwan, and Thailand). Membership has been steadily increasing, with Singapore participating in
2014, and Vietnam in 2016. The aim of the collaborative efforts is
to identify significant ADRs in the region so that we can identify
genomic biomarkers associated with their risk, information that
could lead to a reduction in severe ADRs. We are now focusing on

the identification of genomic biomarkers associated with cutaneous
ADRs induced by the anticonvulsant phenytoin and the antibiotic
co-trimoxazole, as well as with hepatic injury induced by anti-tuberculosis agents. In addition, we also aim to understand how theidentified genomic biomarkers lead to the ADRs by performing
functional studies. It is hoped that the discoveries from our collaborative efforts will identify useful biomarkers that can be used to
predict the risk of ADRs, leading to the establishment of “stratified
medicine” based on pharmacogenomic-guided drug therapeutics.

Figure: South East Asian Pharmacogenomics Research Network (SEAPharm)

65

Project

International Cancer Genome Consortium (ICGC)
Laboratory for Genome Sequencing Analysis

Laboratory for Medical Science Mathematics

The ICGC was organized to launch and coordinate a large number of research projects that have the common aim of comprehensively elucidating the genomic changes present in many
types of cancers. Its primary goals are to generate comprehensive
catalogues of genomic abnormalities in ~25,000 cases of different
cancer types and to make the data available to the entire researchcommunity with minimal restrictions. At the end of 2016, 78 cancer genome projects across 16 countries and the EU were ongoing,
and the ICGC released genomic data from 16,246 donors as Release23 (December, 2016). The RIKEN group has been involved withvirus-related liver cancer, which is one of the most common anddeadly cancers worldwide, especially in Japan and Asia. We performed whole genome sequencing (WGS) and RNA-Seq on 300
liver cancer samples and called their somatic mutations by using
our in-house pipeline (Nat Genet 2016). ICGC also launched a“pan-cancer” whole genome project (PCAWG) in 2014, in which
WGS data + RNA-Seq of 2834 donors were analyzed in uniform
pipelines within the same computational environment. Approximately 1000 researchers and technical staff are involved world-widein 16 theme working groups. We are contributing to this ambitious
project as a member of a technical working group arranging ten

Laboratory for Digestive Diseases“cloud” data centers worldwide, PI and researchers for driver gene
analysis, mutational signatures, immunogenomics, and mitochondrial genomics, as well as by providing sample data from 270 liver cancers to the PCAWG (10% contribution 270/2834), which is
the most productive within the ICGC/TCGA (Figure). In 2016, wecompleted the alignment and mutation calls by the three pipeline
approach and established and validated certain pipelines for immunogenomic profiling from WGS and RNA-Seq, such as HLA genotyping, HLA mutation, and neo-antigen prediction, which were
performed in collaboration with the group from The University ofTokyo. More than 95% of donors had their unique HLA genotype
determined from whole genome sequencing data (Figure)

Figure: Specimen count of PCAWG (“Pan-CAncer” Whole Genome project)samples in ICGC/The Cancer Genome Atlas (TCGA) and distribution of their
HLA-A genotypes
WGS data + RNA-Seq of 2834 donors were analyzed in PCAWG. RIKEN data (270 livercancer samples) were the most abundant in the PCAWG sample set. The RIKEN HLA/
immunogenomics working group determined unique HLA genotypes of more than
95% of donors of 2834 WGS of PCAWG donors.

Commissioned Research

The Biobank Japan ProjectI

n 2013, the Japanese government started a new initiative forlife sciences called Healthcare and Medical Strategy. This initiative consists of nine top priority research fields: 1) Drug development, 2) Development of medical devices, 3) Translational research, 4) Regenerative medicine, 5) Genomic medicine,6) Cancer, 7) Brain science, 8) Infectious diseases, and 9) Rarediseases. For the genomic medicine research field, in 2015 thegovernment decided to promote rapid and publically visible
clinical applications of genomic research findings in parallelwith strengthening of the basic infrastructure of human genome
research. Based on this concept, the government is planning toestablish a network of existing biobanks [Biobank Japan (BBJ),
Tohoku Medical Megabank (Tohoku MMB), and National
Center Biobank Network (NCBN)]. The government also established priority disease areas in 2015. Rare (hereditary) diseases,
cancer, dementia, infectious diseases and pharmacogenomics
were selected as the first priority disease areas because these diseases are thought to be very close to implementation of genomic
information for actual medical practice. Common multifactorialdiseases such as diabetes and cardiovascular diseases are positioned as the second priority disease area, which needs further

66

basic genomic research to precisely elucidate the relationships
between genetic variations and disease.
The Biobank Japan project was started as a MEXT-commissioned project in 2003, and the final goal of this project is theimplementation of personalized medicine. To pursue this aim,
the project constructed a large disease biobank, named Biobank
Japan, in the Institute of Medical Science, the University of Tokyo. This biobank has already collected DNA and clinical information from 253,000 patients suffering from 51 target diseases.
The IMS Core for Genomic Medicine has been working as themain infrastructure of genomic research for this project since
the beginning. Since the primary area of this project is common
multifactorial disease, we are performing large-scale genomic
research including GWAS and whole-genome sequencing-based
association studies using Biobank Japan samples. We are alsocollaborating with various international research groups using
this important resource.

Part 4
Events

Events

RIKEN
IMS
Summer
Program (RISP)2016

The IMS-JSIInternational
Symposium on
Immunology
2016

I

T

MS was delighted to again successfully organize the 11th RISP
(RIKEN IMS Summer Program). RISP began as the RCAI International Summer Program (RISP) in 2006 and has been continued by IMS, beginning 3 years ago. The aim of this activity
is to provide networking opportunities on a broad international
scale for young scientists, as well as to encourage future collaboration and postdoctoral training experiences in Japan. Due tothe broadened research activities of the new IMS, the RISP 2016
program was expanded to include topics in genomic studies andchemical biology to understand human health and diseases, inaddition to its original focus on immunology. The internshipprogram, in which the participants perform research in IMS laboratories, has also been maintained. RISP 2016 was co-organizedby the Chiba University Leading Graduate School Program.
We were pleased to welcome 31 graduate students and postdoctoral fellows from thirteen countries, together with 9 students
from Chiba University and 2 students from Tokyo University,and 1 student from Okinawa Institute of Science and Technology
Graduate University (OIST), who gathered at Yokohama from
June 10 to 17. The scientific sessions were composed of 16 lectures, including IMS PIs and other invited senior scientists fromJapan and abroad. RISP students also presented their researchin both oral and poster sessions. The RISP program ended withparticipation in a two day International Symposium on Immunology, co-organized by IMS and the Japanese Society for Immunology.
RISP2016 was again a success; we received excellent feedback
comments in the evaluation survey, and all students indicatedthat they would recommend this program to colleagues. From
the other perspective, many lecturers commented on how impressed they were with the quality of the RISP students. IMS willagain organize RISP in 2017, including broader topics as necessary to keep pace with recent developments in this rapidly moving life science field.

68

he IMS-JSI International Symposium on Immunology,
hosted by the RIKEN Center for Integrative Medical Sciences (IMS) in conjunction with the Japanese Society for Immunology (JSI), was held June 16-17 at the Pacifico Yokohama
Conference Center. The symposium, entitled “Immune homeostasis and diseases”, included 18 internationally-recognizedspeakers presenting their cutting-edge research and attracted
close to 400 participants. There were four sessions: (1) Microbiota, Fecal bacteriotherapy, (2) Cancer immunology, (3) Genome and epigenome, and (4) Cell death and inflammation. In
the “Microbiota, Fecal bacteriotherapy” session, it became clearthat microbiota changes, for example, by transfer of fecal bacteria, alter populations of macrophages as well as T cells. Theseobservations may provide clues to understand various humandiseases and provide a rational basis for their therapy. The “Cancer immunology” session updated our knowledge of checkpoint
inhibitors, newly developed tools such as chimeric antigen receptor (CAR) T cells, and cell-based therapy. The “Genome andepigenome” session broadened our view by revisiting non-coding SNP GWAS and linking it to tissue- or cell type- specificity. Identification of super-enhancers in Treg-specifying genes
provided important insight into autoimmune diseases. A combination of single-cell mRNA sequencing and CyTOF defineda unique dendritic cell population, allowing more precise lineage mapping of potential target cells for therapy. In the “Celldeath and inflammation” session, the dynamics and regulationof the plasma membrane during cell death turned out to be a
key for clearance of apoptotic cells. Dysregulation of DNA sensing impacts the immune system and can lead to autoimmune
diseases. Studies of programed necrotic cell death, called pyroptosis, are bringing new insight into inflammatory diseases. A
unique role of resident CD169+ macrophage as guards during
tissue injury in the experimental colitis model was revealed.

Events

13th
PGRN-RIKEN
Strategic
Alliance
Meeting

IMS
Advisory
Council2016

TT

he Global Alliance for Pharmacogenomics (GAP), a collaborative program between the former RIKEN Center
for Genomic Medicine (now RIKEN IMS) and the US National Institute of Health (NIH) Pharmacogenomics Research
Network (PGRN), was formed in 2008, with the objective ofidentifying the relationship between genetic variants and individual responses to drugs, including efficacy and side effects.
PGRN-RIKEN strategic alliance meetings, held alternately inJapan and the United States, allow for face-to-face discussions
about the progress of ongoing projects and future directions for
the PGRN-RIKEN collaboration.
On April 20-21, 2016, the 13th PGRN-RIKEN Strategic Alliance Meeting was held at the Hyatt at Fisherman’s Wharf, in
San Francisco, United States, hosted by PGRN. At the openingof the meeting, Prof. Kathy Giacomini, PGRN Program Leader, introduced the importance of the collaboration, noting that
PGRN-RIKEN contributed to as many as 41 out of all 150 Pharmacogenomics GWAS studies conducted by NIH from 2005 toJune 2015.
The meeting provided a valuable forum for exchanging information on ongoing collaborative activities: “Carboplatin-paclitaxel with bevacizumab (ICON-7 study) in ovarian cancer”,“Response to inhaled corticosteroids in large patients population
with asthma”, and “Functional genomic studies from two large
clinical trials involving aromatase inhibitors for breast cancer”.
From the RIKEN side, Dr. Mayumi Tamari, an IMS Team Leader, presented the genetics of allergic diseases.
In addition to these presentations, participants also exploredfour new research proposals from PGRN and RIKEN, ultimately
deciding to adopt three of them as additional collaboration projects, and then the meeting concluded successfully.

he IMS Advisory Council (AC) was held on September
28–30, 2016. The AC began with an Assembly Meeting during which overall Center Activities were described. Dr. Tadashi
Yamamoto gave an overview of the Center and this was followed
by a description of Core Projects for Genomic Medicine by Dr.
Michiaki Kubo and Representative Programs in the Core for
Homeostatic Regulation by Dr. Haruhiko Koseki. Finally Dr.
Yamamoto gave a description of IMS Future Plans. This session
was followed by Block reviews, where individual laboratories
were evaluated. Dr. Yamamoto also explained the Terms-of-reference from the new President of RIKEN, Dr. Hiroshi Matsumoto. At the end of the meeting, Dr. Max Cooper as chair of the AC
provided oral comments and advice, summarizing the discussions and opinions of the AC panel.
In general, the AC commented that IMS is a thriving, international recognized research institute, particularly in genomic
medicine and immunology. IMS PIs continue to publish seminal
papers in top quality journals, participate actively in international collaborations and are invited to prestigious international scientific meetings. There are however external and internal forces that may be disruptive of this success. First, the AC was very
concerned by the continued decrease in the IMS budget. The
second issue was the integration of genomic and biological science at IMS. These are two quite different scientific cultures, theformer thriving on team science to answer large-scale questions
and the latter on individual/small group science to investigate
basic issues that may have clinical applications. The merger ofthese two groups was expected to be difficult and there have beensome small successes. Dr. Yamamoto has inherited these two
problems and he is very committed to efforts that will increase
the IMS budget and to promoting the successful merger of the
genomic and biologic scientific cultures.

69

Events

Harvard
SummerSchool2016

RIKEN
IMS-IITU
Joint
WorkshopI

T

MS offers a summer internship program for undergraduate
students from Harvard University. In this program students
do a research internship in IMS laboratories, have basic biomedical sciences lectures by IMS PIs and attend a Japanese language
course. They also participate in the RIKEN IMS Summer Program (RISP) and the RIKEN IMS-JSI International Symposium
on Immunology. The participants receive a letter grade from
IMS and course credit from Harvard. In 2016 from June 6 to August 14, there were four students from Harvard University, Jessica Yamada, Andrew Chang, Christopher Tse, and Bovey Rao.
Ms. Yamada conducted her research project in the Laboratory for Disease Systems Modeling (Dr. Kitano), Mr. Chang in the
Laboratory for Developmental Genetics (Dr. Koseki), Mr. Tse in
the Laboratory for Skin Homeostasis (Dr. Amagai), and Mr. Raoin the YCI Laboratory for Immune Regeneration (Dr. Ikawa).
During their internships, the students had numerous discussions with IMS researchers and, at the end of the program, theygave oral presentations describing their research results.
In addition, they visited Yokohama Science Frontier HighSchool and met with students who want to become scientists.
On this occasion, they also experienced Japanese culture andlearned Sado (tea ceremony) from the local high school students.

70

he first joint workshop between RIKEN IMS and the Institute of Immunology in Tsinghua University (IITU) was
held on September 13–14, 2016. Three IMS Group Directors,
Drs. Tomohiro Kurosaki, Hiroshi Ohno and Ichiro Taniuchi visited IITU to introduce IMS and discussed possible future collaborations with researchers and students.
Three months later, on December 2–3, 2016, eleven IITU researchers visited RIKEN IMS for the second RIKEN IMS-IITU
workshop. The meeting started with the introduction of each
institution by IMS Director, Dr. Tadashi Yamamoto, and IITU
Director, Dr. Chen Dong. The introductions were followed bypresentations from 16 researchers, eight PIs from IITU and eightresearchers from IMS. The topics covered wide areas of immunology, including Th17, IL-17, Tfh, DC, SLE, rheumatology,
mucosal immunity, skin biology, metabolism, inflammation andneuroinflammatory responses.
On the second day, there were two sessions of one-on-one
discussions. Researchers had time to discuss their interests witheach other and seek further collaborations. From these discussions, three projects were launched under a collaboration grantfrom RIKEN IMS. By the end of the workshop, Directors Chen
Dong and Tadashi Yamamoto agreed to expand research collaborations in 2017 and meet again at the third joint workshop in
Beijing.

Events

The Academia Sinica
IBMS - RIKEN IMS
Joint Workshop
2016

Adjunct
Professorship
Programs

TI

he Academia Sinica IBMS - RIKEN IMS Joint Workshop,
hosted by the RIKEN Center for Integrative Medical
Sciences (IMS), was held on March 23 at the RIKEN Yokohama
Campus. Six leading scientists from the Institute of BioMedical
Sciences (IBMS) of Academia Sinica, the national academic institute of Taiwan, including Dr. Fu-Tong Liu, the current directorof IBMS and the vice president of Academia Sinica, visited IMS
and gave lectures about their cutting-edge research. Six IMS scientists also gave lectures. The topics broadly covered the fields
of immunology, infection and inflammation, ranging from viral
host modulation, lipid biology, glycoprotein biology, allergic andautoimmune diseases, microbiome and mucosal immunity, andinnate lymphoid cells. During and after the lectures, active discussions took place between both groups of scientists, not only
about the data presented in the lectures but also about potential
collaborations. This Joint Workshop was a valuable opportunity to begin fostering international cooperation in education andresearch between RIKEN IMS and the Academia Sinica IBMS.

IMS collaborates with and accepts graduate students from 8
domestic university graduate schools. There are now a total of
28 adjunct professors/associate professors in IMS (Table), and 72
students studied at IMS in 2016. On September 10th, IMS helda briefing session on adjunct graduate school programs to provide an opportunity for students to visit and talk directly withlab leaders and to consider their future directions.

Table: Joint graduate school programs
Graduate Program

Affiliated IMS Investigator

Graduate School of Medicine,
Osaka University

Takashi Saito (Visiting Professor),
Takashi Tanaka (Visiting Professor)

Department of Immunology,
Graduate School of Medicine,
Chiba University

Takashi Saito (Visiting Professor),
Haruhiko Koseki (Visiting Professor),
Hiroshi Ohno (Visiting Professor),
Ichiro Taniuchi (Visiting Professor),
Shin-ichiro Fujii (Visiting Professor),
Fumihiko Ishikawa (Visiting Professor)

School of Biomedical Science,
Tokyo Medical and Dental University

Takashi Saito (Visiting Professor)

Graduate School of Medicine,
Yokohama City University

Michiaki Kubo (Visiting Professor),
Shiro Ikegawa (Visiting Professor),
Mayumi Tamari (Visiting Professor),
Hidewaki Nakagawa (Visiting Professor), Taisei Mushiroda (Visiting Professor), Yukihide Momozawa (Visiting
Associate Professor), Yoichiro Kamatani(Visiting Associate Professor)

Graduate School of Medical Life
Science, Yokohama City University

Hiroshi Ohno (Visiting Professor),
Makoto Arita (Visiting Professor),
Takaharu Okada (Visiting Professor),
Kazuyo Moro (Visiting Professor),
Hidehiro Fukuyama (Visiting Associate
Professor)

Research Institute of Biological Sciences, Tokyo University of Science

Masato Kubo (Professor), Shohei Hori
(Visiting Associate Professor), Katsuyuki Shiroguchi (Visiting Associate
Professor)

Graduate School of Medicine,
Kyoto University

Fumihiko Ishikawa (Visiting Associate
Professor)

Graduate School of Medicine,
Keio University

Masayuki Amagai (Professor),
Kenya Honda (Professor),
Shigeo Koyasu (Visiting Professor),
Haruhiko Koseki (Visiting Professor)

71

Guest lectures 2016
Table: Guest Lectures Jan–Dec, 2016
Date

Speaker

Affiliation

Country

Title

4-Jan-16

Dr. Ruka Setoguchi

Graduate School of Medicine, Kyoto University

Japan

IL-15 reverses functionally compromised state of human memory
CD8+ T cells

8-Jan-16

Dr. Yasuo Mori

Graduate School of Engineerin, Kyoto University

Japan

Redox-Sensitive TRP channels: their Roles in Normal Physiological
Functions and Diseases

12-Jan-16

Dr. Robert Feil

Medical Institute of Bioregulation, Kyushu University

Japan

Epigenetic mechanisms in mammalian genomic imprinting

14-Jan-16

Dr. Cornelis Murre

University of California San Diego

USA

The contraction of space and time in immunity and gene regulation

19-Jan-16

Dr. Tri Glang Phan

Garvan Institute of Medical Research

Australia

Localisation and reactivation of memory T and B cells in subcapsularproliferative foci in the lymph node28-Jan-16

Prof. Jean Pieters

Biozentrum, University of Basel

SwitzerlandNaïve T cells, autoimmunity and resistance against intracellularpathogens; maintaining the balance

5-Feb-16

Dr. Masashi Maekawa

Ehime University Graduate School of MedicineJapan

A novel cholesterol biosensor, D4H, reveals mutual requirements ofphosphatidylserine (PS) and cholesterol in the plasma membrane

5-Feb-16

Prof. Mamoru Watanabe

Tokyo Medical and Dental University

Japan

Adult tissue stem cell therapy in inflammatory bowel disease

17-Feb-16

Dr. Shinya Ikematsu

Okinawa National College of Technology

Japan

Potential usage of biological resources in Okinawa for increasing
average life expectancy

17-Feb-16

Dr. Elena Ezhkova

Icahn School of Medicine at Mount Sinai

USA

Epigenetic regulation of skin development and skin stem cells: Focus
on the Polycomb complex

22-Feb-16

Dr. Antoine H.F.M. Peters

Friedrich Miescher Institute for Biomedical Research (FMI)

Switzerland

Histone variants and proteolysis during spermatogenesis

22-Feb-16

Dr. Kiyotsugu Yoshikawa

Graduate School of Medicine, Kyoto University

Japan

Targeting epithelial-mesenchymal transition from breast cancer toglioblastoma

4-Mar-16

Dr. Kazuya Iwamoto

Graduate School of Medical Sciences, Kumamoto University

Japan

Somatic mutations and epigenetic alterations in the postmortem
brains of patients with psychiatric disorders

14-Mar-16

Dr. Mikael Sigvardsson

Linkoping University

SwedenShaping up a cellular lineage: Lessons from normal and malignant
B-lymphocyte development

18-Mar-16

Dr. Tamiki Komatsuzaki

Research Institute for Electronic Science, Hokkaido University

Japan

Molecular data science perspectives for individuality covering frommolecules to cells

18-Mar-16

Dr. John R. Stanley

University of Pennsylvania School of Medicine

USA

Proteomic analysis of pemphigus autoantibodies indicates divergent
and polyclonal repertoires with changing expression over time

31-Mar-16

Dr. Hideyuki Yoshida

Harvard Medical School

USA

Transcriptional network of interferon and its disease associations-new insights from a global view-

1-Apr-16

Dr. Tomoharu Yasuda

Max Delbrück Center for Molecular Medicine

Germany

Mouse model of a fatal infectious disease and CRISPR/Cas9-mediatedgene therapy

11-Apr-16

Dr. Koichi S Kobayashi

Texas A&M Health Science Center College of Medicine

USA

NLRC5/CITA: a critical regulator of MHC class I and cancer

13-Apr-16

Dr. Yumiko Imai

Akita University Graduate School of Medicine

Japan

Novel therapeutic targets in the severe respiratory RNA virus infection
from the aspect of the virus-host nuclear interactions

14-Apr-16

Dr. Adrian Bracken

Trinity College Dublin

UK

The PRC2 complex in cellular proliferation and cancer

8-Jun-16

Dr. Jun R. Huh

University of Massachusetts Medical School

USA

Maternal IL17 pathway promotes autism-like phenotypes in offspring

9-Jun-16

Dr. Toru Suzuki

Okinawa Institute of Science and Technology Graduate University

Japan

Cell fate determination by mRNA decay in various biological processes

20-Jun-16

Dr. Takahisa Nakamura

Cincinnati Children's Hospital Medical Center

USA

Role of dsRNA pathways and miRNA-regulatory machinery in obesity

20-Jun-16

Dr. Tomoaki Hishida

Salk Institute for Biological Studies

USA

Cancer initiation and progression from Sox2+ cells

9-Aug-16

Dr. Kazuki Okuyama

Linköping University

SwedenProximity interactome analysis of transcription factors involving inhematopoietic cell fate determination

23-Aug-16

Prof. Shin-ichiro IMAI

Washington University School of Medicine

USA

Achieving productive aging in Japan: the systemic regulatory mechanism of mammalian aging and longevity and anti-aging intervention

2-Sep-16

Prof. Kenji Kabashima

Graduate School of Medicine, Kyoto University

Japan

Mechanism of skin immune responses to external stimuli andhomeostatic Ig extravasation: Proposal of inducible skin-associatedlymphoid tissue (iSALT)

26-Oct-16

Dr. Akiyoshi Uezumi

Institute for Comprehensive Medical Science, Fujita Health
University

Japan

Role of mesenchymal progenitor cells in the muscle stroma for theskeletal muscle homeostasis28-Oct-16

Dr. Nozomu Yachie

Research Center for Advanced Science and Technology (RCAST),The University of TokyoJapan

Let's break the walls of measuring cellular and molecular dynamics
using DNA barcodes

1-Nov-16

Prof. James Di Santo

Institute Pasteur

FranceTranscriptional regulation of innate lymphoid cell development andplasticity

7-Nov-16

Dr. Shinichi Tomizawa

School of Medicine Medical Course Histology and Cell Biology,
Yokohama City University

Japan

Epigenome dynamics during mammalian germ cell differentiation

7-Nov-16

Dr. Mayumi Oda

Keio University School of Medicine

Japan

Gene body epigenetic domains and gene-length play a role for cell
type-specific gene expression

7-Nov-16

Dr. Hisato Kobayashi

NODAI Genome Research Center, Tokyo University of AgricultureJapan

DNA methylome analysis in mouse and rat germ cells

7-Nov-16

Dr. Ferdinand von Meyenn

Babraham Institute

UK

Global epigenetic reprogramming: Mechanistic insights from humanand mouse ESCs and PGCLCs

11-Nov-16

Dr. Ryo Yamada

Graduate School of Medicine and Faculty of Medicine,
Kyoto University

Japan

Genetics for immune system

25-Nov-16

Dr. Katsuhiko Shirahige

Research Center for Epigenetic Disease, Institute of Molecular andCellular Biosciences, The University of TokyoJapan

How Cohesin regulates transcription?
–Human genetic diseases link cohesin to transcription machinery–

29-Nov-16

Dr. Emilia Dimitrova

University of Oxford

UK

Role of the CXXC protein Fbxl19 in regulation of gene expression

29-Nov-16

Dr. Julie Brind’Amour

University of British Columbia

CanadaImpact of maternally inherited histone modifications on DNA methylation maintenance and enhancer activation in the early embryo

1-Dec-16

Dr. Chyi-Song Hsieh

Washington University School of Medicine

USA

Interactions of the immune system with commensal bacteria

1-Dec-16

Dr. Markus R Wenk

National University of Singapore

Singapore

Natural variation of blood plasma lipids in healthy Asian individuals

1-Dec-16

Dr. Kenta Yashiro

Graduate School of Medicine, Osaka University

Japan

GFRA2, a GDNF receptor family member, identifies cardiac progenitorsand mediates cardiomyocyte differentiation via an alternative signal
pathway

15-Dec-16

Dr. Takeshi Egawa

Washington University School of Medicine

USA

Regulation of lymphocyte clonal expansion and tumor-suppression by
a c-MYC initiated transcription factor cascade72

Part 5
Data andStatistics

Publlications 2016

1 Aihara, H., Nakagawa, T., Mizusaki, H., Yoneda, M., Kato, M., Doiguchi, M., Imamura,Y., Higashi, M., Ikura, T., Hayashi, T., Kodama, Y., Oki, M., Nakayama, T., Cheung, E.,
Aburatani, H., Takayama, K. I., Koseki, H., Inoue, S., Takeshima, Y. & Ito, T. Histone H2A
T120 Phosphorylation Promotes Oncogenic Transformation via Upregulation of Cyclin
D1. Mol Cell 64, 176–188 (2016).
2 Aizawa, S., Okamoto, T., Sugiyama, Y., Kouwaki, T., Ito, A., Suzuki, T., Ono, C., Fukuhara, T., Yamamoto, M., Okochi, M., Hiraga, N., Imamura, M., Chayama, K., Suzuki, R.,
Shoji, I., Moriishi, K., Moriya, K., Koike, K. & Matsuura, Y. TRC8-dependent degradationof hepatitis C virus immature core protein regulates viral propagation and pathogenesis. Nat Commun 7, 11379 (2016).

Table: IMS Publlications Jan-Dec, 2016

JournalIF (2015)

Number ofPapers 2016

Nat Rev Drug Discov

47.1

1

Nat Rev Immunol

39.4

1

Nature

38.1

4

JAMA

37.7

1

Science34.7

2

Nat Genet

31.6

4

Nat Med

30.4

1

Cell

28.7

1

Lancet Oncol

26.5

1

Cell Stem Cell

22.4

2

Nat Immunol

19.4

6

Nat Cell Biol

18.7

1

Gastroenterology

18.2

1

Eur Heart J

15.1

1

Mol Cell

14.0

1

Mol Psychiatry

13.3

1

Trends Biochem Sci

12.8

1

J Clin Invest

12.6

1

J Allergy Clin Immunol

12.5

3

Ann Rheum Dis

12.4

2

Blood

11.8

2

Nat Commun

11.3

11

Genome Biol

11.3

1

J Exp Med

11.2

2

Biol Psychiatry

11.2

1

Am J Hum Genet

10.8

4

J Hepatol

10.6

1

Genes Dev

10.0

1

Immunol Rev

9.5

2

Proc Natl Acad Sci U S A

9.4

6

Nucleic Acids Res

9.2

2

Diabetes

8.8

1

Clin Cancer Res

8.7

2

Cancer Res

8.6

4

Elife

8.3

1

Cell Rep

7.9

2

Kidney Int

7.7

1

Clin Pharmacol Ther

7.3

4

PLoS Pathog

7.0

1

Haematologica

6.7

1

Cancer Immunol Res

6.7

1

PLoS Genet

6.7

4

Biotechnol Biofuels

6.4

1

J Neurol Neurosurg Psychiatry

6.4

1

J Infect Dis

6.3

1

Allergy

6.3

1

Diabetologia

6.2

1

Mucosal Immunol

6.1

1

Development

6.1

2

Hum Mol Genet

6.0

6

Arterioscler Thromb Vasc Biol

6.0

1

Others

183

TOTAL

289

74

3 Aleman, F., Yazaki, J., Lee, M., Takahashi, Y., Kim, A. Y., Li, Z., Kinoshita, T., Ecker, J. R.
& Schroeder, J. I. An ABA-increased interaction of the PYL6 ABA receptor with MYC2
Transcription Factor: A putative link of ABA and JA signaling. Sci Rep 6, 28941 (2016).
4 Alexandrov, L. B., Ju, Y. S., Haase, K., Van Loo, P., Martincorena, I., Nik-Zainal, S.,
Totoki, Y., Fujimoto, A., Nakagawa, H., Shibata, T., Campbell, P. J., Vineis, P., Phillips, D.
H. & Stratton, M. R. Mutational signatures associated with tobacco smoking in humancancer. Science 354, 618–622 (2016).
5 Aly, H. H., Suzuki, J., Watashi, K., Chayama, K., Hoshino, S., Hijikata, M., Kato, T. &Wakita, T. RNA Exosome Complex Regulates Stability of the Hepatitis B Virus X-mRNA
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263 Wattanapokayakit, S., Mushiroda, T., Yanai, H., Wichukchinda, N., Chuchottawon, C.,
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264 Wheeler, H. E., Gamazon, E. R., Frisina, R., Perez-Cervantes, C., El Charif, O., Mapes, B.,
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265 Yagihara, N., Watanabe, H., Barnett, P., Duboscq-Bidot, L., Thomas, A. C., Yang, P.,
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266 Yakushiji-Kaminatsui, N., Kondo, T., Endo, T. A., Koseki, Y., Kondo, K., Ohara, O., Vidal,
M. & Koseki, H. RING1 proteins contribute to early proximal-distal specification of theforelimb bud by restricting Meis2 expression. Development 143, 276–285 (2016).
267 Yamada, D., Iyoda, T., Vizcardo, R., Shimizu, K., Sato, Y., Endo, T. A., Kitahara, G.,
Okoshi, M., Kobayashi, M., Sakurai, M., Ohara, O., Taniguchi, M., Koseki, H. & Fujii, S.
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Anti-Tumor Activity In Vivo. Stem Cells 34, 2852–2860 (2016).
268 Yamada, H., Masuko, H., Yatagai, Y., Sakamoto, T., Kaneko, Y., Iijima, H., Naito, T.,
Noguchi, E., Konno, S., Nishimura, M., Hirota, T., Tamari, M. & Hizawa, N. Role of Lung
Function Genes in the Development of Asthma. PLoS One 11, e0145832 (2016).
269 Yamamoto, S., Fukuhara, T., Ono, C., Uemura, K., Kawachi, Y., Shiokawa, M., Mori,
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270 Yamasaki, S., Shimizu, K., Kometani, K., Sakurai, M., Kawamura, M. & Fujii, S. I. In
vivo dendritic cell targeting cellular vaccine induces CD4+ Tfh cell-dependent antibody
against influenza virus. Sci Rep 6, 35173 (2016).
271 Yamazumi, Y., Sasaki, O., Imamura, M., Oda, T., Ohno, Y., Shiozaki-Sato, Y., Nagai, S.,
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Koyasu, S. & Akiyama, T. The RNA Binding Protein Mex-3B Is Required for IL-33 Induction in the Development of Allergic Airway Inflammation. Cell Rep 16, 2456–2471(2016).
272 Yan, W., Dai, J., Xu, Z., Shi, D., Chen, D., Xu, X., Song, K., Yao, Y., Li, L., Ikegawa, S.,
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273 Yarwood, A., Viatte, S., Okada, Y., Plenge, R., Yamamoto, K., Barton, A., Symmons, D.,
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274 Yasuda, T., Fukada, T., Nishida, K., Nakayama, M., Matsuda, M., Miura, I., Dainichi, T.,
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83Budget, personnel and patents

IMS Budget FY2016

Personnel FY2016
JPY Million

IMS Budget FY2016

Government funding for operations

2,656

Category

NumberDirector

1

Commissioned research

896

Senior Advisor

2

External competitive funding

805

Deputy Director

2

4,357

Group Director

9Team Leader

22

Coordinator

2

Partnership-Promotion Coordinator

1

Total

Patents
There were 28 patents filed from January to December in 2016.
Patents

2016

Total

International
patents (PCT)28

16

Deputy Team Leader

3

Domestic patents(Japan)Senior Scientist

22

12

Research Scientist

39

Postdoctoral Researcher

11

Special Postdoctoral Researcher

6

Foreign Postdoctoral Researcher

2

Research Fellow

6

Senior Technical Scientist

3Technical Scientist

11

Technical Staff I

56

Technical Staff II

61

International Program Associate

3

Research Associate

9

Junior Research Associate

24Student Trainee

68

Assistant

28

Part-time Staff

24

Research Consultant

3

Senior Visiting Scientist

9

Visiting Scientist
Visiting Technical Scientist

3

Visiting Researcher

2

Senior Research Scientist

1

Temporary Staffing

7

Total

84

170

610

RIKEN Center for Integrative Medical Sciences
Organization Chart

Access to RIKEN Yokohama Campus

Local Access

RIKEN President
By BusTake the #08 bus from Platform 8 at the East Exit of
Tsurumi Station (also accessible from the West Exitof Keikyu Tsurumi Station) and get off at the RIKEN
Shidai Daigakuin Mae bus stop. The institute is acrossthe street. All buses from this platform are bound for
Fureyu.

Hiroshi Matsumoto

Director

Buses depart Tsurumi every 5-15 minutes. It takes
about 15 minutes to arrive at RIKEN Yokohama. The
fare is 220 yen.

Tadashi Yamamoto

Integrative
Medical Sciences
Planning Office

By Train
A 15-minute walk from JR Tsurumi-Ono Station (JRTsurumi Line), which is directly accessible by transferfrom JR Tsurumi Station.

Senior Advisor

Trains run about every 10 minutes during morning andevening rush hour, but less frequently at other times.

Masaru Taniguchi
Shizuo Akira

Searchable train timetables in English are available at
http://www.hyperdia.com/en/

By Taxi
Use the taxi stand at the East Exit of JR Tsurumi Stationor the West Exit of Keikyu Tsurumi Station. The trip
takes about 10 minutes and costs around 1,200 yen.

RIKEN Center for
Integrative
Medical Sciences
Advisory Council
Deputy Director
Michiaki Kubo
Haruhiko Koseki

Max Cooper (chair)Mark Lathrop (vice chair)
Hiroyuki Aburatani
Rudi Balling
Riccardo Dalla-Favera
Michel Georges
Ronald N. Germain
Hajime Karasuyama
Yutaka Kawakami
Paul W. Kincade
Bernard MalissenJohn O’Shea
Fiona Powrie
Peter Sorger
Kiyoshi Takatsu
Katsushi Tokunaga
Dale Umetsu
Arthur Weiss

From the Airport
From Haneda Airport

From Narita Airport

Route 1
Take the Keikyu Railways Airport Express* (blue kanjisign) for Yokohama and get off at Keikyu Tsurumi
Station (27-29 minutes). Airport Express trains run
every 10-15 minutes between 9:30 a.m. and 9:30 p.m.
Next, follow the Local Access directions above to get to
RIKEN Yokohama.

From Narita Airport Station take the JR Sobu Line
(Rapid Express), Airport Limousine Bus or JR Narita
Express to JR Shinagawa Station. (JR Sobu Line is themost inexpensive option and takes about 1 hour and 15
minutes). From JR Shinagawa Station take the JR Keihin
Tohoku Line (Yokohama direction) to JR Tsurumi Station(18 minutes). Next, follow the Local Access directions
above to get to RIKEN Yokohama.

Route 2
Take any train marked with a green (express), red or
dark grey kanji sign to Keikyu Kamata Station. Transfer
to the Keikyu Main Line and take a local train* toward
Yokohama until Keikyu Tsurumi Station* (12 minutes).

* A reserved seat express that requires payment of asurcharge in addition to train fare.
Searchable train timetables in English are available at
http://www.hyperdia.com/en/*Only Airport Express (blue kanji sign) and local trains(dark grey kanji sign) stop at Keikyu Tsurumi Station.
Note that Keikyu Tsurumi Station and JR Tsurumi Station
are two different railway stations and are separated bya bus rotary (the stations are about 150 meters apart).

85

http://www.ims.riken.jp/english/
RIKEN Yokohama Campus
1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama City,
Kanagawa, 230-0045, Japan
Tel : 045-503-9111
Fax : 045-503-9113
Email : yokohama@riken.jp

RIKEN IMS
Annual Report 2016

RIKEN IMS Annual Report 2016 RIKEN Center for Integrative Medical Sciences

RIKEN Center for
Integrative Medical Sciences

RIKEN IMS
Annual Report 2016
RIKEN Center for Integrative Medical Sciences

RIKEN 2017-030