RIMD
Osaka University

Research Institutefor Microbial Diseases

RIMD
Osaka University

Research Institutefor Microbial Diseases

2016-2017

Published by＞

Public Relations Committee RIMD, Osaka University
3-1 Yamadaoka, Suita, Osaka 565-0871, Japan
Tel +81-6-6877-5111
e-mail biken-pr@biken.osaka-u.ac.jp

http://www.biken.osaka-u.ac.jp/english/index.html

2016-2017

RIMD
Research Institute for Microbial Diseases

Research Institute for Microbial Diseases,
Osaka University
Research Institute for Microbial Diseases (RIMD), Osaka University is aworld's foremost institute for basic biological researches including microbiology, immunology and oncology.
We dedicate to stimulate fundamental biological researches as a MEXT
Joint usage/research center.

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Message from the Director
In 1934, the Research Institute forMicrobial Diseases (RIMD), the first
institute attached to Osaka University,
was established for the study of microbial diseases. For more than 80 years
since its foundation, the RIMD hasconcentrated on basic researches in
infectious diseases, immunology, andoncology and made significant contributions to the control of infectious
diseases through the identification of
new pathogens, the elucidation ofpathogenesis of microbes, and thedevelopment of vaccines and diagnostics based on these basic researchfindings. In addition, the RIMD has
achieved an outstanding contribution in
the progress of life sciences throughthe discovery of oncogenes and cellfusion phenomena and the elucidationof innate immune system.
The RIMD is cer tified as the Joint
Usage/ Research Center by the Ministry of Education, Culture, Sports, Science and Technology Japan (MEXT).
In addition to the collaborative
research conducting in our facilities,
the RIMD is also responsible to support for international scientific community through the provision of the bacteria stored at our Pathogenic Microbes
Repository Unit. Faculties at the RIMD
are also af f iliated to the Graduate
Schools of Medicine, Frontier Biosciences, Science, and Pharmaceutical
Scienc es at Osaka Universit y andac c epted many graduate students
worldwide for contribution to the development of human resource in the next
generations.

Contents
The RIMD produces world-leading
research on infectious diseases frombasic research to clinical applications
through tight collaborations with
BIKEN, public interest incorporated
foundation engaged in development,
production and supply of a wide varietyof vaccines based on the researchfindings in the RIMD, and the Immunology Frontier Research Center (IFReC),
established by the great efforts of 4
researchers in the RIMD to develop
new research projects designed toclarify immune responses in a spatiotemporal manner in the body.
The RIMD will continue to dedicate
our efforts in the progress of the basic
research of infectious diseases, immunology, oncology, developmental biology, and cell biology based on the past
outstanding achievements, and to
focus on the development of young
researchers either in Japan or abroad
who are highly motivated to discoverand establish new scientific paradigms
by themselves.

Organization ……………………………………………………………………………… 4
Division of Infectious Disease ………………………………………………………… 6
Department of Molecular Bacteriology
Department of Viral Infections
Department of Molecular Virology
Department of Immunoparasitology
Division of Host Defense …………………………………………………………… 14
Department of Immunoregulation
Department of Host Defense
Department of Cell Biology
Department of Immunochemistry
Division of Cellular and Molecular Biology ……………………………………… 22
Department of Molecular Microbiology
Department of Molecular Genetics
Department of Oncogene Research
Department of Signal Transduction
Department of Cellular Regulation
Research Center for Infectious Disease Control ……………………………… 32
Department of Bacterial Infections
Department of Molecular Protozoology
International Research Center for Infectious Diseases ……………………… 36
Laboratory of Clinical Research on Infectious Diseases
Laboratory of Combined Research on Microbiology and Immunology
Laboratory of Viral Replication
Pathogenic Microbes Repository Unit
Genome Information Research Center …………………………………………… 40

Yoshiharu Matsuura D.V.M., Ph.D.
Director
Research Institute for Microbial Diseases
Osaka University

Department of Experimental Genome Research
Department of Infection Metagenomics
Research Collaboration Center for Infectious Diseases …………………… 46
Section of Bacterial Infections
Section of Viral Infections
Section of Bacterial Drug Resistance Research
Section of Antiviral Research
Mahidol-Osaka Center for Infectious Diseases
BIKEN Endowed Department of Dengue Vaccine Development …………… 52
BIKEN Innovative Vaccine Research Alliance Laboratories ………………… 54
Vaccine Creation Project
Mucosal Vaccine Project
Vaccine Dynamics Project
Animal Resource Center for Infectious Diseases ……………………………… 58
DNA-chip Development Center for Infectious Diseases …………………… 59
Ofﬁce of Combined Program on Microbiology and Immunology ………… 60
Common Research Facilities……………………………………………………… 62

About RIMD……………………………………………………………………………… 64
For Students and Researchers who wants to study in RIMD ……………… 70
Grad Students Studying in RIMD………………………………………………… 72

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Organization

Special Research
Facilities

Research Divisions

To overcome infectious diseases

Research Center for Infectious Disease Control
Bacterial Infections
Iida Lab

Division of Infectious Disease
Dept. ofDept. ofMolecular Bacteriology

Viral Infections

Horiguchi Lab

Shioda Lab

Dept. ofDept. ofTo explore the pathogenesis of microbes

Molecular Protozoology
Horii LabTo understand our body system from genetic information

Genome Information Research Center
Dept. ofDept. ofDept. ofDept. ofMolecular Virology

Immunoparasitology

Matsuura Lab

Yamamoto Lab

Experimental Genome Research
Ikawa LabTo explore the mechanisms that protect against microbes

Division of Host Defense
Dept. ofDept. ofMolecular
Immunology

Immunoregulation
Kinoshita Lab

Dept. ofDept. of

Host Defense

Immunochemistry

Akira Lab

Arase Lab

RI MD

Research Institute
Osaka University

for Microbial Diseases,

Infection Metagenomics
Horii LabTo develop new therapeutic approaches to infectious diseases

International Research Center for
Infectious Diseases
Lab. of Clinical Research on

Lab. of Combined Research on

Infectious Diseases
Kamitani Lab

Microbiology and Immunology
Nagai Lab

Lab. ofLab. of Pathogen Detection

Viral Replication
Kobayashi Lab

and Identiﬁcation
Nakamura Lab

Pathogenic Microbes
Repository Unit

Animal Resource Center for
Infectious Diseases
To explore regulatory mechanisms in cancer cells

DNA-chip Development Center for
Infectious Diseases

Division of Cellular and Molecular Biology
Dept. ofDept. ofDept. ofMolecular Microbiology

Molecular Genetics

Oncogene ResearchHara Lab

Nojima Lab

Okada Lab

Dept. ofDept. of

Signal Transduction

Cellular Regulation

Takakura Lab

Miki Lab

Research Institute for Microbial

thorough advanced researchestablished as a researchand the development ofcenter for microbiology,

human resources. Now, we

immunology and oncology inare also developing new1934. We have performed

research fields such as gene

outstanding researches in theseengineering, genome

field and we also contributeresearch and always

extensively to growth in the

Common Research
Facilities
CentralInstrumentation
Laboratory

Central Laboratory for
Biological Hazardous
Microbes

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Radioisotope
Laboratory

Library

Ofﬁce of Combined
Program on
Microbiology
and Immunology
Research Promotion
Group
Education Promotion
Groupbasic sciences in Japan

Diseases (RIMD) was

exploring breakthrough inbiological science.

Research
Collaboration Centers inOverseas
Research Collaboration Center for
Infectious Diseases
RCC-ERI
Domestic Labs
Mahidol-Osaka Center for
Infectious Diseases

BIKEN Endowed
Department of
Dengue Vaccine
Development

To develop novel vaccines with high safety and efﬁcacy

BIKEN Innovative Vaccine Research
Alliance Laboratories
Vaccine Creation Project
Yoshioka Lab

Mucosal Vaccine Project
Sato Lab

Vaccine Dynamics Project
Aoshi Lab

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Division of Infectious Disease

Department of
Molecular Bacteriology

Some pathogenic bacteria cause specific disease symptoms
including flaccid/spastic paralyses, paroxysmal coughing, skin
exfoliation, and osteogenesis imperfecta, besides general symptoms such as fever and inflammation. Our major questions are as
to how these specific symptoms appear in response to bacterial
infections or what kinds of bacterial virulence factors are involvedin them. We hope that we will understand the nature of bacterial
infections by answering these questions.

Prof.

Yasuhiko Horiguchi

ProfileDr. Horiguchi received his Ph.D. from
Osaka Prefecture University in 1987.
After working at the Kitasato Institute for 3 years, he has worked for
RIMD since 1990. He took his positon as Postdoc in 1990, a Research
Associate in 1992, an Associate Professor in 1998. He was appointed
current position in 2001.

To understand the mechanism of infection

Bordetella pertussis, which is one of our research subjects
is a representative pathogenic bacteria of Bordetella andcauses whooping cough. In addition to B. pertussis, B.
parapertussis and B. bronchiseptica are categorized in thegenus Bordetella. Although these pathogenic organisms
share homologous virulence factors and commonly cause
respiratory infections with characteristic coughing, their hostspecificities and the course of disease manifestation arequite different: B. pertussis is a strict human pathogen causing
the acute disease whereas B. bronchiseptica infects a wide
range of mammals and causes chronic infections. We aretrying to understand what determines host specificities anddistinct disease manifestations in Bordetella infections.
Understanding the molecular mechanism by which thebacteria cause coughing in hosts is our another goal of the
Bordetella research.
We are also studying on virulence factors of Burkholderia
pseudomallei, which causes melioidosis, an endemic
disease in Southeast Asia, northern Australia, and so on.

Analyzing the structure-function relationshipof bacterial protein toxins.
 Bacterial protein toxins cause a variety of specific symptoms manifested in bacterial infections. Many bacterial
protein toxins are essentially multifunctional biomolecules,
which travel in a host body, bind to target molecules or cells,
and modify target molecules with high specificity. Some
bacterial toxins are known as the most poisonous substances on the earth. We are analyzing the structure-function
relationship of these bacterial protein toxins to understandhow they exert such powerful toxicities on target cells andintoxicated animals. We believe that these results should
give an insight into the mechanism causing specific symptoms observed in bacterial infections.
To achieve the above-mentioned goals, we are conducting
the research work by using every experimental techniquebased on bacteriology, molecular and cellular biology,
biochemistry, medical and veterinary science.

Publication(1) The bvg-repressed gene brtA, encodingbioƒNO-CssociCted sWrHCce Cdhesin, iseZpressed dWring host inHection b[
Bordetella bronchiseptica. 0ishiMCYC 5,et CN Microbiol Immunol (2016) 60:
93-10
(2) 2oN[OorphisOs +nHNWencing 'ZpressionoH &erOonecrotic ToZin in Bordetella
bronchiseptica. 1MCdC -, et CN PLoS
ONE (201) 10:e011660

(3) 5Yine Atrophic 4hinitis %CWsed b[
Pasteurella multocida Toxin and
Bordetella &erOonecrotic ToZin*origWchi ; Curr Topics Microbiol
Immunol (2012) 361:113-29() %r[stCN strWctWre oH Clostridium
perfringens enterotoZin dispNC[sHeCtWres oH betC-pore-HorOing toZins
-itCdoMoro -, et CN J Biol Chem (2011)26:199-() %hCrCcteri\Ction oH the OeObrCnetCrgeting %1 doOCin in Pasteurella
multocida toZin -COitCni 5, et CN J Biol
Chem (2010) 2:26-
(6) Clostridium perfringens enterotoxininterCcts Yith cNCWdins viC eNectrostCtic
CttrCction -iOWrC ,, et CN J Biol Chem(2010) 2:01-

Staff
Assis. Prof.：Naoaki Shinzawa ／Postdoc：Aya Fukui ／ Grad. Student: 4

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Division of Infectious Disease

Viruses are simple organisms composed of proteins, nucleic
acids, and, in some cases, lipids. Nevertheless, they interact
with several host factors and ultimately cause disease in
humans. Our laboratory focuses on the molecular mechanisms underlying viral diseases, including human immunodeficiency virus (HIV).

Department ofViral Infections

Antiviral host factors and their application
to a cure for HIV infection
HIV does not establish a productive infection in any monkey
other than the chimpanzee. Also, the sensitivity of HIV
infection and rate of disease progression vary from individual
to individual. To date, several anti-HIV host restriction factors,
LQFOXGLQJ 75,0Į DQG 75,0&\S KDYH EHHQ LGHQWLILHG DV
responsible for these phenomena. Currently, we are trying toelucidate the molecular mechanism(s) underlying the anti-HIV
activity of these factors. We also aim to cure HIV infection by
establishing novel reproductive medicine-based strategies,
including iPS cells from HIV patients.

Prof.

Tatsuo Shioda

Analysis of HIV-1 genome RNA dimerization

The genome of retroviruses such as HIV-1 always exists as
a dimer; genome dimerization plays an important role atvarious stages of the viral life cycle, including genome
packaging and reverse transcription as well as the genome
recombination processes involved in viral diversification.
Therefore, genome dimerization is a novel target for anti-HIV
therapies. We are currently analyzing computer-assisted
structural models of HIV-1 Dimer Initiation Sequences, which
are the most important factors involved in genome dimerization, to get a more complete picture of HIV genome dimerization. We are also analyzing genome recombination in HIV-1
by constructing a novel system to measure recombination
efficiency.

Human genome analysis of HIV-associatedneurocognitive disorders

ProfileDr. Shioda obtaind his B. Sc. fromthe University of Tokyo in 1982 andhis Ph.D. form the same institution
in 1990. After working at the Instituteof Medical Science in the Universityof Tokyo and University of California San Francisco, he was appointed as Associate Professor in 1997.
He took his current position as a
Professor of RIMD in 2000.

Despite successful antiretroviral therapy, nearly a quarter of
HIV patients develop mild-to-severe neurocognitive disorders(HAND). We aim to undertake genome analysis of HAND
patients to elucidate the underlying molecular mechanisms,
thereby developing therapeutic strategies to treat/prevent thisdisease.

Publication(1) A naturally occurring single amino acidsubstitution in human T4+/¶ linMer
region affects its anti-HIV-1 activity andsusceptibility to HIV-1 infection.
0aMayama '.'., et al. AIDS Res Hum
Retroviruses. (2013) 29(6):919-24.

(3) Genome-wide association study ofHIV-related lipoatrophy in Thai patients:
Association of a DLGAP1 polymorphism with fat loss. 7ttayamaMul 5., etal. AIDS Res Hum Retroviruses. (2015)
Aug;31(8):792-6.

(2) 2roperties of human immunodeƒciency virus type 1 reverse transcriptase
recombination upon infection. 5aMuragi
5., et al. J Gen Virol. (2015) Nov;96(11):3382-8.

(4) Impact of T4I/5¶ in vivo. NaMayama'.'., et al. AIDS. (2015) Sep 10;29(14):1733-43.
(5) A Single-Nucleotide Polymorphism in

ABCC4 Is Associated with Tenofovir-Related Beta2-Microglobulinuria in
Thai Patients with HIV-1 Infection.
LiManonsaMul S., et al. PLoS One (2016)
Jan 25;11(1):e0147724.

Fig. 2.Newly identified structure of DIS.
A. Comparison of the current model(brown) with the previously proposedmodel (yellow). B. Structure in a dimericform. The newly identified model shows
more extended DIS structure possibly
allowing stronger molecular interaction.

(6)Novel mutant human immunodeƒciency virus type 1 strains with high degreeof resistance to cynomolgus macaque
TRIMCyp generated by randommutagenesis. Sultana T., et al. J Gen
Virol. (2016) Apr;97(4):963-76.

Fig. 1.Host factors involoving in early replication steps of HIV

Staff
Assoc. Prof.：Emi E. Nakayama ／Assis. Prof.：Jun-ichi Sakuragi ／ Undergrad. Student 1

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Division of Infectious Disease

Molecular biology of hepatitis viruses
Viruses “know” cells better than human beings and have
evolved to replicate in living cells. We are working to understand the molecular mechanisms underlying the interplay
between viruses and host cells through research on hepatitis
viruses, flaviviruses, and insect viruses.

Department of
Molecular Virology

Prof.

Yoshiharu Matsuura

Hepatitis C virus (HCV) infects over 170 million individuals
worldwide and is one of the most common etiologic agents ofchronic liver disease, including liver cirrhosis and hepatocellular carcinoma (HCC). Although novel innovative anti-HCV
drugs that act directly on viral proteins have achieved asustained virological response in hepatitis C patients,
drug-resistant viruses emerge easily. Therefore, host factors
necessary for HCV replication are ideal targets for the development of new therapeutics for chronic hepatitis C; suchdrugs will lessen the possibility of drug-resistant breakthrough viruses emerging because the frequency of mutation
is much lower than that in the viral genome.
Upon infection with HCV, viral RNA is directly translated into
viral proteins. Viral RNA replicates in the cytoplasm using
various host factors and organelles. Viruses replicate in living
cells, and some of them, including HCV, are pathogenic to
the host. We are focusing on trying to understand the molecular mechanisms underlying the interaction between thevirus and host by identifying the host factors involved in the
propagation and pathogenicity of HCV. We have shown thatthe HCV core protein participates not only in the assembly ofviral particles but also in the development of liver steatosis

ProfileDr. Matsuura received his PhD from
Hokkaido University in 1986 andworked at Research Institute ofDaiichi Seiyaku Co. Ltd for . He wasappointed Professor in RIMD in
2000 after working at NERC Instituteof Virology in Oxford University as
a postdoctoral fellow and at the
National Institute of Infectious Diseases as a head of Laboratory of
Hepatitis Viruses in Department of
Virology II. He serves as Director of
RIMD from 2015.

and HCC. We have also shown that host proteins, including
molecular chaperones and apolipoproteins, participate inviral replication and in the formation of infectious particles.
Novel therapeutic agents targeting the host factors crucial for
propagation and pathogenesis of HCV could be available if
we can elucidate the molecular mechanisms underlying
infection and replication of HCV. We are also working on
hepatitis B virus and Japanese encephalitis virus, also members of the Flaviviridae.

Development of baculoviral vectors
Development of viral vectors capable of safely transducing foreign genes into target cells is essential for future genetherapy. We are working on developing the insect baculovirus, Autographa californica nucleopolyhedro virus, as aversatile viral vector for gene delivery. Baculovirus is capableof entering a variety of mammalian cells and facilitates expression of foreign genes under the control of mammalianpromoters; however, the viral genome does not replicate. We
are working on developing viral vectors that have the advantages and characteristics of baculovirus.

Publication(1) TRC8-dependent degradation ofhepatitis C virus immature core protein
regulates viral propagation andpathogenesis. Aizawa S. & Okamoto T.,
et al. Nat. Commun. (2016),
doi: 10.1038/ncomms11379.

(3) Amphipathic ¶-Helices inapolipoproteins are crucial to theformation of infectious hepatitis C
virus particles. Fukuhara T., et al. PLoS
Pathogens (2014),
doi: 10.1371/journal.ppat.1004534.

(2) Lipoprotein receptors redundantly
participate in entry of hepatitis C virus
Yamamoto S. & Fukuhara T.,et al. PLoS
Pathog. (2016), doi: 10.1371/journal.
ppat.1005610.

(4) Biological and immunologicalcharacteristics of hepatitis E virus-likeparticles based on the crystal
structure. Yamashita T. & Mori Y., et al.
PNAS (2009) 106:12986-91.

(5) Critical role of PA28¸ in hepatitis C
virus-associated steatogenesis andhepatocarcinogenesis. Moriishi K., etal. PNAS, (2007) 104:1661-6.
(6) Hepatitis C virus RNA replication isregulated by FKBP8 and Hsp90.
Okamoto T., et al. EMBO J, (2006) 25:
5015-25.

Staff
Assis. Prof.：Takasuke Fukuhara ／ Assis. Prof.：Toru Okamoto ／Postdoc：Chikako Ono ／ JSPS PD ：Tomokazu Tamura ／Undergrad. Student 1 ／ Grad. Student 9

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Division of Infectious DiseaseIn our immunoparasitology laboratory, we use the apicomplexan protozoan parasite Toxoplasma gondii as a model for exploring host defense systems and pathogenesis. Our research goal
is to elucidate the molecular mechanisms underlying the interface between the host and pathogen.

Dept. of Immunoparasitology

Prof.

Masahiro Yamamoto
ProfileBorn at Kumamoto (1979). Received B. Sc. from the University ofTokyo (2001) and Ph.D. from Osaka
University (2006). Promoted to an
assistant professor (2006), an associate professor (2010) at thegraduate school of medicine in
Osaka University, an independentassociate professor (2012) at RIMD
in Osaka University. Appointed current position as a Professor of
RIMD in 2013.

Antiviral host factors and their application
to a cure for HIV infection
Toxoplasma gondii is an obligatory intracellular protozoan
pathogen that causes lethal toxoplasmosis in humans and animals. One third of the global population is thought to be infected with this pathogen, making it the “most successful parasite.”T. gondii infects virtually all nucleated cells in warm-blooded animals. The parasite forms a special membranous structure
called a “parasitophorous vacuole (PV).” The host-parasite interaction takes place through the PV. In response to T. gondii,
the host immune system produces inflammatory cytokines
such as interleukins, chemokines, and interferons. Interferon-Ȗ (IFN-Ȗ) is the most important host factor for inducing anti-T. gondii responses, which suppress and kill the parasites.
One of the main projects in our laboratory is to identify the
IFN-Ȗ-induced anti-T. gondii host defense mechanisms involved in innate and adaptive immunity. Recently, we foundthat IFN-Ȗ-inducible GTPases called GBPs are important forT. gondii PV disruption, and that their function in anti-T. gondii
responses requires autophagy proteins; this suggests an unexpected link between IFN-Ȗ-induced immunity and autophagic pathways.

On the other hand, virulent T. gondii suppress IFN-Ȗ-induced
host immunity and even manipulate host immune cells to maximize the virulence of the parasite. Another main project in ourlaboratory is to identify novel virulence mechanisms used by
T. gondii. For example, we recently showed that a T. gondii-secreting virulence factor, GRA6, directly activates the host transcription factor NFAT4 to induce chemokines and recruit neutrophils to eradicate the parasite. Thus, our laboratory is focusing on host-parasite interactions via immunoparasitologicalmechanisms.

Dense granule

Publication(1) p62 plays a speciƒc role ininterferon-¸-induced presentation of a
Toxoplasma vacuolar antigen. Lee Y., etal. Cell Rep. (2015) 13:223-33.
(2) RabGDI¶ is a negative regulator ofinterferon-¸-inducible
GTPase-dependent cell-autonomous
immunity to Toxoplasma gondii.
Ohshima J., et al. Proc Natl Acad Sci
USA. (2015) 112:E4581-90.

(3) Selective and strain-speciƒc NFAT4
activation by the Toxoplasma gondii
polymorphic dense granule protein
GRA6. Ma J.S., et al. J Exp Med. (2014)
211:2013-32.

(5) A cluster of interferon-¸-inducible p65
GTPases plays a critical role in hostdefense against Toxoplasma gondii.
Yamamoto M., et al. Immunity (2012)
37:302-13.

(4) Role of the mouse and human
autophagy proteins in IFN-¸-induced
cell-autonomous responses against
Toxoplasma gondii. Ohshima J., et al.
J Immunol. (2014) 192: 3328-35.

(6) ATF6· is a host cellular target of the
Toxoplasma gondii virulence factor
ROP18. Yamamoto M., et al. J Exp Med.
(2011) 208:1533-46.

Rhoptry

Toxoplasma gondii
Pathogenic proteins are secreted from Dense granules and Rhoptry.

Toxoplasma gondii (green) proliferating inside macrophages (red).

Staff
Assis. Prof.：Miwa Sasai ／ SA Assis.Prof.：Hironori Bando ／SA Assis.Prof.：Jisu Ma ／ Grad. Student 2

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Division of Host Defense

Glycosylphosphatidylinositol (GPI) is a glycolipid that anchors
proteins to plasma membranes. GPI-anchored proteins(GPI-APs) have various and important physiological functionsin the body. Why do proteins have this peculiar structure? Our
research goal is to elucidate the biogenesis, transport, andremodeling of GPI-APs and to understand the physiologicalsignificance of this linkage.

Dept. of Immunoregulation

Prof.

Taroh Kinoshita (SUP)

ProfileDr. Kinoshita received his Ph.D.
from Osaka University in 1981.
After working at Department ofBacteriology in Osaka University
Medical School and Department of
Pathology in New York University
School of Medicine, he was appointed Professor in RIMD in 1990
(current position). He served as a
Director of RIMD from 2003 to
2007. He concurrently works for
Osaka University Immunology Frontier Research Center as a Professor from 2007.

How are GPI-APs regulated?

GPI anchors are synthesized in the endoplasmic reticulum(ER) and attached to the C terminus of proteins during
post-translational modification. GPI-APs are transported
from the ER to the Golgi and then to the cell surface in a waythat is regulated according to the features of GPI. Recently,
we identified the enzyme that cleaves GPI anchors andshowed that GPI-APs are secreted and act on distant tissues. This result indicates that GPI anchors enable our bodysystems to regulate where and when these proteins work.
We are currently studying the molecular mechanism that
controls the functions of GPI-APs. In addition, GPI anchors
have specific carbohydrate side-chains; intriguingly, the chainvaries among cells and proteins. We are interested in thephysiological significance of this carbohydrate chain andhow it is synthesized in cells.

Molecular mechanisms underlying
GPI deficiency
We found that paroxysmal nocturnal hemoglobinuria (PNH) iscaused by somatic mutation of the X-linked PIGA gene, a genenecessary for GPI-anchor biosynthesis. A recent report showedthat atypical PNH is caused by somatic mutation of one allele of
the PIGT gene, a gene responsible for GPI-anchor attachment,in combination with a germline mutation in the other allele. Now,
we are studying how unused GPI anchors are involved in thepathogenesis of atypical PNH to try to find a cure for thisdisease.
We also identified a disease called inherited GPI deficiency(IGD), which is caused by mutation of the GPI-anchor synthesizing enzyme, PIGM. Recent whole exome sequencing analysis using a next-generation sequencer identified 14 GPI-related
gene mutations responsible for IGD.
To elucidate the molecular mechanisms underlying the pathogenesis of this disease, we developed a system to analyze GPI
biosynthesis and modification. This system contributes to IGD
research worldwide. Our aim is to elucidate how GPI anchors
are involved in IGD and to find a way to treat this disease.

Publication(1) Pathogenic variants in PIGG cause
intellectual disability with seizures andhypotonia. Makrythanasis P., et al. Am.
J. Hum. Genet. (2016) 98:615-26.
(2) Post-Golgi anterograde transport
requires GARP-dependent endosometo-TGN retrograde transport. Hirata, T.,et al. Mol. Biol. Cell (2015) 26: 3071-84.

(3) The alpha helical region in p24¸2
subunit of p24 cargo receptor is pivotal
for the recognition and transport ofglycosylphosphatidylinositol-anchoredproteins. Theiler, R., et al. J Biol. Chem.
(2014) 289:16835-43.
(4) Null mutation in PGAP1 impairs GPIanchor maturation in patients withintellectual disability andencephalopathy. Murakami, Y., et al.
PLoS Genet. (2014) 10(5):e1004320.

(5) Glycosylphosphatidylinositol (GPI)
anchor deƒciency caused by mutations
in PIGW is associated with Westsyndrome and hyperphosphatasia withmental retardation syndrome.
Chiyonobu, T., et al. J. Med. Genet.
(2014) 51:203-7.
(6) Signiƒcance of GPI-anchored protein
enrichment in lipid rafts for the control
of autoimmunity. Wang, Y., et al. J. Biol.
Chem. (2013) 288:25490-9.

Staff
Assoc. Prof.：Yusuke Maeda ／ Assoc. Prof.：Yohiko Murakami(SUP) ／SA Assis.Prof.：Noriyuki Kanzawa ／ Postdoc：Tetsuya Hirata ／Grad. Student 3

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GPI-anchor biosynthesis and the transport/remodeling of GPI-APs.

15

Division of Host Defense

Dept. of Host Defense

Innate immunity is a defense system triggered by patternrecognition receptors, which recognize various pathogens
such as bacteria, fungi, and viruses, and induce the production of inflammatory factors to trigger immune responses. To
gain comprehensive understanding of the molecular mechanisms responsible for innate immunity in vivo, our lab focuseson the following themes:Prof.

Shizuo Akira (SUP)

ProfileDr. Akira graduated Osaka University Medical School in 1997 and received his Ph.D. from the same institution in 1984. He spent theperiod from 1985 to 1987 as a Research Fellow in the Department ofMicrobiology and Immunology, California University. He returned to
the Institute for Molecular and Cellular Biology in Osaka University
where he remained until 1996. He
was appointed his current positionin 1999 after working in Hyogo College of Medicine for 3 years. He
concurrently serve as the Director
for WPI Immunology Frontier Research Center, Osaka University
from 2007.

Exploration of the relationship between
immune responses and mechanisms that
ensure mRNA stability
We have been investigating the comprehensive innate
immune response induced by pattern recognition receptors thatsense various pathogens. Recently, by studying the Toll-likereceptor (TLR)-signaling pathway we found that a novel regulatory mechanism, “mRNA stability,” controls inflammatory
responses. Regnase-1, an endoribonuclease, constitutively
degrades mRNAs encoding inflammatory cytokines. Once the
TLR pathway is activated, the synthesis of mRNAs encoding
inflammatory cytokines is promptly induced along with the degradation of Regnase-1 itself. As a result, mRNAs encoding
inflammatory cytokines are stably expressed, resulting in an ongoing inflammatory response. Thus, endogenous Regnase-1
negatively regulates the stability of mRNAs encoding inflammatory cytokines in normal immune cells. Once pathogens invade
cells, Regnase-1-mediated negative regulation is released,leading to inflammation. Moreover, TLR signals also suppress
excessive inflammation by inducing synthesis of mRNA encoding Regnase-1. Currently, we are examining the molecularmechanism underlying the activity of Regnase-1 family
member proteins and other related molecules to understandthe relationship between mRNA stability and innate immune
responses.

Uncovering the novel functions of new
macrophage subsets
Macrophages are leukocytes that eliminate invading bacteria by phagocytosis. There are two types of macrophage: M1
and M2. We identified a new type of M2 macrophage that is involved in various pathological disorders such as allergic
response and metabolic syndromes. So far, we have identified a variety of “disease-specific macrophage subsets” thatare independently associated with arteriosclerosis, autoimmune disease, cancer, and fibrosis. Currently, we are focusing on the role of these novel disease-specific macrophage
subsets in different pathologies and are attempting to define
these new macrophage subsets in detail.

Publication(1) Malt1-Induced Cleavage of Regnase-1
in CD4+ Helper T Cells Regulates
Immune Activation. Uehata T., et al.
Cell. (2013) 153(5):1036-49.

(3) Regnase-1, a Ribonuclease Involved in
the Regulation of Immune Responses.
Akira S. Cold Spring Harb Symp Quant
Biol. (2013) 78:51-60.

(2) Critical role of Trib1 in differentiation oftissue-resident M2-like macrophages.
Satoh T., et al. Nature. (2013)495(7442):524-8.

(4) The ǏB kinase complex regulates thestability of cytokine-encoding mRNA
induced by TLR-IL-1R by controllingdegradation of regnase-1. Iwasaki H.,
et al. Nat Immunol. (2011) 12(12):1167-75.

(5) The Jmjd3-Irf4 axis regulates M2
macrophage polarization and host
responses against helminth infection.
Satoh T., et al. Nat Immunol. (2010) 11(10):936-44.
(6) Zc3h12a is an RNase essential forcontrolling immune responses byregulating mRNA decay. Matsushita K.,
et al. Nature. (2009) 458(7242):118590.

Staff

Regnase-1 in CD4+ T cells suppresses autoimmune disease in mice.

One subtype of disease-specific macrophage.

Assoc. Prof.：Kazuhiko Maeda ／ Assis. Prof.：Takashi Satoh ／Postdoc：Kanako Kuniyoshi ／ Undergrad. Student 5 ／Grad. Student 10

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Division of Host Defense

For multicellular organisms, intercellular communication isan indispensable mechanism underlying homeostatic regulation of the whole organism. Cells deliver and receive
various signals via extracellular factors, including growth
factors and adhesion molecules. We are focusing on
HB-EGF, a membrane-anchored growth factor, and its role inmechanisms underlying cell growth, morphogenesis. andcancer malignancy.

Department of Cell Biology

Prof.

Eisuke Mekada

Physiological role of HB-EGF and associatedregulatory mechanisms
HB-EGF is a membrane-anchored growth factor that is
proteolytically cleaved from the cell surface to yield a form (Fig.
1). Soluble HB-EGF functions in various biological processes
involved in cell proliferation, differentiation, and inflammation.
Analyses of HB-EGF knockout (KO) mice reveal that HB-EGF
is expressed in several tissues and plays pivotal roles in many
physiological processes. In particular, we are focusing on therole of HB-EGF in mouse cardiac valve development (Fig. 2). In
this process, HB-EGF plays a role as a growth-inhibitory factor,
though it is generally considered a growth-promoting factor incancers. We are now analyzing the regulatory mechanismsthat switch HB-EGF functions between growth promotion andgrowth inhibition.

Role of HB-EGF in cancer malignancy

HB-EGF is highly expressed in ovarian cancer tissues. Usinga mouse xenograft model (Fig. 3), we found that over-expression of HB-EGF promotes tumor formation by ovarian cancer
cells, and that inhibiting its expression suppresses tumor formation. Moreover, administration of CRM197, which neutralizes
HB-EGF activity, suppresses tumor formation. Thus, factors
inhibiting HB-EGF activity may be potent anti-ovarian cancer
drugs. We are developing a new anti-ovarian cancer drug,
called BK-UM (CRM197), which targets HB-EGF. Pre-clinicaland clinical studies are underway.

ProfileDr. Mekada graduated Yamagata
University in 1974 and received his
Ph.D. from Osaka University in 1982.
After working at RIMD, Institute for
Molecular and Cellular Biology inOsaka university and Institute of Life
Science in Kurume university, he wasappointed Professor in RIMD in 2000.
He served as the Director of RIMD
from 2011 to 2015.

Fig. 1.
(Left) HB-EGF comprises an extracellulardomain, a transmembrane domain, andan intracellular domain. HBF is cleaved
and secreted from the cell surface. (Right)
HB KO mice have an enlarged heartcompared with wild-type mice.

Publication(1) Tetraspanin is required for generationof reactive oxygen species by the dual
oxidase system in Caenorhabditis
elegans. Moribe H., et al. PLos Genet.
(2012) Sep;8(9):e1002957.

(3) Heparin-binding EGF-like growth factor
is a promising target for ovarian cancer
therapy. Miyamoto S., et al. Cancer Res.
(2004) Aug 15;64(16):5720-7.

(2) HB-EGF and PDGF mediate reciprocalinteractions of carcinoma cells withcancer-associated ƒbroblasts to
support progression of uterine cervicalcancers. Murata T., et al. Cancer Res.
(2011) Nov 1;71(21):6633-42.

(4) Mice with defects in HB-EGF
ectodomain shedding show severe
developmental abnormalities.
Yamazaki S., et al. J Cell Biol. (2003)
Nov 10;163(3):469-75.
(5) Requirement of CD9 on the egg

Staff
Assoc. Prof.：Ryo Iwamoto ／plasma membrane for fertilization.
Miyado K., et al. Science (2000) Jan
14;287(5451):321-4.
(6) Heparin-binding EGF-like growth factor,
which acts as the diphtheria toxin
receptor, forms a complex withmembrane protein DRAP27/CD9,
which up-regulates functional
receptors and diphtheria toxin
sensitivity. Iwamoto R., et al. EMBO J.
(1994) May 15;13(10):2322-30.

Fig. 2.
A murine cancer model. Cancer cells canbe visualized using fluorescent proteins.

Assis. Prof.：Hiroto Mizushima

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Division of Host Defense

We have been studying interactions between pathogens andvarious paired receptors. In addition, we found that MHC class II
molecules function as molecular chaperones to transport misfolded proteins to the cell surface. Analyses of misfolded proteinstransported to the cell surface revealed that they are involved in
autoimmune diseases by acting as a target for autoantibodies.

Dept of ImmunochemistryProf.

Hisashi Arase (SUP)Interaction between immune receptors andpathogens

Misfolded proteins complexed with MHC
class II molecules trigger autoimmune disease

Immune cells express “paired” activating and inhibitory
receptors that are highly homologous. The inhibitory receptors recognize self-antigens and downregulate immune
response to the self. On the other hand, we found that someinhibitory receptors are used by pathogens for immune
evasion (Fig. 1). By contrast, we found that LILRA2, anorphan activating receptor expressed on human myeloid
cells, recognizes abnormal immunoglobulins cleaved by
microbial proteases but not normal immunoglobulins.
Because immunoglobulins are important for host defense,
their degradation is very dangerous in terms of immunity(Fig. 2). In this way, paired receptors play an important role
not only in immune regulation but also in host defense
against pathogens.

Allelic polymorphisms in MHC class II molecules arestrongly associated with susceptibility to many autoimmune
diseases. However, it is unclear how MHC class II molecules are involved in autoimmune disease susceptibility. We
found that misfolded cellular autoantigens are rescued fromprotein degradation by MHC class II molecules. Furthermore, we found that misfolded proteins complex with MHC
class II molecules and become targets for autoantibodies.
Autoantibody binding to misfolded proteins that are transported to the cell surface by MHC class II molecules
correlated strongly with susceptibility to autoimmune disease,suggesting that misfolded proteins, which normally would
not be presented to the immune system, can be targets for
autoantibodies by acting as “neo self” antigens, which are
involved in the pathogenicity of autoimmune diseases(Fig.3).

ProfileProf. Arase received M.D. from Hokkaido University School of Medicine
at 1990 and received Ph.D from Hokkaido University at 1994. Thereafter
he worked as an assistant professor
at Chiba University School of Medicine (1994), a research fellow at University of California San Francisco(2000), an associate professor at
Chiba University (2002) and an associate professor at Research Institutefor Microbial Diseases (2004). he isworking as current position from
2006.

Fig. 1. Immune evasion of herpesviruses via paired inhibitory receptors.
Inhibitory receptors play an important role in immune regulation, whereas pathogens exploit inhibitory receptors for immune evasion. Especially, we have found that inhibitory receptors are used by herpesviruses to enter into host cells.

Publication(1) LILRA2 is an innate immune sensor for
microbially cleaved immunoglobulins.
Hirayasu K., et al. Nature Microbiology.
(2016) 6:16054.
doi: 10.1038/nmicrobiol.2016.54.

(3) Autoantibodies to IgG/HLA class II
complexes are associated withrheumatoid arthritis susceptibility. Jin
H., et al. Proc. Natl. Acad. Sci. USA.
(2014) 111: 3787-92.

(5) Myelin-associated glycoprotein
mediates membrane fusion and entryof neurotropic herpesviruses. SuenagaT., et al. Proc. Natl. Acad. Sci. USA(2010) 107:866-71.

(2) Rheumatoid Rescue of Misfolded
Cellular Proteins by MHC Class II
Molecules: A New Hypothesis for
Autoimmune Diseases. Arase H. Adv.
Immunol. (2015) 129:1-23.

(4) Neutrophil inƒltration during
inflammation is regulated by PILR¶ via
modulation of integrin activation. WangJ., et al. Nat. Immunol. (2013) 14:34-40.

(6) PILR¶ is a herpes simplex virus-1 entry
co-receptor that associates withglycoprotein B. Satoh T., et al. Cell
(2008) 132:935-44.

Staff

Fig. 2. Activating paired receptors play a role in host defense against bacterial infection
Activating paired receptor, LILRA2, recognizes immunoglobulin cleaved by bacterial protease activate innate immune cells(Hirayasu et al. Nat. Microbiol. 2016).

Fig. 3. Misfolded proteins complexed with MHC class II molecules
are targets for autoantibodies.
Misfolded cellular proteins are transported to the cell surface without
being processed to peptides by associating with MHC class II moleculesin the ER. Furthermore, misfolded proteins complexed with MHC class
II molecules encoded by disease-susceptible alleles are specificallyrecognized by autoantibodies. This suggests that misfolded proteinscomplexed with MHC class II molecules are natural autoantigens for
autoantibodies, which affect susceptibility to autoimmune diseases(Arase Adv. Immunol. 2016).

Assoc. Prof.：Tadahiro Suenaga ／ Assis. Prof.：Masako Kohyama ／Postdoc：Fumiji Saito ／ Postdoc：Kazuki Kishida ／Postdoc：Wataru Nakai ／ Undergrad. Student 2 ／ Grad. Student 7

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Division of Cellular andMolecular Biology

It has become apparent that aging has a major impact on theincidence of cancers. However, the underlying mechanisms areunclear. We think that cellular senescence plays a key role. In ourlaboratory, we are aiming to understand the roles and mechanismsof cellular senescence in vivo. We believe that understanding themolecular mechanisms underlying cellular senescence in vivo will
provide valuable insight into the development of aging-associateddiseases such as cancer, and open up new possibilities for theircontrol.

Department of
Molecular Microbiology

Prof.

Eiji Hara

Cellular senescence is the state of irreversible cell cycle
arrest that can be induced by a variety of potentially oncogenic stimuli and has, therefore, long been considered tosuppress tumorigenesis. We reported that p16INK4a andp21Waf1/Cip1, both cyclin-dependent kinase inhibitors, play
crucial roles in both the onset and establishment of cellularsenescence in cell culture and in mouse models. Recently,
we generated transgenic mice expressing firefly luciferase
under the control of the p16INK4a or p21Waf1/Cip1 gene promoters. Using these senescence response reporter mice in
combination with knockout mice, we are investigating thetiming and, hence, the likely roles and mechanisms, ofcellular senescence in vivo.

ProfileDr. Hara received his Ph.D. from
Tokyo University of Science in 1993.
After working at Imperial Cancer Research Fund Laboratories, U.K. (Postdoctoral Fellow), Cancer Research
UK-Paterson Institute, U.K. (Group
Leader) and the Institute for Genome
Research, University of Tokushima(Professor), he was appointed Division Chief in the Cancer Institute,Japanese Foundation for Cancer Research in 2008. He took his current
position at RIMD from 2015.

Publication(1) Ablation of the p16INK4a tumour
suppressor reverses ageing
phenotypes of klotho mice. Sato S., etal. Nature Communications (2015)
6:7035.ǡ

(3) DNA damage signaling triggers
degradation of histone
methyltransferases through APC/CCdh1
in senescent cells. Takahashi A., et al.
Molecular Cell (2012) 45:123-31.ǡ

(5) Mitogenic signalling and the p16INK4aRb pathway cooperate to enforce
irreversible cellular senescence.
Takahashi A., et al. Nature Cell Biology(2006) 8:1291-7.

(2) Obesity-induced gut microbial
metabolite promotes liver cancer
through senescence secretome.
Yoshimoto S., et al. Nature (2013)
499:97-101.ǡ

(4) Real-time in vivo imaging of p16Ink4a
reveals cross-talk with p53. YamakoshiK., et al. Journal of Cell Biology (2009)
186:393-407.

(6) Opposing effects of Ets and Id proteinson p16INK4a expression during cellularsenescence. Ohtani N., et al. Nature(2001) 409:1067-70.

Staff
Assoc. Prof.：Sugiko Watanabe ／ Assis. Prof.：Shimpei Kawamoto ／JSPS PD：Masaki Takasugi ／ Postdoc：Masahiro Wakita ／Understanding the molecular mechanisms
underlying inflammatory diseases induced
by senescence-associated secretory
phenotypes (SASPs)

Exploring the physiological roles andmechanisms underlying cellular senescencein vivo

In addition to stable cell cycle arrest, senescent cells also
develop senescence-associated secretory phenotypes(SASPs), which contribute both positively and negatively tothe onset of inflammatory diseases such as cancer (depending on the biological context). Despite considerable progressin understanding the biological roles of SASPs, far less isknown about how they are induced.
Thus, a greater understanding of the underlying molecularmechanisms will lead to novel therapeutic strategies forvarious aging-associated diseases, including cancer.
Similar to aging, obesity is associated with cancer. However, the underlying mechanisms are not well understood.
Recently, we traced the association between obesity andincreased cancer risk to gut microbiota communities that
produce DNA-damaging bile acid. We found that DNA-damaging bile acid promotes development of obesity-associatedliver cancer by inducing SASPs in hepatic stellate cells. We
are now focusing on the potential clinical implications ofthese findings.

Fig. 1.
Cellular senescence initially inhibits proliferation of damaged cells, thereby acting as a fail-safe
mechanism. However, in the long term, senescent cells may eventually promote tumorigenesis via SASPs.

6 weeks

12 months

20 months

Fig. 2.Real-time bioluminescence imaging of p16INK4a gene expression during aging(Journal of Cell Biology 186: 393-407. 2009).

Grad. Student 2

22

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Division of Cellular andMolecular Biology

Lats group
We are studying the molecular mechanisms underlying Lats1(large tumor suppressor 1), Lats2, cyclin G, and GAK (cyclin
G-associated kinase) activity to understand how chromosomal
instability occurs in cancer cells.

Department of
Molecular Genetics

Prof.

Hiroshi Nojima
ProfileDr. Nojima received his PhD from
Tokyo University in 1979 and worked
as a postdoc at Stanford University,
School of Medicine for 3 years before
returning to Japan to be employed by
Jichi Medical School. He spent 7
years at Jichi Medical School as Assitant (1981-1982) and Lecturer(1982-1988). He was appointed as Associate Professor at RIMD in 1988
and became Professor in 1995.

Lats1 and Lats2 are pivotal serine/threonine kinases that
regulate the cell cycle checkpoint, whereas the Hippo
pathway mainly regulates organ size and cell growth. We
found that Lats1/2 kinases play pivotal roles in chromosome
stability through stringent regulation of cell proliferation andmitotic progression. For example: (A) Mouse embryonic
fibroblasts (MEFs) derived from Lats1 knockout (KO) and
Lats2 KO mice display severe mitotic defects, such as
centrosome amplification/fragmentation, abnormal chromosome segregation, and aberrant cytokinesis; (B) Lats1
regulates precise centrosome duplication and chromosome
segregation by regulating the stability of a protein phosphatase, Cdc25B; (C) MEFs from mice in which Lats1 is knocked
out by disrupting its N-terminal region (Lats1DN/DN) display
chromosome missegregation, polyploidy (an increase in thenumber of multinucleated cells), and anchorage-independent
growth, which is a hallmark of malignancy; (D) we identified
two novel Lats2-mediated signaling pathways, the ALB and
CLP pathways (Fig. 1); (E) recently, we reported that Lats1/2
phosphorylate a kinesin-like motor protein, CHO1, which
regulates cytokinesis on the centrosome; (F) the activatedform of Lats2 phosphorylates p21/CDKN1A, inducing apoptosis under conditions of severe DNA damage; and (G) weare currently analyzing the molecular mechanisms by which
Lats1/2 regulate EMT and cancer stem cells to identify new
diagnostic markers for invasive cancers.

Cyclin G (CycG1 and CycG2) group

We previously discovered that CycG1 associates with theB’Ȗ subunit of protein phosphatase 2A (PP2A) to recruit
PP2A to target proteins such as p53 and MDM2. A 29
amino acid peptide, named ELAS1 [i:lȁz wȁn], triggers
apoptosis in cancer cells by promoting de-phosphorylationof Mdm2-pT216 and p53-pS46 via inhibition of CycG1-B’Ȗ
association (Fig. 2). At present, our basic research is
focused on clinical application of ELAS1.

GAK (cyclin G-associated kinase) group

GAK plays an essential role in membrane trafficking byregulating clathrin uncoating in the cytoplasm. We discovered the following additional functions of GAK: (A) GAK
forms a complex with CycG1 and B’Ȗ and phosphorylatesB’Ȗ–pT104 to regulate PP2A activity; (B) GAK localizes atthe centrosome and nucleus to regulate centrosome maturation and chromosomal condensation/alignment, respectively; (C) GAK kinase dead knockout mice (GAK-kd-/-)exhibit neonatal lethality, with pulmonary dysfunction (Fig.
3); and (D) recently, we discovered several phosphorylation targets of GAK and found that these play importantroles in proper progression of M phase.

Fig1. Lats1/2 signal transduction pathway. Hippo
pathway to inhibit cell growth, CLP pathway to inhibit
traslation and ALB pathway to regulate chromosome
segregation and cytokinesis.

Publication(1) Lats1 suppresses centrosome
overduplication by modulating thestability of Cdc25B. Mukai S., et al. Sci
Rep (2015) 5:16173.

(3) Phosphorylation of CHO1 by Lats1/2
regulates the centrosomal activation of
LIMK1 during cytokinesis. Okamoto A.,
et al. Cell Cycle (2015) 14(10):1568-82.

(5) N-terminal truncation of Lats1 causes
abnormal cell growth control andchromosomal instability. Yabuta N., etal. J Cell Sci (2013) 126(Pt 2):508-20.

(2) ELAS1-mediated inhibition of the cyclin
G1-B ¸ interaction promotes cancer cell
apoptosis via stabilization andactivation of p53. Ohno S., et al.
Oncogene (2015) 34(49):5983-96.

(4) Lats2 phosphorylates p21/CDKN1A
after UV irradiation and regulates
apoptosis. Suzuki H., et al. J Cell Sci(2013) 126(Pt 19):4358-68.

(6) Neonatal lethality in knockout miceexpressing the kinase-dead form of thegeƒtinib target GAK is caused bypulmonary dysfunction. Tabara H., etal. PLoS One (2011) 6(10):e26034.

Staff
Assoc. Prof.：Yabuta Norikazu ／Assis. Prof.：Daisuke Okuzaki (SUP) ／Postdoc：Yoko Naito ／ Grad. Student 3

24

Fig. 2. ELAS1 targets 2 molecules

Fig. 3. GAK Kinase KO mice shows developmental failure in the lung25

Division of Cellular andMolecular Biology

Cancer develops due to accumulation of mutations within a cell,which can then become malignant through immortalization andtransformation. The malignant traits of cancer cells occur as
they evade cancer inhibitory mechanisms such as apoptosis
and senescence and acquire capacity for autonomous proliferation. In addition, cancer cells acquire invasive and metastatic
characteristics through the loss of intercellular communication
and altered cell morphology.

Department of
Oncogene ResearchProf.

Masato Okada
ProfileThe molecular mechanism underlying
p18/Ragulator and mTOR nutrient signaling

Src and cancer development

Src is a signaling molecule that localizes to the sub-membrane and was the first oncogene to be discovered. Normal
tissues retain morphology by maintaining contact between
neighboring cells via cell-cell junctions; however, cancer cells
exhibit altered morphology (shown in Fig. 1) and undergo invasion and metastasis by secreting growth factors and proteases. Our laboratory has examined the role of Src as anactivator of signaling pathways that control remodeling of thecytoskeleton, which contributes to motility by inducing morphological changes. In addition, Src is involved in cell membrane-mediated signaling pathways that promote expressionof genes encoding proteases, thereby leading to malignancy.
We aim to further elucidate the detailed molecular mechanisms by which Src affects cancer cell invasion and metastasis.

mTOR is responsible for nutrition- and growth-related signaling in cells, and is involved in various biological phenomena. Our laboratory discovered that the p18 protein acts as
an adaptor for molecules involved in regulating mTOR, andthat it plays a crucial role in activating mTOR. We will continue
our research into the molecular mechanism by which p18
regulates mTOR using protein structural analysis and by
studying the molecular interactions between other factors involved in mTOR regulation.
In addition, we are studying the molecular mechanisms underlying cancer defense in naked mole rats (NMR). NMR are
rodents that are similar to mice; however, they are unique inthat they have an exceptionally long life span (up to 10 times
that of mice). They also exhibit significant resistance to aging
and cancer. Our laboratory focuses on identifying the molecular mechanisms that allow NMR to acquire these traits.

Interestingly, unlike other oncogenes, Src harbors no mutations. We found that Src is involved in a phenomenon called“cell competition,” in which cells interact and compete witheach other, producing a “winner” based on their relative fitness. We anticipate that revealing the relationship between
Src and cell competition will increase our understanding of
the function of Src in cancer development; these studies are
ongoing.

Dr. Okada graduated Kyoto University School of Science in 1981 andreceived his Ph.D. from Osaka University in 1988. He worked as a Research Associate at the Institutefor Protein Research in Osaka University and became an Associate
professor in 1996 in the same institution. He was appointed current
position as a Professor of RIMD in
2000.

EMT

Collective
Invasion

Invasion

Publication(1) Polarization of M2 macrophages
requires Lamtor1 that integrates
cytokine and amino-acid signals.
Kimura T., et al. Nat Commun. (2016)
7:13130, doi: 10.1038/ncomms13130.

(3) p18/LAMTOR1: a late endosome/lysosome-speciƒc anchor protein forthe mTORC1/MAPK signaling pathway.
Nada S., et al. Methods Enzymol (2014)
535:249-63(2) The Rho guanine nucleotide exchange
factor ARHGEF5 promotes tumor
malignancy via epithelial-mesenchymaltransition. Komiya Y., et al. Oncognesis(2016) 5: e258

(4) The novel lipid raft adaptor p18
controls endosome dynamics byanchoring the MEK-ERK pathway tolate endosomes. Nada S., et al. EMBO
J. (2009) 28:477-89(5) The lipid raft-anchored adaptor protein
cbp controls the oncogenic potential ofc-Src. Oneyama C., et al. Mol Cell
(2008) 30:426-36(6) Transmembrane phosphoprotein Cbp
regulates the activities of Src-familytyrosine kinases. Kawabuchu M., et al.
Nature (2000) 404:999-1003

Intravasation

Fig. 1. Invasion and metastasis of cancer cells

Staff
Assoc. Prof.：Shigeyuki Nada ／ Assis. Prof.：Kentaro Kajiwara ／Postdoc：Akira Ogawa ／ Undergrad. Student 5 ／ Grad. Student 14

Fig. 2.Src activation induces morphological changes andincreases cell mobility.
26

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Division of Cellular andMolecular Biology

Tissue-specific stem cells continuously produce terminally differentiated functional cells and maintain organ integrity. Bloodvessels supply oxygen and nutrients to all tissues; tissues andorgans cannot develop without blood vessel formation. Our aim
is to elucidate the cellular and molecular mechanisms underlying
vascular formation (particularly those involving stem cells) andto develop strategies to manage patients with vascular diseases.

Department of
Signal Transduction

Prof.

Mechanism of vascular formation

Tissue homeostasis in all organs is maintained via a highly
hierarchal architecture of blood vessels, which is precisely
regulated in an organ-specific manner. We are examining
how blood vessel diversity is regulated, focusing on theprocesses of angiogenesis and blood vessel maturation.
Our recent studies clarified that arterial-venous alignment isregulated by the apelin/APJ system and is critical forthermoregulation (Kidoya, Dev Cell 2015).

Stemness and vascular niche

Nobuyuki Takakura

Stem cells localize in perivascular areas in many organs.
Cells that comprise such a vascular niche regulate the“stemness” of stem cells. In our cancer stem cell (CSC)
model based on PSF1 promoter activity, we found that
CSCs proliferate and survive in the vascular niche (Nagahama, Cancer Res 2010, Kinugasa, Stem Cells 2014). Regulation of the vascular niche is a promising approach toinhibiting tumor growth. Because blood vessels developing

ProfileDr. Takakura obtained his Ph.D in
Graduate School of Medicine, Kyoto
University in 1997. He was appointed Professor of RIMD in 2006 afterworking at Kumamoto University as
an assistant professor for 4 years
and Kanazawa University as a professor for 5 years.

E 14.5E 15.0in the tumor microenvironment are immature and abnormal,
normalization of blood vessel development must control
CSCs in the vascular niche. Vascular normalization also
improves anti-tumor immunity and drug delivery. Therefore,
we are seeking ways to normalize blood vessels withintumors.

Development of tissue regeneration methods
based on endothelial stem cells
We have identified endothelial stem cells in pre-existingblood vessels and showed their utility for vascular regeneration (Naito, EMBO J 2012). Recently, we found that suchendothelial stem cells affect the resistance of cancer cells toanti-angiogenic therapy (Naito, Cancer Res 2016). We are
examining how endothelial stem cells develop and how they
are maintained during development with a view to using thiscell population to treat vascular disease.

E 15.0Publication(1) Endothelial side population cells
contribute to tumor angiogenesis andantiangiogenic drug resistance. Naito
H., et al. Cancer Research (2016) 76(11):3200-10.

(3) Identiƒcation and characterization of aresident vascular stem/progenitor cell
population in preexisting blood vessels.
Naito H., et al. EMBO J (2012) 31(4):842-55.

(2) APJ Regulates Parallel Juxtapositional
Alignment of Arteries and Veins in theskin. Kidoya H., et al. Dev Cell (2015) 33(3):247-59.

(4) Spatial and temporal role of theapelin/APJ system in the caliber size
regulation of blood vessels during
angiogenesis. Kidoya H., et al. EMBO J(2008) 27(3):522-34.

(5) A role for hematopoietic stem cells inpromoting angiogenesis. Takakura N.,
et al. Cell (2000) 102(2):199-209.

Fig. 1.Vascular development in mouse embryos. Hierarchal architecture of blood vessels accompanied by arterial (yellow)–venous(blue) alignment. Green; endothelial cells.

(6) Critical role of the TIE2 endothelial cell
receptor in the development ofdeƒnitive hematopoiesis. Takakura N.,
et al. Immunity (1998) 9(5):677-86.

Staff
Assis. Prof.：Hiroyasu Kidoya ／ Assis. Prof.：Hisamichi Naito ／Postdoc：Weizhen Jia ／ Undergrad. Student 1 ／ Grad. Student 11

28

Fig. 2.Endothelial cells (blue) and CSCs (green) in a tumor.
CSCs localize at the perivascular area, the so called “vascular niche.”

29

Division of Cellular andMolecular Biology

Most cancers originate from epithelial cells. Normal epithelial
cells form a sheet-like tissue structure in which cells are tightly
attached to each other and to the basement membrane.
Through malignant progression, cells proliferate and expand by
invading surrounding tissues. Furthermore, cells metastasize to
distant organs via blood vessels, forming often incurable
tumors. Our aim is to elucidate the mechanism underlying thismysterious process of cancer development.

Department of
Cellular Regulation

Prof.

Hiroaki Miki
ProfileDr. Miki received his Ph.D from University of Tokyo in 1998. He was appointed Professor in RIMD afterworking at the Institute of Medical
Science, Universuty of Tokyo and at
the Institute for Protein Research,
Osaka University.

Role of PRL in malignant progression ofcancers

Functional analyses based on organoidculture of intestinal epithelia

PRL is highly expressed in malignant tumors and promotes
cancer metastasis. We discovered that PRL associates with
CNNM4, a Mg2+ transporter, and inhibits its Mg2+ transporting activity. Moreover, we also found that intestinal polyps
became malignant and invaded the surrounding muscle
tissue when CNNM4 was disrupted. At present, we areinvestigating the functional relationship between cancer
malignancy and Mg2+ dyshomeostasis caused by CNNM4
inhibition.
In normal epithelial tissues, cells are attached to eachother and collectively maintain their structure; these characteristics are disrupted in cancer tissues. Forced expressionof PRL in epithelial cells cultured on matrix gels induced amarked change in their morphology; some cells invaded
into the matrix only when PRL-expressing cells weresurrounded by non-expressing cells. These results suggestthat physical interaction between PRL-expressing cells andnon-expressing cells stimulates invasive behavior during
malignant progression. We are trying to clarify the molecular mechanism underlying this function of PRL.

A method of culturing intestinal epithelia in 3D matrix gelsthat mimic in vivo situations was recently developed; thesystem is called organoid culture. In this system, intestinal
epithelial cells differentiate and form a structure comprisinga monolayer sheet of cells. We are using this organoidculture system to investigate the role of PRL/CNNM in cellproliferation, differentiation, and cancerous transformation.
Many oncogenes and anti-oncogenes involved in regulating cell proliferation and survival have been identified. By
contrast, characteristics involved in transformation ofepithelial cells in a 3D space, which accompany architectural changes (such as invasion and metastasis) in tissues,
remain unclear. For example, how do cancer cells exit theepithelial tissue in which they are “born” and expand theirterritory by invading surrounding tissues? We are tacklingthese problems and trying to identify the mechanisms
underlying cancer development.

Publication(1) Mg2+ Extrusion from Intestinal Epitheliaby CNNM Proteins Is Essential for
Gonadogenesis via AMPK-TORC1
Signaling in Caenorhabditis elegans.
Ishii T., et al. PLoS Genet. (2016) 12(8):e1006276(2) Membrane protein CNNM4-dependent
Mg2+ efflux suppresses tumor
progression. Funato Y., et al. J Clin
Invest. (2014) 124(12):5398-410(3) Basolateral Mg2+ extrusion via CNNM4
mediates transcellular Mg2+ transport
across epithelia: a mouse model.
Yamazaki D., et al. PLoS Genet. (2013)9(12):e1003983(5) Nucleoredoxin sustains Wnt/·-cateninsignaling by retaining a pool of inactive
dishevelled protein. Funato Y., et al.
Curr Biol. (2010) 20(21):1945-52(4) Thioredoxin mediates
oxidation-dependent phosphorylationof CRMP2 and growth cone collapse.
Morinaka A., et al. Sci Signal. (2011) 4(170):ra26

Staff
Assis. Prof.：Daisuke Yamazaki ／ Assis. Prof.：Yosuke Funato ／Postdoc：Osamu Hashizume ／ Undergrad. Student 3 ／ Grad. Student 630

Fig. 1. Macroscopic images of theintestine (left) and histological images of polyps (right) in the indicatedgenetically engineered mice. CNNM4-deficient mice develop adenocarcinomas that invade the muscle
layer (arrows).

Fig. 2. Organoid culture of intestinal epithelia from the indicatedgenetically engineered mice.
CNNM4 deficiency causes morphological abnormalities.

31

Research Center for Infectious
Disease Control

Department ofBacterial Infections

In our laboratory, we are conducting research and collecting genomic
information to understand how bacterial pathogens infect the host andcause disease. In addition, by developing new pathogen detection
methods using high-throughput DNA sequencers, we aim to identify
novel pathogens and reveal the pathogenesis of unknown infectious
diseases.

Prof.

Tetsuya Iida

ProfileDr. Iida received his Ph.D. from
Osaka University in 1991. He wasappointed SA Professor at the International Research Center for Infectious Diseases in RIMD after working as a Research Associate for six
years and an Associate Professor
for seven years. He took his current
position as a Professor of RIMD in2015.

We performed whole genome sequencing of Vibrio
parahaemolyticus, a bacterium that causes acute gastroenteritis in humans, and revealed that the type III secretion
system T3SS2 is essential for pathogenicity. T3SS2 directly
injects bacterial proteins (effectors) into target host cells.
We demonstrated that injection of those effectors by T3SS2
from V. parahaemolyticus leads to inflammation of theintestinal mucosa and diarrhea. Currently, we are analyzing
the molecular mechanism by which those effectors cause
the symptoms of acute gastroenteritis.
Also, we revealed that expression of the genes encoding
T3SS2 is induced by bile. In fact, chemical substances that
adsorb and remove bile suppressed symptoms caused by
V. parahaemolyticus in animal models, suggesting thatthese substances may be new therapeutic agents for V.
parahaemolyticus infection. This is an example of “anti-virulence therapy” rather than antimicrobial therapy. This kind of

approach is expected to provide novel therapeutic strategies for various bacterial infections.
Furthermore, based on findings obtained from our researchon pathogenicity, we aim to explore the life cycle of bacterial
pathogens in their natural environments.

Development of methods to diagnosebacterial infections based on genomics
and metagenomics
Emerging and re-emerging infectious diseases cause
many problems worldwide. In many cases of such infection,
the causative agent is unknown and/or the pathogenic
mechanism is not yet clear. To identify the agents that causesuch infections, and to understand the underlying pathogenesis, we are developing a high-throughput DNA sequencing-based system to detect pathogens and analyze theirvirulence traits.

Fig. 1. The genomes of bacteria belonging to genus Vibrio
comprise two distinct circularchromosomes.

Publication(1) A repeat unit of Vibrio diarrheal T3S
effector subverts cytoskeletal actin
homeostasis via binding to interstrand
region of actin ƒlaments. Nishimura M.,
et al. Sci Rep. (2015) 5:10870.

(3) A cytotoxic type III secretion effector of
Vibrio parahaemolyticus targets vacuolar
H+-ATPase subunit c and ruptures hostcell lysosomes. Matsuda S., et al. PLoS
Pathog. (2012);8(7):e1002803.

(2) Interaction between the type III effector
VopO and GEF-H1 activates the
RhoA-ROCK pathway. Hiyoshi H., et al.
PLoS Pathog. (2015) 11(3):e1004694.

(4) VopV, an F-actin-binding type III secretion
effector, is required for Vibrio
parahaemolyticus-induced enterotoxicity.
Hiyoshi H., et al. Cell Host Microbe. (2011) 10(4):401-9. doi: 10.1016/j.chom.2011.08.014.

Staff
Assoc. Prof.：Toshio Kodama ／Assis. Prof.：Shigeaki Matsuda

32

Identifying the mechanism(s) underlying
bacterial infection and pathogenesis(5) Metagenomic diagnosis of bacterial
infections. Nakamura S., et al. Emerg
Infect Dis. (2008) 14(11):1784-6.
(6) Genome sequence of Vibrio
parahaemolyticus: a pathogenic
mechanism distinct from that of V
cholerae. Makino K., et al. Lancet.
(2003) 361(9359):743-9.

Fig. 2. Bile in the small intestine induces expression ofgenes encoding T3SS2, whichis the major virulence factor
produced by Vibrio parahaemolyticus, the causative agentof diarrhea.

33

Research Center for Infectious
Disease Control

Malaria is widespread in tropical and subtropical regions,and millions of people, particularly in Africa, remain at riskof disease and death despite substantial progress inmalaria control. No effective malaria vaccine has beendeveloped or licensed. Our laboratory is undertaking basic
research on our own candidate vaccine antigen gene andconducting clinical trials.

Department of
Molecular Protozoology

Prof.

Toshihiro Horii

ProfileDr. Horii received his Ph.D. from
Osaka University in 1981. After
working at School of Science inOsaka University as Research Associate, he was appointed Associate Professor at RIMD in 1991 andbecame Professor in 1999.

Malaria vaccine targeting SERA5

The treatment of malaria patients is completely dependent
on the efficacy of anti-malaria drugs; however, drug-resistantparasites are emerging. Although a malaria vaccine is theideal weapon against this pathogen, vaccine development ishampered by genetic polymorphisms in candidate antigen
genes.
We have been focusing on the SERA5 molecule of P.
falciparum (the malaria parasite) and developed malaria
vaccine BK-SE36 by utilizing a recombinant SERA5 protein.
SERA5 is a protein expressed by newly “born” malaria
parasites that then invade red blood cells to produce the next
generation. Epidemiological studies in malaria hyper-endemic areas showed that children with antibodies against SERA5
are resistant to symptomatic malaria infection, although suchchildren are a minority.
It was surprising that Ugandan adults that suffered numerous malaria infections did not respond to vaccination with
BK-SE36. By contrast, malaria-naïve Japanese adults
produced high levels of antibodies. However, we observed
good antibody responses in young Ugandan children thatexperienced fewer malarial infections. Indeed, we obtained
72% protective efficacy 1 year post-vaccination in a follow-up
study of 6-20 years old in the phase Ib trial. This suggeststhat our vaccine provides better protection in younger individuals. We are currently conducting Phase Ib clinical trials in
Burkina Faso in West Africa involving toddlers aged 1–5
years.

Molecular mechanism underlying the use of
antigenic genes to treat malaria
The malaria parasite develops highly sophisticated strategies to evade the human immune system. One of the mostdifficult phenomena encountered by those developing vaccines
is genetic polymorphism of vaccine candidate genes; that is,field-isolated parasites harbor different sequences from thevaccine candidate genes. Fortunately, SERA5 is highly homologous among malaria parasites worldwide. We have been
undertaking genetic analysis of SERA5 epitopes and formulating new vaccines with novel adjuvants to ensure better
efficacy.

Fig. 1. Clinical trial of the BK-SE36 malaria vaccine.
The vaccine was produced under GMP (Good Manufacturing
Procedure) conditions at the Kanonji Institute of The Research
Foundation for Microbial Diseases of Osaka University.

Publication(1) Antibody titres and boosting after naturalmalaria infection in BK-SE36 vaccine
responders during a follow-up study in
Uganda. Yagi M., et al. Sci Rep. (2016)
6:34363. doi: 10.1038/srep34363.

(3) Protective epitopes of the Plasmodium
falciparum SERA5 malaria vaccine
reside in intrinsically unstructured
N-terminal repetitive sequences. Yagi
M., et al. PLoS One (2014) 9(6):e98460.

(2) Immunogenicity and protection frommalaria infection in BK-SE36
vaccinated volunteers in Uganda is not
influenced by HLA-DRB1 alleles.
Tougan T., et al. Parasitol Int. (2016) 65(5 Pt A):455-8.

(4) Phase 1b randomized trial andfollow-up study in Uganda of theblood-stage malaria vaccine candidate
BK-SE36. Palacpac N. M., et al. PLoS
One (2013) 8(5):e64073.

(5) Plasmodium falciparum serine repeat
antigen 5 (SE36) as a malaria vaccine
candidate. Palacpac N. M., et al.
Vaccine (2011) 29(35):5837-45.
(6) Evidences of protection against
blood-stage infection of Plasmodium
falciparum by the novel protein vaccine
SE36. Horii T., et al. Parasitol Int. (2010)
59(3):380-6.

Staff
Assis. Prof.：Nobuko Arisue ／ Assis. Prof.：Takahiro Tougan ／SA Professor：Eisaku Kimura ／ SA Associate Professor：Nirianne M. Q. Palacpac ／Postdoc：Jyotheeswara Reddy Edula34

Fig. 2.
Kaplan-Meier curves for 6- to 20-year-olds at 130–365
days post-second vaccination with BK-SE36.
35

International Research Center for
Infectious Diseases

Laboratory of Clinical Researchon Infectious DIseases

Laboratory of Combined Researchon Micobiology and Immunology

SA Assoc. prof.

Wataru KamitaniInternational Research Center for
Infectious Diseases

Assoc. prof.

ProfileProfileDr. Kamitani received his Ph.D. from Osaka University in
2003. After working at RIMD for one year, he spent theperiod from 2004 to 2009 as a postdoctoral fellow at the
University of Texas Medical Branch at Galveston. He returned to the RIMD and became an Associate professor for
Global COE program. He took his current position in 2013.

Dr. Nagari received his Ph.D. from Kyoto University in 1991.
He joined Institute for Virus Research in Kyoto university
until 1992 and became a Research Associate in National
Institute of Genetics in the same year. He worked as a
Postdoctoral Associate and Associate Research Scientist
at Yale School of Medicine from 199 to 2004 before he appointed SA Associate Professor at RIMD in 2004. He took
his current position in 2011.

Hiroki Nagai

Publication(1) Nonstructural protein p39 of feline calicivirus suppresses
host innate immune response by preventing IRF-3
activation. Yumiketa Y., et al. Vet. Microbiol. (2016)
185:62-7

Publication(2) Japanese encephalitis virus core protein inhibits stressgranule formation through an interaction with Caprin-1
and facilitates viral propagation. Katoh H., et al. J. Virol.
(2013) 87(1):489-502(2) Molecular and structural analysis ofǡLegionellaǡDotI gives
insights into an inner membrane complex essential for
type IV secretion. Kuroda T., et al. Sci Repǡ(2015) 5:10912(3) Severe Acute Respiratory Syndrome Coronavirus nsp1
Facilitates Efƒcient Propagation in Cells through aSpeciƒc Translational Shutoff of Host mRNA. Tanaka T.,
et al. J. Virol. (2012) 86(20):11128-37

Coronaviruses infect many different animals, including
human, and cause them to have respiratory and gastrointestinal diseases. Newly emerged Middle East respiratory
syndrome coronavirus (MERS-CoV) and severe acute respiratory syndrome coronavirus (SARS-CoV) infect respiratory
tract and cause severe pneumonia disease. Our researchgroup focuses on these coronaviruses and studies themolecular biology and host cell-virus interaction of thesecoronaviruses.
SARS-CoV is the etiological agent of a newly-emerged
human respiratory disease that originated in southern
China in 2002 and spread worldwide in the 2003 epidemic.
After 10 years of the epidemic of SARS-CoV, novel coronavirus, MERS-CoV, has been reported in Middle East region.
MERS-CoV spreads to North America, Europe, China, andKorea. No effective treatment against MERS-CoV. Our
research group studies about non-structural protein 1(nsp1), that is one of pathogenicity factor in Coronavirus.
Nsp1 of SARS-CoV induces host protein synthesis
suppression through binding to 40S ribosome complex. The
nsp1 enhance viral replication through binding to viral RNA.
Our research group utilizes a Bacterial Artificial chromosome (BAC)-based revers genetics system for thesecoronaviruses. Our group try to understand the mechanism
36

of Coronaviruses replication and pathogenesis for development therapeutic targets against Coronaviruses using the
BAC-based revers genetics system.

(1) The Type IVB secretion system: an enigmatic chimera.
Kubori T., et al. Curr Opin Microbiolǡ(2016) 29:22-9(3) Hijacking the host proteasome for the temporaldegradation of bacterial effectors. Kubori T., et al.
Methods Mol Biolǡ(2014) 1197:141-52

Many pathogenic and endosymbiotic bacteria survive within
eukaryotic host cells. Legionella pneumophila is one such
intracellular bacteria, ubiquitously found in fresh water environments. In nature, Legionella survives and replicates within
free-living amoeba (e.g., Acanthamoeba). When inhaled byhumans, Legionella enters alveolar macrophages and establishes a replicative niche. Infections eventually cause a severe
form of pneumonia, known as Legionnaires’ disease.

role in pathogenesis/endosymbiosis; however, the underlying molecular basis remains largely unknown. We havebeen working to address the question of how intracellularbacteria like Legionella establish replicative niches inhostile environments, such as those inside eukaryotic cells.

Deciphering the molecular basis uponwhich Legionella survive within cells

Fig. Gene expression control by SARS-CoV nsp1
①Nsp1 binds to 40S ribosome, and then induces translational shutoff.
The nsp1-40S binding induces cleavage of mRNA.
②Nsp1 binds to viral mRNA, and then the interaction enhances viral
replication.

The type IV secretion system (T4SS) is a bacterial secretion system that transports biological macromolecules suchas nucleic acids and proteins and is evolutionally closely
related to bacterial conjugation systems. Legionella utilizes
Dot/Icm T4SS to deliver hundreds of bacterial “effector
proteins” into host cells, a process essential for intracellularsurvival and pathogenesis. Phylogenetic analyses of theorder Legionellales, which includes Legionella species anda zoonotic pathogen called Coxiella burnetii, demonstratethat a common ancestor acquired a T4SS on its chromosome, and that T4SSs are maintained by vertical genetransfer. Thus, T4SSs and effector proteins play a criticalFig. 1. Legionella pneumophilaFig. 2. T4SS core complex

37

International Research Center for
Infectious Diseases

International Research Center for
Infectious Diseases

Laboratory of Viral Replication

SA Assoc. prof.

Takeshi Kobayashi

ProfileDr. Kobayashi received his Ph.D. from Osaka University
in 2000. He became a Research associate at RIMD in
the same year. He spent for five years as a postdoctoral
fellow in Vanderbilt University, USA before returning toJapan to work at the Institute for Virus Research in Kyoto
University. He was appointed SA Associate Professor of
RIMD in 2012.

The facility is part of the National BioResource Project(NBRP) directed by the Ministry of Education, Culture, Sports,
Science and Technology, Japan, and is a member of theJapan Society for Culture Collections (JSCC); therefore, wecollect and preserve pathogenic bacterial strains. Thesestrains are distributed to investigators in and outside thiscountry upon request. Our collection is listed on the website
for this facility (http://rceid.biken.osaka-u.ac.jp) and that of the
NBRP.

Pathogenic Microbes
Repository Unit

Staff
Head (SUP): Tetsuya Iida, Ph.D.
Associate Professor (SUP): Toshio Kodama, Ph.D.

Publication(1) Reverse genetics for fusogenic bat-borne orthoreovirus
associated with acute respiratory tract Infections in humans:role of outer capsid protein sigmaC in viral replication andpathogenesis. Kawagishi T., et al. PLoS Pathog. (2016)
12:e1005455(2) Rapid whole genome sequencing of Miyazaki-Bali/2007
Pteropine orthoreovirus by modiƒed rolling circular
ampliƒcation with adaptor ligation - next generation
sequencing. Singh H., et al. (2015) Sci. Rep. 5:16517(3) Imported case of acute respiratory tract infection associatedwith a member of species Nelson Bay Orthoreovirus.
Yamanaka Y., et al. (2014) PLoS One 9:e92777

1) Oncolytic viral therapy using reovirus
Mammalian orthoreoviruses (reoviruses) are members of thefamily Reoviridae and contain a 10-segmented genome comprising
double-stranded (ds) RNA. Reoviruses are highly tractable experimental models for studies of dsRNA virus replication and pathogenesis. In the last decade, reoviruses have been evaluated in bothanimal models and humans as oncolytic agents against a variety oftumors, including head and neck, colon, breast, and pancreaticcancers. This is based on the observation that reoviruses induce cell
death and apoptosis in tumor cells with an activated Ras signaling
pathway. Wild-type reovirus-based oncolytic therapies are safe, butshow limited efficacy. We are developing safer and more effective
reovirus-based cancer therapeutics via genetic modification.
2) Highly pathogenic bat-borne reovirus
Bats are a natural reservoir for many important zoonotic
viruses, including Hendra virus, Nipah virus, and possibly Severe
Acute Respiratory Syndrome (SARS) coronavirus and Ebola virus.
Nelson Bay reovirus (NBV) was isolated from a flying fox in 1968,
but was not associated with any disease. Recently, Melaka virus,
which is genetically closely related to NBV, was isolated from ahuman patient in Malaysia suffering from an acute respiratory tract
infection (RTI). Subsequently, other related strains of bat-associated orthoreoviruses have been isolated in Malaysia, Indonesia, andChina. We also isolated and characterized another new NBV,
38

called Miyazaki virus, from a patient with acute RTI after returning to Japan from Indonesia in 2007. These isolates raise concerns
about bat-transmitted orthoreovirus infections in humans. We are
currently investigating how NBV replicates and causes diseaseusing a combination of genetic, biochemical, and biophysicalapproaches with an aim to developing vaccines, diagnostic tools,
and therapeutic agents.
3) Other reoviruses
Members of the family Reoviridae include rotaviruses (important
gastrointestinal pathogens in both humans and animals) and avian
reoviruses (economically important diseases in commercial poultry
flocks). We are also studying these viruses to better understand theirreplication, protein expression/function, and effect on cell biology.

Strategy to generate reovirus
from cloned cDNAs
The ten reovirus cDNAs aretransfected into murine L cells
expressing T7 RNA polymerase.
39

Genome Information Research Center

Department of Experimental
Genome Research

In the past, animals harboring natural mutations have beenused to elucidate the mechanisms underlying various diseases.
In the “post-genome project era,” genetically modified animals
play a key role in basic molecular biological investigations andact as models of human disease. Our laboratory studies themechanisms underlying mammalian reproductive systems
through genetic manipulation of animal models.

Prof.

Masahito Ikawa (SUP)
ProfileAnother tool improved by work in our laboratory is lentiviral(LV) vector-mediated genetic manipulation in vivo. We devel-

Publication(2) Structural and functional insights into
IZUMO1 recognition by JUNO inmammalian fertilization. Kato K., et al.
Nat Commun. (2016) 7:12198.

We were the first laboratory in the world to produce genetically modified mice that express green fluorescent protein(GFP) throughout the body (Fig. 1). These green fluorescent mice are useful for many types of research project.
Indeed, we used these animals to label sperm with fluorescent protein and visualize the fertilization process (ExpAnim. 2010; JCS. 2010, 2012; PNAS. 2012, 2013) (Fig. 2).
Recently, we found that calcineurin (PPP3CC/PPP3R2) isessential for sperm motility and male fertility (Science.
2015). Inhibiting sperm calcineurin may lead to the development of a reversible male contraceptive.

Development of new technologies for
producing genetically modified animals

Dr. Ikawa received his Ph.D. from
Osaka University in 1997. After working as JSPS postdoctoral fellow anda Research Associate at Genome Information Research Center in Osaka
University, he spent 2 years at The
Salk Institute in the USA as a Research Associate. After returning toOsaka University in 2002, he becamean Associate Professor in 2004 andwas appointed to the current position in 2012. He was awarded JSPS
Prize in 2013. His lifework is to study
mammalian reproductive systems
using genetically engineered mice.

(1) CRISPR/Cas9 mediated genome
editing in ES cells and its application
for chimeric analysis in mice. Oji A., etal. Sci Rep. (2016) 6:31666.

Analysis of molecular mechanisms involvedin mammalian gametogenesis, fertilization,and implantation(3) Genome engineering uncovers 54
evolutionarily conserved andtestis-enriched genes that are notrequired for male fertility in mice.
Miyata H., et al. Proc Natl Acad Sci USA.
(2016) 113(28):7704-10.
(4) Generation of Hprt-disrupted rat
through mouse←rat ES chimeras.
Isotani A., et al. Sci Rep. (2016)
6:24215.

(5) Sperm calcineurin inhibition prevents
mouse fertility with implications formale contraceptive. Miyata H., et al.
Science. (2015) 350(6259):442-5.

oped the technique of placenta-specific gene manipulation
by transducing blastocyst stage embryos with LV vectors(Nat Biotechnol. 2007; PNAS. 2011). Using this technique,
we are trying to elucidate the mechanism underlying implantation and placentation.
We also established rat embryonic stem (ES) cells andJHQHUDWHG PRXVHļUDW FKLPHULF DQLPDOV :H ZRXOG OLNH WR
use this animal model to study body/organ size control in
vivo; indeed, this method may enable derivation of variousorgans from ES or iPS cells (Genes Cell. 2011; Sci Rep.
2016).
Our recent interest is using the CRISPR/Cas9 system togenerate genetically modified animals to study fertilization,
implantation, and placentation. We have had success inmice and rats using sgRNA/Cas9-expressing plasmids (Sci
Rep. 2013,2016; DGD. 2014; PNAS.2016; Nat. Commun.
2016).
Our laboratory and the Animal Resource Center for Infectious Diseases (http://www.arcid.biken.osaka-u.ac.jp/) offer
support services such as generation of genetically modified
animals, in vitro fertilization, and cryopreservation of mousestrains. For more information, please visit our homepage(http://www.egr.biken.osaka-u.ac.jp/index.php).

Fig. 1. GFP-expressing mice are useful for many types of research project.
Genetically modified animals play a key role in basic molecular biology-based
investigations and are good models for studying human disease.

(6) Generation of mutant mice bypronuclear injection of circular plasmid
expressing Cas9 and single guided
RNA. Mashiko D., et al. Sci Rep. (2013)
3:3355.

Staff
Assis. Prof.：Yuhkoh Satouh (SUP) ／ Assis. Prof.：Yoshitaka Fujihara ／ Assis. Prof.：Haruhiko Miyata ／ Assis. Prof.：Masashi Mori ／SA Assistant Professor：Daiji Kiyozumi (SUP) ／ JSPS Postdoc：Taichi Noda ／ Postdoc：Keisuke Shimada ／Postdoc：Julio Manuel Castaneda ／ Postdoc：Kanako Kita ／／ Undergrad 3 ／ Graduate MA1・Ph.D.4 ／Visiting Researcher：Masaru Okabe ／ Guest Professor：Martin M. Matzuk ／ Guest Assoc. Professor：Ayako Isotani

40

Fig. 2. Mouse eggs (left) and sperm (right) labeled with fluorescent proteins. These gametes are used tovisualize the fertilization process. (Figs from Exp. Anim. 59(1), 105-7, 2010)41

Genome Information Research Center

Department of
Infection Metagenomics

Next-generation sequencing (NGS) is a technology that cangenerate enormous amount of genomic information in ashort time and has made huge progress in genomic scienceand infectious disease research. Staffs at the Department of
Infection Metagenomics who specialized in bioinformatics,
microbiology, and infectious diseases gather to conduct research on pathogens and infectious diseases using
NGS-based genomic/metagenomic analysis.

Head
Prof.

Toshihiro Horii (SUP)Prof.

Tetsuya Iida (SUP)

Development of methods for pathogen
detection based on metagenomic analysis
A metagenome is the sum of all genomes of all organisms
inhabiting a particular environment. The emergence of NGS
has enabled comprehensive analysis of genomic information from large numbers of organisms, thereby leading tosignificant advances in metagenomic analysis. For example, comprehensive analysis of microbial genomes in bloodor nasopharyngeal samples from patients suffering from
diseases of unknown cause makes it possible to identify thepathogens causing these symptoms and the genetic factors
responsible for pathogenesis. This method, unlike conventional pathogen-specific methods, is applicable to various
types of sample (e.g., blood, nasal swab, stool). It can also
detect multiple pathogens in a single sample. Our laboratory uses metagenomic analysis to develop new methods forthe diagnosis of infectious diseases.

Genomic analysis of microbial pathogens
PacBioMiSeq

Study of gut flora during onset of
infectious disease
It is becoming clear that the gut flora is microbiota that is
involved in various diseases and plays an important role inhost defense. By performing metagenomic analysis ofchanges in and recovery of bacterial gut flora over time incases of diarrhea, our laboratory is studying the relationshipbetween human gut flora and pathogens. Furthermore, notonly is bacterial gut flora related to disease, but it is also
closely related to lifestyle factors. Our research is focusedon how bacterial gut flora is affected by environmental
factors and the physiological state of the individual.
NGS technology had made remarkable progress. New
hardware platforms are being developed. NGS itself reads
only nucleic acid sequences, and further analysis isrequired to handle the enormous amount of data obtained.
It is important to have a broad knowledge of bioinformatics,
microbiology, and genomics in order to select the appropriate model based on the characteristics of each sequencing
platform. At our laboratory, we carry out co-operative
research with specialists in the fields of bioinformatics,
microbiology, and infectious diseases.

The molecular mechanisms underlying the pathogenicity ofmany infectious diseases remain unclear. Our laboratory
conducts genomic analysis-based research to identify genes
responsible for pathogenicity and to identify the molecularmechanisms by which infectious diseases develop.

Ion PGM

454 GS Jr

Publication(1) Herpes zoster laryngitis in a patient
treated with ƒngolimod. Hagiya H., etal. J Infect Chemother. (2016) Aug 20.
(2) Metagenomic Analysis of
Cerebrospinal Fluid from Patients with
Multiple Sclerosis. Perlejewski K., et al.
Adv Exp Med Biol. (2016) 935:89-98(3) Lypd8 promotes the segregation offlagellated microbiota and colonicepithelia Okumura R. et al., Nature.
(2016) Apr 7;532(7597):117-21.
(4) Homo-trimeric structure of the type IVb
minor pilin CofB suggests mechanismof CFA/III pilus assembly in humanenterotoxigenic Escherichia coli.
Kawahara K. J., et al. Mol Biol. (2016)
Feb 11.

(5) Metagenomic Analysis Reveals
Dynamic Changes of Whole Gut
Microbiota in the Acute Phase of
Intensive Care Unit Patients. Ojima M.,
et al. Dig Dis Sci. (2015) Dec 29.
(6) Performance comparison of secondand third-generation sequencers using
a bacterial genome with two
chromosomes. Miyamoto M., et al. BMC
Genomics. (2014) Aug 21;15(1):699.

Fig. 1.Large scale computer system for NGS data analysis.

Fig. 2.

Staff
SA Associate Professor：Shota Nakamura (SUP) ／Associate Professor：Naohisa Goto (SUP) ／Genomic analysis of Vibrio parahaemolyticus using four models of next-generation
sequencer: ■454 GS Jr（Roche）、■IonPGM（Life Technologies）、■MiSeq（Illumina）、■Pacific Biosciences RS System (PacBio)
GS Jr, MiSeq, and IonPGM produce short reads. Therefore, they require assembly of
short fragments. The third generation sequencer, PacBio, on the other hand can produce
long reads and assemble them into two long sequences with lengths equivalent to two
chromosomes. However, PacBio has low accuracy with respect to sequence information.
Although the read length of MiSeq is far shorter than that of PacBio, it has a much higher
yield. Thus, to conduct a proper analysis it is necessary to understand the characteristics
of each sequencing platform.

SA Assis. prof.：Daisuke Motooka ／ Postdoc：Mizue Anda

42

43

Genome Information Research Center

Laboratory of Genome Research

Germ Cell Group

Assoc. prof.

Takeshi Miwa

Assoc. prof.

ProfileProfileDr.Takeshi Miwa received his Ph.D. from Osaka
University in 1983. He appointed as a Assoc. prof. in
RIMD after working at The University of Tokyo andStanford University.

Dr. Nozaki received his Ph.D. from Osaka University
in 1986. He took his current position at RIMD in 2007
after working as a Research Associate and an Associate Professor in the same institution.

Publication

Publication(1) Connexin45 contributes to global cardiovasculardevelopment by establishing myocardial impulse
propagation. Nishii K., et al. Mech Dev. (2016) 140:41-52(1) Cetuximab-resistant oral squamous cell carcinoma cells
become sensitive in anchorage-independent culture
conditions through the activation of the EGFR/AKT
pathway. Ohnishi Y., et al. Int J Oncol. (2015) 47(6):2165-72(2) A novel heart failure mice model of hypertensive heartdisease by angiotensinⅡ infusion, nephrectomy, and saltloading. Tsukamoto Y., et al. Am J Physiol Heart Circ
Physiol. (2013) 305:1658-67(3) Interleukin-16 promotes cardiac ƒbrosis and myocardial
stiffening in heart failure with preserved ejection fraction.
Tamaki S., et al. PLoS One (2013) 8(7):e68893(4) L-Carnitine prevents the development of ventricularƒbrosis and heart failure with preserved ejection fraction
in hypertensive heart disease，Omori Y., et al. J
Hypertens. (2012) 30:1834-44

We are currently using animal models to investigate the molecularbiological mechanisms involved in human disease, particularly cardiovascular disease.
1) We have established a diastolic heart failure model using Dahl salt-sensitive rats. This model showed that left ventricular (LV) fibrosis and stiffening
play crucial roles in the development of heart failure with preserved ejection
fraction (HFpEF). Digitalis-like factors and the subsequent activation of the
Na+/Ca2+ exchanger may play important roles in the development ofhypertensive HFpEF and also regulate the effect of carnitine when administered to the HFpEF model. In addition, serum interleukin-16 (IL-16) levels
are elevated both in patients with HFpEF and in the rat model. Increased
cardiac expression of IL-16 in transgenic mice induces cardiac fibrosis and
LV myocardial stiffening, which is accompanied by increased macrophage
infiltration (Fig. 1).
2) To understand the cellular and molecular aspects of vascular smoothmuscle (SM) cell growth in atherosclerotic plaques, we characterized themechanisms responsible for transcription of SM-specific genes, particuODUO\WKHKXPDQ60DOSKDDFWLQ 6PĮ$ JHQH )LJ 6HYHUDOFLVDFWLQJ
'1$ HOHPHQWV DQG WUDQVFULSWLRQDO QXFOHDU IDFWRUV HVVHQWLDO IRU 6PĮ$
H[SUHVVLRQKDYHEHHQLGHQWLILHG6LQFH6PĮ$LVDOVRH[SUHVVHGLQPDQ\tissues during acute inflammation, we are examining expression of the6PĮ$DQGLWVIXQFWLRQ V 

44Masami Nozaki(2) Interaction between basigin and monocarboxylatetransporter 2 in the mouse testes and spermatozoa. ChenC., et al. Asian J Androl (2016) 18 (4):600-6(3) Isolation and propagation of neural crest stem cells frommouse embryonic stem cells via cranial neurospheres.
Minamino Y., et al. Stem Cells Dev (2015) 24 (2):172-81(4) The apoptotic initiator caspase-8: Its functional ubiquity
and genetic diversity during animal evolution. Sakamaki
K., et al. Mol Biol Evol. (2014) 31 (22):3282-301

Our body is derived from a single fertilized egg. During development, germ cells (and sometimes cancer cells), as well as many
types of tissues, are generated. We in the germ cell group havebeen conducting research focusing on germ cells, the early
embryo, stem cells, and cancer cells. Currently, we study osteogenesis from pluripotent stem cells and are characterizing cancer
stem cells.
Fig. 1. Increased cardiac expression of IL-16 in mice under
FRQWURO RI WKH Į0+& SURPRWHU FDXVHV LQFUHDVHG P\RFDUGLDO
fibrosis and stiffness. (Left) Four-chamber view of hearts from
non-transgenic and transgenic mice. (Right) Sirius Red-stained
sections of the LV from fibrotic areas.

Fig. 2.(PEU\RQLFDRUWD $R H[SUHVVWKHKXPDQYDVFXODU60
ĮDFWLQ SURPRWHU OHIW  EXW WKRVH ZLWK 0 FHQWHU  DQG 0
(right) point mutations in the transcriptional nuclear factor-binding regions do not.

Neural crest cells are derived from neuroepithelium via epithelial-mesenchymal transition (EMT) at the early developmental
stage. These cells migrate extensively and differentiate into
various tissues in the whole embryo. Bone is generated frommesoderm; however, most of the skull is derived from neural crest
cells. Recently, we established a method of differentiating ES cells
into maxillofacial bone via neural crest cells. We are conducting
research aimed at applying our findings to regenerative medicine
using iPS cells, adult tissue stem cells, and ES cells.
EMT is a phenomenon also observed in malignancy. Epithelial
cell-derived cancer cells acquire migration and invasive potency
after EMT, leading to metastasis. Some malignant cancer cells
have stem cell-like properties. Differences in the characteristics ofcancer stem cells and other cancer cells make the disease difficult
to treat. Thus, our aim is to understand the characteristics ofcancer stem cells and establish new treatment strategies.

Fig. Neuro-spheres derived from ES cells are induced to undergo osteogeneVLV E\ WUHDWPHQW ZLWK %03 DQG '(; &DOFLXP GHSRVLWLRQ LV VKRZQ E\
red-staining of alizarin, and upregulation of osteogenic marker genes duringosteogenesis is detected by semi-quantitative RT-PCR.

45

Thailand-Japan Research Collaboration Center
Section of Bacterial Infections

Until recently, it was believed that infectious diseases could
be conquered by developing chemotherapy regimens andvaccines; however, the recent worldwide emergence of new
infectious diseases and the re-emergence of infectious
diseases once considered to be under control have seriously challenged this notion. Since many infectious diseases
spread rapidly across national borders, it is clear that theycannot be controlled by the efforts of individual countries.
To overcome this, Osaka University founded the Research
Collaboration Center on Emerging and Re-emerging Infections (RCC-ERI) at the Thai National Institute of Health(NIH), Department of Medical Sciences, Ministry of Public
Health of Thailand, in 2005. The program is now in the third
phase (2015–2020) and is sponsored by the Japan Agency
for Medical Research and Development, which succeeded
to the second phase program named “the Japan Initiative for

Thailand-Japan Research
Collaboration Center

Global Research Network on Infectious Diseases (J-GRID).”
In addition to basic and applied research into emerging andre-emerging infections, we aim to develop human resources. We also aim to establish an effective system that would(i) provide information that would help prevent the spread of
emerging and re-emerging infections, and (ii) promptly
activate a variety of countermeasures should such a disease
emerge, including the development of therapeutics and/or
vaccines. Finally, we wish to enter into collaboration withlaboratories from nations that neighbor Thailand so that wecan be on the “frontline,” with the capacity to respond quickly
to the global spread of infectious disease.

SA Prof.

Kazuhisa Okada Profile
Dr. Okada received his Ph.D. from Osaka university in2005 and joined RIMD as a postdoctoral fellow in thatsame year. He was appointed Lecturer of Thailand-Japan Research Collaboration Center on Emerging andRe-emerging infections in 2015 after working as a Postdoctoral fellow from 2005 to 2011 and an Asistant Professor from 2011 to 2015 in the same institution.

Shigeyuki Hamada
SA Associate prof.

Kazuhisa Okada

Publication(1) Characterization of 3 Megabase-Sized Circular Replicons
from Vibrio cholerae. Okada K., et al. Emerg Infect Dis.
(2015) 21(7):1262-3.
(2) Cholera in Yangon, Myanmar, 2012-2013. Aung WW., et al.
Emerg Infect Dis. (2015) 21(3):543-4.
(3) Comparative genomic characterization of aThailand–Myanmar isolate, MS6, of Vibrio cholerae O1 El
Tor, which is phylogenetically related to a “US Gulf Coast”
clone. Okada K., et al. PLoS One. (2014) 9 (6):e98120.
(4) Vibrio cholerae O1 isolate with novel genetic background,
Thailand–Myanmar. Okada K., et al. Emerg Infect Dis.
(2013) 19:1015-7.

The collaboration center is located in the campus of the Ministry of Public Health.

BSL-2 and BLS-3 laboratories in the center.

46

Enteric infectious diseases caused by various microbes
occur frequently in Thailand. However, there are nolarge-scale epidemiological studies of the etiology ofgastroenteritis in Thailand. In the Section of Bacterial
Infections, we are trying to develop effective diagnostic
tools to detect bacterial pathogens and devise measures
to prevent enteric infections, including those mediated byVibrio cholerae O1.
We collect and analyze fecal specimens from patientswith severe diarrhea admitted to leading hospitals
located in different parts of Thailand. We then try to detectspecific pathogens by real-time PCR analysis of bacterial genes and culture analysis. We also attempt to identify
unknown pathogens using TOF-MS or NGS. In cases ofcholera in Thailand or Myanmar, we will co-operate withgovernmental authorities to analyze V. cholerae isolates
to elucidate the route of transmission. We will also
undertake molecular and/or genomic characterization of
any identified pathogen. Our research goals are to develop methods for rapid diagnosis and to prepare countermeasures against what is still a challenging pathogen.

Fig. Pulsed-field gel electrophoresis of intact genomic DNA of Vibrio

cholerae isolates and circular representation of the genome of V.
cholerae O1 El Tor TSY216, consisting of 3 chromosomes (Emerg
Infect Dis, 2015).

47

Thailand-Japan Research
Collaboration Center

Section of Viral Infections

Section of Bacterial Drug
Resistance Research

SA Prof.

Masashi Tatsumi

Thailand-Japan Research
Collaboration Center

SA Prof.

ProfileProfileDr. Tatsumi received his Ph.D. from The University ofTokyo. After working at National Institute of Health inJapan, Pasteur Institute and INSERM (French National
Institute of Health and Medical Research) in France, hespent 12 years in National Institute of Infectious Diseases. After retirement, he served as a chief advisor for JICA
until taking his current position at RIMD in 2016.

Dr. Hamada received is Ph.D. from Osaka university
Graduate School of Dentistry in1971. He joined the same
university as an Assistant Professor from 1977 and an
Associate Professor in 1977. He became the Director of
Dental Research, National Institute of Health in 1980. He
was appointed to current position in 2009 after working
as a Professor of Microbiology in Osaka University
School of Dentistry from 1986 to 2005 and a Professorof Nihon University Advanced Research Institute for the
Sciences and Humanities from 2005 to 2009.

Shigeyuki Hamada

Publication(1) Neutralization Activity of Patient Sera Collected during
the 2008-2009 Chikungunya Outbreak in Thailand.
Kishishita N., et al. J Clin Microbiol (2015) 53:184-90.
(2) Occurrence of hepatitis E virus infection in acute hepatitis
in Thailand. Siripanyaphinyo U., et al. J Med Virol (2014)
86:1730-5.
(3) Characterization of chikungunya virus-like particles.
Noranate N., et al. PLoS One (2014) 9:e108169.

Publication(1) Fetal septic meningitis in child caused by Streptococcus
suis serotype 24. Kerdsin, A., et al. Emerg. Infect. Dis.
(2016) 22(8):1519-20.
(2) Molecular and genomic characterization of pathogenic
traits of group A Streptococcus pyogenes. Hamada S., etal. Proc. Jpn Acad. Sci. (2015) B91:539-59.
(3) Characterization of 3 megabase-sized circular replicons
from Vibrio cholerae. Okada K., et al. Emerg. Infect. Dis.
(2015) 21(7):1262-3.

The Section of Viral Infections, in collaboration with the
NIH, Department of Medical Sciences, Ministry of Public
Health of Thailand, focuses on two emerging and re-emerging viral diseases that are prevalent in Asian countries.
One category of viral diseases under study is viral enteric infectious diseases, among which focus is placed on
those caused by norovirus, hepatitis A, and hepatitis E.
Norovirus causes food poisoning and is the etiologic agent
responsible for acute gastroenteritis via human-to-humantransmission. We perform evolutionary analysis based on
the genotyping of strains that cause pandemics. In
addition, our use of recombinant baculoviruses to investigate expression of norovirus virus-like particles, followedby production of monoclonal antibodies, is ongoing. The
ultimate goal is to develop rapid diagnostic tools.
A second category of diseases are mosquito-borne
infections, particularly chikungunya fever, which we investigate from an epidemiological, molecular biological, andimmunological point of view. We have developed a neutralizing antibody-based assay using a lentivirus pseudotyped with chikungunya virus envelope proteins. A recombinant baculovirus system produces chikungunya virus-likeparticles, which are used to examine the possibility of48

developing a chikungunya vaccine. In addition, analysis ofvarious cell libraries to identify and isolate the cellularfactors required for the virus replication is ongoing.

Fig. Upon expression, CHIKV (BSL3) structural proteins self-as-

semble into virus-like particles that are morphologically andantigenically similar to those of native CHIKV. A lentivirus
pseudotyped with CHIKV envelope proteins (BSL2) was also
used in the CHIKV neutralization test.

Carbapenem-resistant Enterobacteriaceae (CRE), including
Klebsiella pneumonia and Escherichia coli, are highly resistantto carbapenems and many other antibiotics. The rapidly
increasing prevalence of CRE over the past decade has
increased concern in healthcare facilities and public healthcommunities worldwide. Japan is no exception, even though
the prevalence of CRE at this time remains low. Our aim is to
examine the epidemiological dissemination of CRE in Southeast Asian countries.
Carbapenem resistance is usually carried by a plasmid(s)that harbors genes encoding carbapenemases, i.e., class A.3&VFODVV%PHWDOORȕODFWDPDVHV LQFOXGLQJ,039,0RU
NDMs), or class D OXA-type enzymes. We have attempted
to isolate CRE from patients admitted to leading hospitals in
Thailand and Myanmar. CRE isolates are assigned to particularspecies by biochemical characterization or MALDI-TOF-MS,
followed by profiling using pulse-field gel electrophoresis(PFGE) and multilocus sequence typing (MLST). We thendetermine the whole genome sequence of CRE isolates to
identify the full plasmid and construct a comprehensive imageof the relationships between isolates based on MLST andphylogeny. By undertaking these genomic epidemiologicalstudies, we increase our understanding of how CRE

infections spread and may be able to identify potential reservoirs.
Whole genome sequence data and epidemiological studies
may contribute to the development of diagnostic tools andimproved public health services.

Fig. PFGE and phylogenetic relationships among 44 isolates of carbap-

enem-resistant Klebsiella pneumoniae obtained from a single hospitalin Thailand. These isolates were separated into three major sequencetypes (ST), and comprised plasmid(s) carrying several carbapenem-resistant genes, including NDM-1, OXA-232, OXA-48, and IMP-14.

49

Thailand-Japan Research
Collaboration Center

Section of Antiviral Research

Prof.

Tatsuo Shioda

Thailand-Japan Research
Collaboration Center

ProfileDr. Shioda obtaind his B. Sc. from the University of Tokyoin 1982 and his Ph.D. form the same institution in 1990.
After working at the Institute of Medical Science in the
University of Tokyo and University of California San
Francisco, he was appointed as Associate Professor in
1997. He took his current position as a Professor of RIMD
in 2000.

The Mahidol-Osaka Center for Infectious Diseases (MOCID)
focuses on several tropical infectious diseases that are ofimportance to human health in Thailand. Mosquito-borne viral
infectious diseases such as dengue fever/dengue hemorrhagic fever and chikungunya fever are of particular interest.
We are currently developing rapid diagnosis kits and are
examining factors that affect disease severity. We would liketo improve the research skills of young scientists and increase
their interest in infectious diseases by collaborating with Mahidol University, which provides clinical samples.

Mahidol-Osaka Center for
Infectious Diseases

Staff
Director of MOCID: Prof. Tatsuo Shioda Ph.D
Associate Professor: Emi E. Nakayama, M.D.,Ph.D

Publication(1) Capsid-CPSF6 Interaction Is Dispensable for HIV-1
Replication in Primary Cells but Is Selected during Virus
Passage In Vivo. Saito A., J Virol. (2016) 11;90(15):6918-35.

Assistant Researcher/Ph.D student: Mr.Aekkachai Tuekprakhon B.SC.
Assistant Researcher: Juthamas Phadungsombat M.SC. (Microbiology)
Assistant Researcher: Narinee Srimark M.SC. (Public Health)(2) Roles of Capsid-Interacting Host Factors in Multimodal
Inhibition of HIV-1 by PF74. Saito A., J Virol. (2016) 27;90(12):5808-23

Publication(3) Macaque-tropic human immunodeƒciency virus type 1:
breaking out of the host restriction factors. Saito A, Akari
H. Front Microbiol. (2013) 9;4:187.

(1) Detection of chikungunya virus antigen by a novel
rapid immunochromatographic test. Okabayashi T., etal. Clin Microbiol. (2015) 53(2):382-8.
(2) Chikungunya virus was isolated in Thailand, 2010.
Sasayama M., et al. Virus Genes (2014) 49(3):485-9.
(3) Monoclonal antibody targeting chikungunya virus
envelope 1 protein inhibits virus release. Masrinoul P.,
et al. Virology (2014) 464-465:111-7.

Recently, climate change and urbanization haveincreased the risk of vector-borne diseases. Dengue andchikungunya viruses cause dengue fever/hemorrhagic
fever and chikungunya fever, respectively. Both viruses
are transmitted by Aedes mosquitoes. In 2014, there
was an outbreak of dengue fever in Tokyo. However, noantiviral drugs are available to treat these infections. Our
aim is to develop antiviral drugs against dengue andchikungunya viruses.

(4) Low levels of antibody-dependent enhancement invitro using viruses and plasma from dengue patients.
Chaichana P., et al. PLoS One. (2014) 18;9(3):e92173.

Diagnostic kit developed by the MOCID.

Four serotypes of dengue virus are distributed acrossthe world, and re-infection with different serotypes ofdengue virus leads to a more severe infection. We aim todevelop novel diagnostic tools that can differentiate
between the four serotypes of dengue virus.

Evaluation of CHIKV detection kit at Safdarjung Hospiral, Dehli,
India

50

Evaluation of CHIKV detection kit at the Institute of Tropical
Medicine Antwerp, Belgium

51

Endowed Chair

BIKEN Endowed Department of
Dengue Vaccine Development

Clinical Training Course on Tropical Infectious
Diseases in the Thailand
Endowed chair prof.

Eiji Konishi

ProfileDr. Konishi received his Ph.D. from Kobe University in
1980. He joined Kobe University medical school and Yale
University medical school after he got Ph.D. He was appointed to the current position in 2011 after working at
Kobe University School of Medicine and Graduate
School of Health Sciences as an Associate Professor.

Publication(1) A mouse monoclonal antibody against dengue virus type1 Mochizuki strain targeting envelope protein domain II
and displaying strongly neutralizing but not enhancing
activity. Yamanaka A., et al. J Virol. (2013) 87(23):12828-37.

The age of global travel means that people can spread pathogens worldwide. Infectious
diseases are now a global problem that extends beyond national borders. In Japan, there is
a compelling need for experienced specialists to study these infectious diseases.
Since 2009, RIMD, together with the School of Medicine at Osaka University, has provided
a clinical training course on Tropical Infectious Diseases in Thailand. The course provides
medical doctors with clinical training on the diagnosis and treatment of infectious diseases
and is supported by hospitals in Thailand. This training course provides a valuable
opportunity for Japanese clinicians to gain clinical experience from hospital staff operating ina high-incidence area. Over 50 doctors participated in this training course, and the alumni
are conducting basic/clinical research into infectious diseases or working for Medicines Sans
Frontieres and various governmental organizations.

(2) A review of successful flavivirus vaccines and the
problems with those flaviviruses for which vaccines are
not yet available. Ishikawa T.et al., Vaccine. (2014) 32(12):1326-37(3) Expression of enhancing-activity-free neutralizing
antibody against dengue type 1 virus inplasmid-inoculated mice. Yamanaka A., et al. Vaccine(2015) Nov 9;33(45):6070-7.

Hospitals participating in clinical training in Thailand.
Doctors can learn directly from local staff.

The BIKEN Endowed Department of Dengue
Vaccine Development was established in the
Faculty of Tropical Medicine, Mahidol University,
Thailand, in 2011 by an endowment from The
Research Foundation for Microbial Diseases ofOsaka University, Osaka, Japan to the ResearchInstitute for Microbial Diseases, Osaka University,
Osaka, Japan. This fiscal year is the last of the 6
year contract between these organizations.

induction of infection-enhancing antibodies and, therefore, maximize vaccine
efficacy. Our department has developed a new antigen that induces neutralizing, but not infection-enhancing antibodies. This technology is applicable toseveral vaccine strategies.

＜Hospitals for clinical training in Thailand＞
Maesot：Bangkok：

Mae Sot General Hospital
Mae La refugee camp
Mae Tao Clinic
Maeramad Hospital
Shoklo Malaria Research Unit

Ramathibodi Hospital, Mahidol University
Queen Sirikit National Institute of Child Health

Udonthani：

Dengue fever is a mosquito-borne viral disease
that affects those living in tropical regions.
However, autochthonous transmission of thisdisease to Japan and European countries intemperate regions was recently reported, making
dengue fever a global infectious disease. An
estimated 3.9 billion people are at risk of infection,with 390 million infections and 100 million cases
reported annually. The recently licensed Sanofi’s
vaccine does not show sufficient protective efficacy. Infection-enhancing antibodies that are induced
simultaneously with neutralizing antibodies may
affect the activity of the latter. Thus, there is aneed to design a vaccine antigen to minimize
52

Doctors gain hands-on experienceduring the training course.

Khon kaen：

Srinagarind Hospital, Khon Kaen University
Khon Kaen General Hospital

Udon Thani Genelas Hospital
A new antigen that induces neutralizing,
but not infection-enhancing, antibodies,
was successfully developed using amouse monoclonal antibody against dengue type 1 virus that possessed enhancing activity only (EAb, right panel). Manipulation of the epitope targeted by thisEAb suppressed induction of EAb. We
also generated a neutralizing-only antibody (NAb, left panel) and are currently
analyzing its epitope, which may be relevant to the development of an effective
dengue vaccine.

An antibody-expressing vaccine is a strategy designed to produce neutralizing-only
antibodies in the host. Heavy (H)- and light(L)-chain genes encoding the antibody-binding (Fab) region of NAb were cloned into
the antibody-expressing vector, pFUSE.
Mice co-inoculated with two plasmids harboring the H- and L-chains produced neutralizing antibodies 3 days later (left). However, no infection-enhancing activity was
detected (right). Mice maintained detectable levels of neutralizing antibodies for atleast 3 months, suggesting that an antibody-expressing vaccine can be used toprotect from dengue fever.

http://tmtc.biken.osaka-u.ac.jp/intention/index.html

53BIKEN Innovative Vaccine Research
Alliance Laboratories

As the recent Ebola virus outbreak in Africa and the worldwide influenza pandemic have powerfully demonstrated,society demands the development, production, and distribution of vaccines against infectious diseases. To meet thisstrong demand, The Research Foundation for Microbial
Diseases of Osaka University (BIKEN) and the ResearchInstitute for Microbial Disease, Osaka University (RIMD),
have established a new research organization, called The
BIKEN Innovative Vaccine Research Alliance Laboratories,
to promote the co-operative development of vaccines. Here,
we are developing new technologies to acquire basic
information required to design next-generation vaccines.

Staff

BIKEN Innovative Vaccine Research
Alliance Laboratories

Vaccine Creation Project

ProfileDrector, SA Prof.
Koichi Yamanishi M.D., Ph.D.

SA Assoc. prof.

Yasuo Yoshioka

Dr. Yoshioka received his Ph.D. from Osaka University in
2004. He took his current positon at RIMD from 2015
after working at National Institute of Health Sciences,
the Center for Advanced Medical Engineering and Informatics and Grad. School of Pharmaceutical Sciencesin Osaka University.

Publication(1) Distribution of Silver Nanoparticles to Breast Milk andTheir Biological Effects on Breast-Fed Offspring Mice.
Morishita Y, Yoshioka Y, et al. ACS Nano. (2016) Aug 15.
(2) Metal nanoparticles in the presence oflipopolysaccharides trigger the onset of metal allergy inmice. Hirai T, Yoshioka Y, et al. Nat Nanotechnol. (2016) 11(9):808-16.
(3) Silica and titanium dioxide nanoparticles cause
pregnancy complications in mice. Yamashita K, Yoshioka
Y, et al. Nat Nanotechnol. (2011) 6(5):321-8.
(4) Interleukin-1 family cytokines as mucosal vaccine
adjuvants for induction of protective immunity against
influenza virus. Kayamuro H, Yoshioka Y, et al. J Virol.
(2010) 84(24):12703-12.

A regular meeting between laboratories.
All members for this research project attend the meeting. Researchers address their research topics in depth andtake part in active discussions.

Most protein antigens such as non-living macromolecules or protein-subunit antigens evoke weak or undetectable adaptive immune responses. Therefore, todevelop effective vaccines it is necessary to develop
vaccine adjuvants and antigen delivery carriers. In
addition, to develop optimal (in terms of efficacy andsafety) vaccines for clinical application, it is important tounderstand the mechanism by which vaccines act on
the immune system. In this regard, our research is
focused on optimizing vaccines through drug delivery
systems and safety science. Our specific research
projects are:1) Development of vaccine adjuvants using comprehensive screening methods.
2) Development of antigen delivery carriers andadjuvants using nanotechnology.
3) To use these adjuvants and delivery carriers todevelop vaccines for infectious diseases.

Experimental laboratories.
Researchers can move freely among three laboratories. All threelaboratories have common equipment.

54

55

BIKEN Innovative Vaccine Research
Alliance Laboratories

BIKEN Innovative Vaccine Research
Alliance Laboratories

Mucosal Vaccine Project

Vaccine Dynamics Project

SA Assoc. prof.

Shintaro Sato

SA Assoc. prof.

ProfileProfileDr. Sato received his master’s degree from Hokkaido university in 1999 and his Ph.D. from Osaka University in
2003. He took his current position in 2015 after working
at RIMD as a Postdoctoral fellow and at University ofTokyo as an Assistant Professor.

Dr. Aoshi received his M.D. from Hamamatsu University
School of medicine in 1999 and his Ph.D. from the same
institution in 2006. He was appointed current positon in2015 after working at Washington University in St. Louis,
NIBIOH, and IFReC in Osaka University.

Publication(1) IL-10-producing CD4+ T cells negatively regulate
fucosylation of epithelial cells in the gut. Goto Y., et al.
Sci. Rep. (2015) 5:15918.
(2) Innate lymphoid cells regulate intestinal epithelial cell
glycosylation. Goto Y., et al. Science (2014) 345(6202):1254009.
(3) Transcription factor Spi-B-dependent and -independent
pathways for the development of Peyer's patch M cells.
Sato S., et al. Mucosal Immunol. (2013) 6(4):838-46.
(4) Indigenous opportunistic bacteria inhabit mammalian
gut-associated lymphoid tissues and share a mucosal
antibody-mediated symbiosis. Obata T., et al.
Proc. Natl. Acad. Sci. U S A. (2010) 107(16):7419-24.

Because most pathogens invade and infect their host
via mucosal tissues, mammals have established astrictly organized and dynamic immune system at mucosal surfaces. This system, named the mucosal immune
system, combats infectious pathogens. The development of mucosal vaccines, which activate mucosal andsystemic immune responses, is receiving increasing
attention. We have focused on epithelial cells, particularly M cells, which are professional antigen uptake cells
located in areas that come into contact with non-self-antigens. Our main research themes are:1)To identify new M cell-specific genes and elucidate
the mechanism of M cell antigen uptake.
2)To understand the aged mucosal immune systemand develop effective mucosal vaccines for older
people.
3)To establish an in vitro culture system for normal
and functional primary mucosal epithelial cells anduse it to screen for candidate mucosal vaccine
antigens and adjuvants.

56

Taiki Aoshi

Publication(1) Development of non-aggregating poly-A tailed
immunostimulatory A/D-type CpG oligodeoxynucleotides
applicable for clinical use. Aoshi T., et al. J Immunol Res.
(2015) 2015:316364. doi: 10.1155/2015/316364.
(2) Plasmacytoid dendritic cells delineate immunogenicity of
influenza vaccine subtypes. Koyama S., et al.
Sci Transl Med. (2010) Mar 31; 2(25):25ra24.
(3) The cellular niche of Listeria monocytogenes infectionchanges rapidly in the spleen. Aoshi T., et al.
Eur J Immunol. (2009) Feb; 39(2):417-25.
(4) Bacterial entry to the splenic white pulp initiates antigen
presentation to CD8+ T cells. Aoshi T., et al. Immunity(2008) Sep 19;29(3):476-86.

Vaccination, which utilizes the built-in “immune system”
in the body, is one of the most successful medicine forcontrolling microbial infections. The immune systemcomprises many different immune cells. T cells are oneof the most important immune cells that help to establish
protective immunity against many pathogens. In
addition, T cells are also involved in many non-infectious
conditions such as autoimmunity and cancer.
Although T cells are critically involved in many diseases, current vaccines (with the exception of attenuatedlive vaccines) cannot induce sufficiently strong T cell
responses. We are developing “T cell inducing vaccine”
through the studies of T cell epitope and T cell/antigen
presenting cell interaction. We are also developing
clinically applicable “nucleic acid adjuvants” that not only
induce strong Th1/CTL responses but also retain goodsafety profiles. We believe that the understanding of T
cell epitope, T cell/antigen presenting cell interactions,and the development of new adjuvants will enable us tomake safer and more effective T cell inducing vaccines in
the near future.

57

Animal Resource Center for Infectious Diseases

DNA-chip Development Center for Infectious Diseases

Staff
Head, Prof.: Masahito Ikawa, Ph.D.
Assis. prof.: Yuhkoh Satouh, Ph.D.
Assis. prof. (SUP): Yoshitaka Fujihara, Ph.D.
Assis. prof. (SUP): Haruhiko Miyata, Ph.D.
Assis. prof. (SUP): Masashi Mori, Ph.D.

To study infectious diseases and cancer, it is important toanalyze interactions between pathogenic factors and thehuman body. Animal models are indispensable for biomedical
research, particularly since molecular biology and biotechnology methods can be used to generate genetically modified
mice that aid our understanding of the molecular mechanisms
underlying such diseases. Experimentally infected animals
and genetically engineered animals used for these purposes
should be managed in a suitable, safe, and controlled manner.
The Animal Resource Center for Infectious Diseases is aunique facility that was established in 1967 to meet theserequirements.
The center is equipped with pass-through-type autoclave
systems and HEPA filtered air exchange systems to minimize
the risk of contamination so that infected or geneticallyengineered animals are maintained in a safe environment.
The animals are housed in three areas: SPF (specific pathogen free), BSL (biosafety level) 2, and BSL3. Before gaining
access to restricted areas, researchers must take an official
orientation and submit a research plan for committee review.
The condition of the animals is monitored regularly.

SA Assis. prof. (SUP): Daiji Kiyozumi, Ph.D.
Guest Associate prof.; Ayako Isotani, Ph.D.

IVF/ETTG

KO, KI

-2000

261

228

50

2001-2003

443

104

57

2004-2006

331

43

69

2007-2009

216

22

74

2010-2012

388

55

152

2013-2015

580

50

242*

Table 1 *Tg, Transgenic; KO, Knockout; IVF, in vitro
Fertilization; ET, Embryo transfer.
* Includes lines generated using the CRISPR/Cas9
genome editing system.

Our facility provides the following services: generation ofgenetically manipulated animals, in vitro fertilization, andcryopreservation of mouse strains (Table 1). The facility
provides these services in co-operation with the Department
of Experimental Genome Research.

Facility Management: The DNA-chip Development Center for
Infectious Diseases is a unique facility that was established in 2004 to
analyze the transcriptional dynamics and variations involved in
infectious diseases. We provide the following contract analysis
services and various research supports such as use briefings andtraining courses of installed apparatus. We also conduct the following
research projects in this facility.
(A) Contract analysis service(1) Transcriptome analysis using DNA-chip analyzers.
The high density DNA microarray system in this facility permits
comprehensive transcriptional analysis of gene expression in thehuman or mouse host, and in various pathogenic organisms. Two
DNA microarray systems, namely the Agilent-type and the Affymetrix-type, are available in this center.
(2) Nano-counter Analysis System (NanoString Technologies):
This system is also useful for more accurate quantitative analysisof the transcriptional levels of particular genes.
(3) Proteome analysis using mass spectrometry.
The MS/MS spectrometer installed in this facility enables theanalysis of the expression, interactions and modifications ofproteins from humans, mice, and pathogenic organisms. Thiscenter is also capable of recent technical innovations, such as themass spectrometric detection of pathogenic organisms thatfacilitates the development of novel diagnostic systems for
infectious diseases.
(4)Gene expression analysis using Next Generation Sequencer(collaboration with Dept of Infection Metagenomics)(5) Gene data analysis.
Gene data analysis using IPA and NextBio is also available in thiscenter.
(B) Research Projects
Based on DNA microarray analysis on mRNA from the peripheral bloodmononuclear cell (PBMC) of patients suffered from autoimmune
diseases such as vasculitis, we have identified several genes that are
specifically up-regulated in PBMC of these patients. IPA and NextBio
softwares we used are useful data mining tools to detect such disease
specific genes from big data obtained comprehensive gene analysis
without deep knowledge on bioinformatics.

Staff
Head, Prof. (SUP): Hiroshi Nojima, Ph.D.
Assoc. prof. (SUP): Norikazu Yabuta, Ph.D.
Assis. prof.: Daisuke Okuzaki, Ph.D.

DNA microarraynCounterSystem

Publication(1) IFI27 Is a Useful Genetic Marker for Diagnosis of
Immunoglobulin A Nephropathy and Membranous
Nephropathy Using Peripheral Blood. Nagasawa Y., et al.
PLoS One. 2016 Apr 21;11(4):e0153252.

Biosafety level 3 room.
The room is used for research involving disease model animals at
biosafety level 3. Hemorrhagic fever with renal syndrome-causing
virus (HFRSV) was isolated in this area. In addition, animal experiments involving Zika virus, SARS, and Acquired Immune Deficiency Syndrome (AIDS), can be handled in this facility.

58

Buildings at the Animal Resource Center.
Building A (in front of the chimney, built in 1967, two-story).
Building B (rear right of the chimney, built in 1978, four-story).
Building C (on the right side of Building A, built in 2009, four-story).

(2 )Interleukin-18-deƒcient mice develop dyslipidemia resulting in
nonalcoholic fatty liver disease and steatohepatitis. YamanishiK., et al. Transl Res. 2016 Mar 19.
Next Generation Sequencer HiSeq(3) Microarray and whole-exome sequencing analysis of familial
Behçet's disease patients. Okuzaki D., et al. Sci Rep. 2016 Jan
20;6:19456. doi: 10.1038/srep19456.
(4) CAWS administration increases the expression of interferon
gamma and complement factors that lead to severe vasculitis
in DBA/2 mice. Nagi-Miura N., et al. BMC Immunol. 2013 Sep
24;14(1):44.

59

Office of Combined Program on Microbiology
and Immunology

Education Promotion Group

Our institute and the Immunology Frontier Research Center are world-class institutes for microbiological and immunologicalresearch, respectively. To take maximal advantage of this situation and to foster intellectual exchange between these two
institutes, we have developed a combined microbiology and immunology program.

Research Promotion GroupTo facilitate research on microbiology and immunology, we havebased our multidisciplinary graduate program on microbiology andimmunology. We dedicate to planning and management of thegraduate program and also organize Open House sessions andprovide guidance for new graduate students and post-docs. The
aim of these activities is to promote communication between
researchers and to develop human resources with expertise covering both microbiology and immunology.

Associate Professor:

Hodaka Fujii,
M.D., Ph.D.

Our aim is to promote collaborative research, information exchange, andpeople-to-people exchange between laboratories via:

Associate Professor:

Yoshiko Murakami,
M.D., Ph.D.

1. Organization of the Awaji International Forum on Infection and
Immunity (annually in September). http://awaji-forum.com
2. Organization of the Biken monthly seminar for reporting research
progress.
3. Organization of the Annual Research presentation program and thecompetition of research presentations.

Assistant Professor:

Toshitsugu Fujita,
Ph.D.

Lecture series for graduate students

Awaji International Forum

Awaji Yumebutai
International Conference Center

Joint Orientation for RIMD and IFReC

Research Project(Chromatin Biochemistry Research Group)

Biochemical analysis of molecularmechanisms of genome functions
using the locus-speciﬁc chromatin
immunoprecipitation technologies

A seminar held in December

Academic Prize awardee

Research projects
Dr. Murakami concurrently serves as the leader of the PNH group in the Dept. of Immunoregulation andundertake the following profects.
1. Investigation of the pathogenesis of acquired glycosylphosphatidylinositol (GPI) deficiency and PNH.
2. Investigation of the pathogenesis of IGD.
See the Dept. of Immunoregulation and website (http://igd.biken.osaka-u.ac.jp) for details.

Publication(1) Pathogenic variants in PIGG cause
intellectual disability with seizuresand hypotonia. Makrythanasis P., etal. Am. J. Hum. Genet. (2016)
98:615-26.

60(2) Null mutation in PGAP1 impairs
GPI-anchor maturation in patientswith intellectual disability andencephalopathy. Murakami, Y., et al.
PLoS Genet. (2014) 10(5):e1004320.

(3) Glycosylphosphatidylinositol (GPI) anchordeƒciency caused by mutations in PIGW is
associated with West syndrome andhyperphosphatasia with mental retardation
syndrome. Chiyonobu, T., et al. J. Med. Genet.
(2014) 51:203-7.

A comprehensive understanding of the mechanisms
underlying genome function, such as transcription andepigenetic regulation, requires identification of molecules that bind to the genomic regions of interest in vivo.
To facilitate the biochemical and molecular biologicalanalysis of specific genomic regions, we have developed locus-specific chromatin immunoprecipitation
technologies (iChIP and enChIP) to purify the genomic
regions of interest. We will then apply these locus-specific
ChIP methods to elucidate the molecular mechanisms
underlying important epigenetic phenomena such as
regulation of tumor suppressor gene expression, DNA
repair, and the lineage commitment of lymphocytes.

Publication(1) Efƒcient sequence-speciƒc isolation of DNA
fragments and chromatin by in vitro enChIP
technology using recombinant CRISPR
ribonucleoproteins. Fujita T., et al.
Genes Cells (2016) 21:370-7.

(2) Identiƒcation of non-coding RNAs
associated with telomeres using a
combination of enChIP and RNA
sequencing. Fujita T., et al. PLoS One (2015)
10:e0123387.

(3) Identiƒcation of proteins associated withan IFN¸-responsive promoter by a retroviral
expression system for enChIP using
CRISPR. Fujita T., et al. PLoS One (2014) 9:
e103084.

61

Common Research Facilities

Common Research Facilities

Central Instrumentation
Laboratory

Head, Professor:
Hiroaki Miki, Ph.D.
Associate professor:
Shinji Higashiyama, Ph.D.
Associate professor:
Naohisa Goto, Ph.D.
Assistant professor:
Kazunobu Saito, Ph.D.
SA Assistant professor:
Miki Morimatsu, Ph.D.

The Central Instrumentation Laboratory was established in 1959. When equipment was lacking in many laboratories, researchers brought their machines
together and co-operated with each other. Now, various pieces of precisionapparatus and high performance machines are available in the laboratory at all
times. These include ultracentrifuges, transmission and scanning electron microscopes, a Biacore system, cell analyzer/sorters, an DNA sequencers, and mass
spectrometers. Also, large cell storage tanks equipped with automatic liquid
nitrogen supply systems and a specified chemical treatment room are alsopresent. In addition, professional technicians are employed to maintain andmanage these devices, as well as to provide services, education, and training fornewcomers. They also provide in-house services such as cell sorting, mass
spectrometry-based protein identification, electron microscope image capture,and DNA sequencing. As experimental machines become more and more
complicated, the services provided by specialist staff are essential for ongoing
research at the institute.

Central Laboratory for Biological
Hazardous Microbes

Central Instrumentation Laboratory

Radioisotope Labolatory

Head, Professor:
Hiroaki Miki, Ph.D.
Associate professor:
Shinji Higashiyama, Ph.D.

62

The radioisotope (RI) laboratory was established in 1967 and was designed forbiomedical experiments involving RIs. Now, RIMD researchers perform RI
experiments in the RI Laboratory at the Immunology Frontier Research Center at
9F, the Central Laboratory for Biological Hazardous Microbes at 1F, and theradiation exposure room in the North building at 1F. Facilities include a RI stockroom, a distribution room, a tissue culture room, and an area for RI measuring
equipment. Safety requirements are met by a stringent security system thatinvolves the use of ID cards and the computerized management of RIs.

This BSL-3 laboratory was set up in1983 to ensure the safe handling of
biologically hazardous microbes such as
hemorrhagic fever with renal syndrome(HFRS) virus. All experimental studies
using hazardous microbes, such as HIV,should be handled in this laboratory. The
laboratory is a three-story building with
550 m2 of floor space. The facilities aredesigned to protect researchers from
pathogenic infection and to prevent the
spread of biohazardous pathogens
outside the building. The supply of fresh
air is regulated to keep the room interiors
at negative pressure. High-quality filters
are installed on the exhaust outlet tominimize microbial contamination of theenvironment. Each room is equippedwith safety cabinets and autoclaves tosterilize used material before disposal.
Researchers must be approved by the
Biosafety Committee before they use
this laboratory. Various microbes, including HIV, SARS corona virus, and scrapie
agent, are studies in this facility.

The RIMD library contains academic books and journals about microbiology,
immunology, oncology, and other related scientific fields such as cell biology,
genetics, histology, developmental biology, biochemistry, pharmacology, andpathology. In particular, we have collected rare books on parasitology thatcannot be found at other institutions; these books are frequently accessed byvisitors to the RIMD library. Now, the RIMD library owns 6,800 books anddozens of scientific journals, some of which are donated by the BIKEN foundation. Most of the books and journals in the RIMD library are registered on theindex at the main Library of Osaka University, which can also be accessed bylibraries throughout Japan via the Interlibrary Loan (ILL) system. One librarian
handles the RIMD library, and two professors and two associate professors are
members of the RIMD library committee.

General Affairs Section
Accounting Section
Research Cooperation Section
Planning Ofﬁce
PR Ofﬁce

Central Laboratory for Biological
Hazardous Microbes

Head, Professor:
Tatsuo Shioda, Ph.D.

RIMD libraryHead, Professor:
Tetsuya Iida, Ph.D.

Administration andResearch Support

63

About RIMD

RIMD History

Elucidationof the Innate
Immune System
Shizuo Akira

Discovereda viraloncogene

Research Institute for Microbial
Diseases opened
RIMD was founded through amerger of the Research Centerfor Communicable Diseases (Osaka Medical School), the Takeo
Tuberculosis Institute (donatedby Mr. Jiemon Takeo), and the
Osaka Leprosy Institute (donated by an anonymous benefactor).

Kumao Toyoshima

2000
2010

1980

Discovered
cell fusion
Yoshio Okada

Approved as a Joint
Usage/ResearchCenter by
MEXT

1990

Developed
a malaria
vaccine1967

Toshihiro Horii

1950

1970
1960

1930
1940
Discovered
Vibrioparahaemolyticus
Tsunesaburo Fujino

Developed
a chickenpox
vaccineMichiaki Takahashi

RIMD moved to
the Suita Campus

Developed
a measles
vaccineYoshiomi Okuno

RIMD buildings in 1967

Institute construction wascompletedWith ﬁnancial support from Gendo
Yamaguchi, a benefactor from Osaka, the
RIMD main building was constructed in
Dojima, Osaka.

Tenji Taniguchi

Biken
History Museum

Professor of Bacteriology at
the Osaka Medical school. He
played a huge role in the foundation of RIMD as he emphasized the need for a research
institute in the KANSAI areathat focused on microbial or infectious diseases.

A model of the HEV-hepatitis
E virus
The first ultracentrifuge rotor,
installed about 60 years ago

Koch’s microscope
Prof. Taniguchi (right) andLab Members

Gendo Yamaguhi
A successful businessmanin the KANSAI area. He
gave back to the community by offering his property
for public beneﬁt services
and temples. He donated200,000 yen to establish
RIMD.

Opening hours: 9:00–17:00 (Mon–Fri)
Location: RIMD Main Building 1F
Fee: Free
Biken History Museum website
http://museum.biken.osaka-u.ac.jp/
english/home.html

A manipulator used for research in RIMD

Prof. Tenji Taniguchi’s Desk

Pictures and materials are provided by Biken History Museum
6465About RIMD

RIMD Awards

66

67

About RIMD
RIMD Joint Usage / Research Center

General Research project, Specific Research project,
Short term
Short term

Research project,Long term

To help victims of
the earthquake

Total
Symposiums

Date

Participants

2010
2011

2

2012

2
Awaji International Forum on

2013

Infection and Immunity

2014
2015
Total

4
France-Japan International Exchange Symposium

RIMD-CVRDC Joint SymposiumInternational Symposium of the Institute Network※

68

Year

2010

2011

2012

2013

2014

2015

Projects

24

42

49

42

32

35

Country

Institute / University

Republic of Uganda

Faculty of Medicine, Gulu University

ThailandBamrasnaradura Infectious Diseases InstituteThailandFaculty of Tropical Medicine, Mahidol University

Indonesia

Faculty of Medicine, Airlangga University

RIMD

Triangle
Relationships
on Infectious Diseases
and Immunology

69

For Students andResearchers who wants
to study in RIMD

> Osaka University COOP guidance book

The Osaka University COOP is a non-profitable organization for students andstaffs in Osaka university. They provide stores, cafeterias andother life supports including housing and traveling.

http://www.osaka-univ.coop/english/index.html

RIMD is one of the world’s foremost institute in immunology, microbiology and cancer research.
We also conduct research in various bioscience related fields including gene engineering,genomic science and bioinformatics. We welcome motivated grad-students and researchers fromaround the world.
The way to join RIMD would be different depends on the situation. Candidate for grad-school
students or post-docs may need to decide the lab to join and then ask PIs how to belong to RIMD.
The Orientation and lab tour would be held in May every year. Please check our website for detail.

http://suishin.biken.osaka-u.ac.jp/setsumeikai/setsumeikai_en.html

Information in Japanese Government or Organization> Study in Japan Comprehensive Guide by The Ministry ofForeign Affairs of Japan
http://www.studyjapan.go.jp/en/index.html> Websites of Japanese Embassies in your country
http://www.mofa.go.jp/about/emb_cons/mofaserv.html> Japan Student Services Organization (JASSO)

An independent administrative institution established under the MEXT,
comprehensively administers support programs forinternational students including scholarship loan programs.

Information in Osaka University website> Osaka University website for Global Affairs
http://www.osaka-u.ac.jp/en/international> Osaka University website for International Students
http://www.osaka-u.ac.jp/en/for-student> Osaka University Support Office for international Students
https://iss-intl.osaka-u.ac.jp/supportoffice/> Osaka University International Students Association
http://ouisa.info/

http://www.jasso.go.jp/en/index.html> Gateway to study in Japan by JASSOInformation in Japanese, English, Chinese, Korean, Indonesia, Thai, Vietnamese

http://www.g-studyinjapan.jasso.go.jp> Japanese Government scholarship by MEXT
http://www.studyjapan.go.jp/en/toj/toj0302e.html> Suita International Friendship Association

A public interest Incorporated Foundation to
support citizen’s international exchange activities and provide assistance toforeigners living or Studying in Suita city.

http://suita-sifa.org/en/> Osaka University Brothers and Sisters Program (BSP)70

An International-exchange circle organized by Osaka University
Students to support international students.

RIMD provides our own scholarships for international students, Taniguchi scholarship. 3 grad

http://www.bsp-ou.net/

studying in RIMD on this scholarship.

students from Indonesia (2 students in master’s course, 1 student in doctoral course) are71

Grad Students Studying in RIMD
Why RIMD?

Research Interest

I have deep interest in learning Japanese language andexperiencing the cultures in Japan. Most importantly,
the thought of involving in a Japanese research teamlead by top university’s department is tremendously
exciting as it offers me the opportunity to grasp moreinsight into biological science field, which I particularly
admire and appreciate. Thus, I applied for Japanese
Government Scholarship (MEXT scholarship), and it
took me almost a year to get through the selection
process, with a bit of luck I managed to secure thescholarship*. Since I am interested in the field of
oncology, I searched for cancer research labs in Japanand came into contact with Professor Okada Professor
Associate Nada via Okada-lab webpage. Bothprofessors were very friendly in guiding and handling myadmission in Department of Oncogene, RIMD, Osaka
University. Since then, I was admitted as a ResearchStudent in October 2015 and few months later I passed
the entrance examination in 2016, I became a graduate
student for the Graduate school of Science, Osaka
University.

Coming to graduate school, I knew I
wanted to study the basic processes
governing cell function and development in a
genetically traceable organism. I also wanted towork under the guidance of an advisor who took a
primary interest in my scientific work. Therefore, myresearch is mainly focused on elucidating andcomparing the regulation of TGF-B signaling pathwaythat confers to cancer resistance in Naked Mole Rat(NMR) and house mouse. In brief, NMR is an organism ofinterest that provides a platform for understanding how
cancer resistance can be achieved in mammals ingeneral since it possess exceptional longevity and isremarkably cancer-resistant. Thus, increased insight
into molecular mechanism at the interface of aging andage-related disease, such as cancer, is crucial formaking progress in improving human health. In anutshell, it is pivotal to elucidate the signaling pathway
of TGF-B in which NMR might rely on this pathway toconfer cancer resistance, as compared to its short-lived
cancer-prone mice that lack the mechanism that lead to
extreme longevity and cancer resistance.

A day in the life
I am taking Special Integrate Science course in the
Graduate school of Science. The course is mainly forinternational students in which the lectures are held in
English. Since some Japanese students are also taking
some lectures in SISC program, from the interactive
conversation with the local students, we can share anddevelop deeper understanding in this extraordinary
biological world. Apart from that, Sensei and laboratory
comrades in the Okada-lab are very friendly. And I
wholeheartedly agree to my friends saying that KANSAI
people are friendlier than people from east part ofJapan. Currently, I really enjoy my daily life in Osaka.

Message for young students.
I hope that young students will set directions of theirown goal, as the direction of one’s goal is the directionof one’s life will move on. Most importantly, let yourself
move towards what is good, valuable, strong and true.
And not to forget to try new things and experience
enticing adventures, keep moving forward as curiosity
surely will lead you down new path and from there youcan embark your amazing journey of life.
*Only 24 students out of several thousand applicants in
Malaysia could get the scholarship!

Why RIMD?

Research Interest

I approached one of my professors in the
East Asian Language and Culture department
at UC Berkeley about my future during my lastyear of college. He insisted that I would be a top
candidate for the MEXT scholarship to pursue my
Master’s degree in Japan given my dual degrees in
Biology and Japanese. I have always been passionate
about women’s health and reproductive biology, so I
reached out to one of my biology professors to ask if she
had any contacts in Japan. She gave me RIMD Professor
Masaru Okabe’s email address and I sent him amessage about my interest. He quickly replied backsaying that while he retired, Professor Ikawa took overthe laboratory and to contact him. From there, it all fell
perfectly into place. Professor Ikawa was incredibly kindand helpful throughout this journey. I was delighted tolearn that the Ikawa lab at Osaka University, one of the
top reproductive biology labs in the field, offered theideal opportunity for me to gain experience and pursue
my career aspirations as a researcher focused on thefemale reproductive system. After securing the MEXT
scholarship with his help, I was admitted as a ResearchStudent in April 2015, passed the Master’s course
entrance exam in August 2015, and became a graduate
student in the Graduate School of Medicine at Osaka
University.

A day in the life

My family history of reproductive diseases initiallymotivated me to pursue a scientific career focused on
the female reproductive system. As I learned about thehealth benefits of female contraception specific toovarian cancer prevention, I became further intrigued
with female fertility factors. As a Master’s student inProfessor Ikawa’s lab, I am currently using the
CRISPR/Cas9 system to create knockout mice to findgenes essential for female fertility. CRISPR/Cas9 iscurrently the leading technology for gene manipulation.
By injecting a signal guide RNA sequence into fertilized
mouse eggs, we can recruit the Cas9 protein to cut ourtarget DNA sequence, rendering the protein functionallyuseless. While the majority of research conducted inthis lab is based on the male reproductive system, my
passion for the female system urges me to uncover the
unknown in female reproduction. For the next two years,
my goal as a Master’s course student is to find specificfemale infertility factors that could ultimately betranslated into contraception.

Message for young students.
While I have been studying Japanese since high school,
it did not prevent the ups and downs of living in a brandnew country. Nonetheless, perseverance, believing inmyself, and not being afraid to ask for help has led me towhere I am. I am always aware of my privilege and am
thankful to those who have supported me. If it were notfor my undergraduate professors, I would not be inJapan right now. Finding strong mentors is key to future
success.

The Graduate School of Medicine Master’s course isspecial in that all of the lectures are packed into the firsttwo and a half months of the program. Because thecourse is mainly for Japanese students, all of thelectures are conducted in Japanese. While it has been achallenge for me to learn biological Japanese
vocabulary in the beginning, I can confidently saythat my vocabulary has increased
tremendously. Not only that, sharing thatexperience with my Japanese classmates
was an amazing opportunity to further
Department of Experimental
develop my Japanese. Now that classes
Genome Research (Ikawa Lab)
have ended, I am back full time in the
Master’s Course in the
Ikawa lab. While continuing to do
Graduate School of Medicine
experiments and furthering the bonds I
BA in Molecular and Cell Biology andhave with my labmembers, I am also
BA in Japanese Language
teaching them English and learning to speakfrom the University of California,
the Kansai dialect!
Berkeley Class of 2014

Ferheen Abbasi

Woei-Yaw Chee
Department of Oncogene Research (Okada Lab)
Masters course in Graduate school of Science, Osaka university
BA: National University of Malaysia / Bournemouth university, UK

72

73

RIMD STAFF (2016.4)

BUILDING AREA
●Graduate Students●Staffs①②

Professor

16

Graduate school of Medicine

436Endowed Chair Professor

1

Graduate school of Science

4

9

SA Professor

4

Graduate school of Pharmaceutical Science

40

Associate Professor

17

Graduate school of Dentistry20

SA Associate Professor

11

Graduate school of Frontier Biosciences

19

0

Assistant Professor

26

Endowed Chair Assistant Professor

1

SA Assistant Professor

5■①Doctoral Program■②Masterʼ s Program

Educational Support Staff

3

Technical Staff

15

Adniministrative Staff

20

Special research students

SA Researcher

37

Research students

3Part-time General & Technical Staff

42

JSPS Research fellows

13●Research Fellows and Research Students
2② South Building① Main Building (left)⑪ Cutting-edge Research
Building for Infectious
Diseases (right)

Total 198
(SA: Specially Appointed)⑦ Central Laboratory for
Biological Hazardous Microbes
and (back)⑤⑥ Animal Resource Center(left,front)

ACCOUNTS
Other Research Grants
99,239Site Area 36,036㎡
Building Area 8,702㎡

Management Expenses Grant (Personnel)
902,862

3

7

Health and Labor Sciences Research Grants
1,775

12

13

6
9

MEXT Research Grants
391,121

8

5

4
10
1

Donations for Research
239,499
Cooperation Research
129,605

11

Total

3,015,216

2

unit: thousand yen

Collaborative Research
757,051

74

Management Expenses Grant(Non-Personnel)
494,064

Gross Floor Area 39,945㎡

Building nameTotal
floor numbers

Building
area (㎡)

Total floorarea(㎡)■①Main Building

7

1,706

6,397

■②South Building

2

409

945

■③North Building

3

492

1,252

■④Annex

2

768

1,548

■⑤Animal Resource Center A

2

640

1,391

■⑥Animal Resource Center B

4

355

1,425

■⑦Central Laboratory for
Biological Hazardous Microbes

3

241

550

■⑧Central Instrumentation Laboratory

2

378

504

■⑨Depository for Dangerous Chemicals

1

160

160

■⑩Integrated Life Science Building

10

1,072

9,258

■⑪Cutting-edge Research Building
for Infectious Diseases

9

973

7,448

■⑫Animal Resource Center C(belonging to IFReC)

4

738

2,482

■⑬IFReC Building

9

770

6,585

75

ACCESSMAP▶Osaka University

Suita CampusrchSupport RIMD Resea in science-

8

Bus Stop
Handai Guchi-Be part of the quest

2
1

3
East Gate

Senri Gate
West Gate

Bus Stop
Handai Honbu
Mae

9

6

4

7

Handai
Byoin Mae5

to find our more

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toational students● Helping intern
ectious Diseases
Inf
al
pic
Tro
g Course on● Helping Trainin
for clinical doctors.
cientistsseminars for non-s
entific lectures and● Organizing sci● Supporting RIM

Kita-Senri

Main Gate

1 Research Institute for Microbial Diseases

4 Graduate School of Medicine

7 Administration Bureau

2 Immunology Frontier Research Center

School of Frontier
5 Graduate
Biosciences

8 The Institute of Scientific

3 Graduate School of Engineering

6 Osaka University Hospital

9 Osaka University Dental Hospital

MINOHCampusKita-Senri
Ishibashi

Hotarugaike

Osaka (Itami)
AirportTOYONAKA
CampusTrain
12-minute walk from “Kita-Senri” Station
on Hankyu Senri Line.

Saito-nishi

SUITA
CampusHandai Byoin Mae

Shibahara Senri-Chuo Yamada

Ibaraki-Shi

IbarakiTenjimbashisuji
6-chomeOsaka(Umeda)Kitahama
Yodo

KyobashiTemma

Nakanoshima yabashi bashi
Namba
CenterKansaiInternational Airport
TennojiTengachaya

nsfer
Credit card, Bank tra
eck the website
For detail please ch

english/ikin.osaka-u.ac.jp/
https://www.miraik

ur doPlease make yonation for follow

ing projects at theWebsitecilities><Institutes and Fa
Diseases
te for Microbialnd■Research Institu
larial Vaccine Fu
Maofnt
Developme■Research and

Bus

Awaji

Nakanoshima

Monorail
20-minute walk from “Handai Byoin Mae” Station
on Osaka Monorail Saito Line.

[How to donate]

MinamiIbaraki

Shin-Osaka

Juso

and Industrial Research

Tsuruhashi

JR Shinkansen
JR Lines
SubwaySubway(mido-suji)
Kita-Osaka Kyuko Line
Osaka Monorail
Hankyu Lines
Other Private Lines
Bus Lines

・From Senri-Chuo Station :Kadoma-shi

5-minute walk from “Handai-Guchi” Bus Stop on Hankyu
Buses heading to “Onohara Higashi” , “Toyokawa-Eki” ,“Fujikasai” .
12-minute walk from “Handai Honbu Mae” Bus Stop on
Hankyu Buses heading to “Handai Honbu Mae” or“Ibaraki Mihogaoka” .

・From Hankyu Ibaraki-shi Station:

12-minute walk from “Handai Honbu Mae” Bus Stopon buses heading to “Handai Honbu Mae”
(via JR Ibaraki Station).
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