Osaka University

Research Institute forMicrobial Diseases

ĳıĲĳȽĳıĲĴContents
History & Outline

Ĳ

Organizationĳ

Former Directors & Professors

ĴDepartment Heads

ĵ

Faculty & StudentsĶ

Research & Activities
Division of Infectious Diseases……………………………………… ķ
Division of Host Defense ……………………………………………… Ĳĵ
Division of Cellular and Molecular Biology …………………………… ĳĵ
Research Center for Infectious Disease Control ……………………… Ĵĳ
Genome Information Research Center ………………………………… ĴķInternational Research Center for Infectious Diseases ……………… ĵĳ

Office of Combined Program on Microbiology and immunology

ĵĺThailand-Japan Research Collaboration Centeron Emerging and Re-emerging Infections

ĶĲ

Mahidol-Osaka Center for Infectious Diseases

Ķĵ

BIKEN Endowed Department of Dengue Vaccine Development

ĶĶGlobal COE ProgramĶķ

Research Facilities

Ķĺ

Immunology Frontier Research CenterķĶ

Accounts & Building AreaķĹ

Map & Accessĸı

Photo by Hiroshi Nojima.

History & Outline

The Research Institute for Microbial Diseases was originally established as a ﬁve-department “Research

History & Outline

Research Institute for Microbial Diseases
Center for Communicable Diseases” in February, 1934 after a donation from Mr. Gendo Yamaguchi at
Dojima in Osaka City. At that time, the Institute was located on Nakanoshima campus of Osaka Medical
School, but then became a part of Osaka University in September, 1934. It was moved to its present site on
Suita campus in 1967, in accordance with the university's relocation program.
Until 1993, the Institute had grown to include nineteen departments spanning a wide range of diverse
subjects, such as infectious disease, immunology, cancer and molecular biology. Additionally, it had three
special research facilities, a central laboratory and a library. In 1997, the Research Center for Emerging
Infectious Diseases was added to the Institute. In 2005, the Institute was reorganized to constitute three
Research divisions; the “Division of Infectious Disease”, the “Division of Host Defence” and the “Division of
Cellular and Molecular Biology”, which represent 15 departments together with three attached centers for
specialized research on infectious disease and genome information. Moreover, the Research Collaboration
Center on Emerging and Re-emerging Infections was founded in Bangkok in collaboration with the National
Institute of Health, Thailand, to defend people against possible emerging and re-emerging infections.
Basic research on infectious disease, immunology, and cell biology is the Institute's principal focus. The
results of research in these ﬁelds at the Institutehave contributed considerably to the diagnosis,
prevention, and treatment of infectious diseases,
immunological diseases and cancer, as well as toprogress in basic biomedical science. The Institutewas selected as one of “the 21st century center of
excellence programs” on the theme of “combined
program on microbiology and immunology” in
2003. Also, it was certiﬁed as “joint usage /research center” by MEXT (the Ministry of education, culture, sports, science and technology) in
The copper plate recalling that the Research Institutefor Microbial Diseases was originally establishedby a donation from Mr. Gendo Yamaguchi.
(At the entrance hall of the main building)2009, and started the mission in April, 2010. Moreover, the institute newly established the “Center for
Genetic Analysis of Biological Responses” toconserve gene resources and to protect intellectual
property.

Mr. Gendo Yamaguchi
Born in Onomichi City in 1863 as the eldest
son of a physician, he moved to Osaka at theage of 15 and became one of western Japanʼsmost successful businessmen. He retired in
1917, and devoted the rest of his life toreligion and the tea ceremony. He donatedmost of the proceeds of his estate to public
enterprise, shrines and temples.

The Institute accepts and trains Master and Ph.D.
candidates in the medical and biological sciences. At
present, more than 200 full, associate, and assistantprofessors, research associates, graduate students
and research fellows pursue studies in microbiology, oncology and molecular biology in state of theart facilities at the Institute.

Ĳ

Organization
Director
Faculty Meeting

Delegate Assembly

Research Division
Division of Infectious Diseases
Department of Molecular Bacteriology
Department of Viral lnfections
Department of Molecular Virology
Department of Pharmacotherapy
Department of Immunoparasitology
Division of Host Defense
Department of Molecular Immunology
Department of Immunoregulation
Department of Host Defense
Department of Cell Biology
Department of Immunochemistry
Division of Cellular and Molecular Biology
Department of Molecular Microbiology
Department of Molecular Genetics
Department of Oncogene Research
Department of Signal Transduction
Department of Cellular Regulation

Special Research Facilities
Research Center for Infectious Disease Control
Department of Bacterial Infections
Department of Molecular Protozoology
Department of Virology
Genome Information Research Center
Department of Experimental Genome Research
Department of Genome Informatics
Department of Infection Metagenomics
International Research Center for Infectious Diseases
Department of Special Pathogens
Department of Infectious Disease Control
Pathogenic Microbes Repository Unit
Animal Resource Center for Infectious Diseases
DNA-chip Development Center for Infectious Diseases
Center for Genetic Analysis of Biological Responses

Office of Combined Program on Microbiology and Immunology
Research Promotion Group
Education Promotion Group

Research Collaboration Center in Overseas
Thailand-Japan Research Collaboration Center on Emerging and Re-emerging Infections
Section of Bacterial Infections
Section of Viral Infections
Mahidol-Osaka Center for Infectious Diseases
BIKEN Endowed Department of Dengue Vaccine Development

Common Research Facilities
Central Instrumentation Laboratory
Radioisotope Laboratory
Central Laboratory for Biological Hazardous Microbes
Library

Administration
General Affairs Section
Accounting Section
Research Cooperation Section

World Premier International Research Center
Immunology Frontier Research CenterĳFormer Directors & Professors<DVKLUR.RWDNH0'3URIHVVRUí

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Organization, Former Directors & Professors

Former Directors

Former Professors
Yashiro Kotake, M.D., Professor

Toshio Miwatani, M.D., Professor

Sadao Yoshida, M.D., Professor

Michiaki Takahashi, M.D., Professor

Arao Imamura, M.D., Professor

Hajime Fujio, M.D., Professor

Yukichi Satani, M.D., Professor

Tetsuo Taguchi, M.D., Professor

Tenji Taniguchi, M.D., Professor

Aizo Matsushiro, D.Sc., Professor

Kota Sera, M.D., Professor

Atsuo Nakata, D.Sc., Professor

Tatsunori Masayama, M.D., Professor

Hiroto Okayama, M.D., Professor

Shohei Otani, M.D., Professor

Kumao Toyoshima, M.D., Professor

Teishiro Seki, M.D., Professor

Teruo Kitani, M.D., Professor

Masami Suda, M.D., Professor

Shin-ichiro Takai, M.D., Professor

Kaoru Morishita, M.D., Professor

Morihiro Matsuda, M.D., Professor

Hisashi Yamaguchi, M.D., Professor

Takashi Kurimura, M.D., Professor

Tsunesaburo Fujino, M.D., Professor

Koichi Yamanishi, M.D., Professor

Masakazu Ito, M.D., Professor

Akira Hakura, D.Sc., Professor

Juntaro Kamahora, M.D., Professor

Tetsu Akiyama, D.Sc., Professor

Shinji Nishimura, M.D., Professor

Takeshi Kurata, M.D.,D.Med.Sci., Professor

Mitsuhiko Kato, M.D., Professor

Shigeharu Ueda, M.D.,D.Med.Sci., Professor

Masahiko Yoneyama, M.D., Professor

Kazunori Shimada, M.D.,D.Med.Sci., Professor

Shigeru Shiba, M.D., Professor

Chihiro Sasakawa, M.D., Professor

Shozo Inoki, M.D., Professor

Akio Sugino, D.Sci., Professor

Mitsuo Hori, M.D., Professor

Hiroshi Kiyono, D.D.S.,Ph.D., Professor

Yoshiomi Okuno, M.D., Professor

Yoshitake Nishimune, M.D., Professor

Shigeyuki Ishigami, M.D., Professor

Toru Nakano, M.D., D.Med. SC., Professor

Tsunehisa Amano, M.D., Professor

Hideo Shinagawa, D.Sc., Professor

Junichi Kawamata, M.D., Professor

Shin-ichi Tamura, Ph.D., Professor

Yoshio Okada, M.D., Professor

Michiyuki Matsuda, M.D., D.Med. SC., Professor

Mitsuo Torii, D.Sc., Professor

Takeshi Honda, M.D., Ph. D., Professor

Konosuke Fukai, M.D., Professor

Naoyuki Taniguchi, M.D., Ph. D., Professor

Tatsuo Mori, M.D., Professor

Tamotsu Yoshimori, M.D., Ph. D., Professor

Tonetaro Ito, M.D., Professor

Kazuyuki Tanabe, M.D., Ph. D., Professor

Takeo Kakunaga, D.Pharm., Professor

Fumio Imamoto, D.Sc., Professor

Shiro Kato, M.D., Professor

Atsushi Kumanogo, M.D., Ph. D., Professor

Toshio Nakabayashi, M.D., Professor

Kazunori Oishi, M.D., Ph. D., SA Professor

Takahisa Yamanouchi, M.D., Professor

Masanori Kameoka, Ph. D., SA ProfessorĴDepartment Heads
2012,04,01

Director
Vice Director
Division of Infectious Diseases
Department of Molecular Bacteriology
Department of Viral Infections
Department of Molecular Virology
Department of Pharmacotherapy
Department of Immunoparasitology
Division of Host Defense
Department of Molecular Immunology
Department of Immunoregulation
Department of Host Defense
Department of Cell Biology
Department of Immunochemistry
Division of Cellular and Molecular Biology
Department of Molecular Microbiology
Department of Molecular Genetics
Department of Oncogene Research
Department of Signal Transduction
Department of Cellular Regulation

Professor Eisuke Mekada Ph.D.
Professor Yoshiharu Matsuura D. V. M., Ph.D.
Professor Yasuhiko Horiguchi D. V. M., Ph.D.
Professor Tatsuo Shioda Ph.D.
Professor Yoshiharu Matsuura D. V. M., Ph.D.
Associate Professsor Masahiro Yamamoto Ph.D.
Professor Hitoshi Kikutani M. D., Ph.D.
Professor Taroh Kinoshita Ph.D.
Professor Shizuo Akira M. D., Ph.D.
Professor Eisuke Mekada Ph.D.
Professor Hisashi Arase M. D., Ph.D.
Professor Hiroshi Nojima Ph.D.
Professor Masato Okada Ph.D.
Professor Nobuyuki Takakura M. D., Ph.D.
Professor Hiroaki Miki Ph.D.

Research Center for Infectious Disease Control
Department of Bacterial Infections
Department of Molecular Protozoology
Department of Viorogy
Genome Infomation Research Center
Department of Experimental Genome Research
Department of Genome Informatics
Department of Infection Metagenomics
International Research Center for Infectious Diseases
Department of Special Pathogens
Laboratory of Clinical Research on Infectious Diseases
Laboratory of Infection Cell Biology
Laboratory of Viral Infection
Department of Infectious Disease Control
Laboratory of Genomic Research on Pathogenic Bacteria
Laboratory of Malariology
Laboratory of Combined Research on Microbiology and Immunology

Head, Professor Toshihiro Horii Ph.D.

Animal Resource Center for Infectious Diseases
DNA-chip Development Center for Infectious Diseases
Center for Genetic Analysis of Biological Responses

Head, Professor Masaru Okabe Ph.D.
Head, Professor Hiroshi Nojima Ph.D.
Head, Professor Masaru Okabe Ph.D.

Office of Combined Program on Microbiology and Immunology
Research Promotion Group
Education Promotion Group

Head, Director Eisuke Mekada Ph.D.
Associate Professsor Yoshiko Murakami M. D., Ph.D.
Associate Professsor Hodaka Fujii M. D., Ph.D.

Thailand - Japan Research Collaboration Center on Emerging andRe-emerging Infections
Section of Bacterial Infections
Section of Viral Infections

Head, SA Professor Naokazu Takeda Ph.D.

Mahidol-Osaka Center for Infectious Diseases

Head,Professor Yoshiharu Matsuura D.V.M., Ph.D.
SA Associate Professor Tamaki Okabayashi D.V.M., Ph.D.

BIKEN Endowed Department of Dengue Vaccine Development

Endowed Chair Professor Eiji Konishi Ph.D.

Central Instrumentation Laboratory
Radioisotope Laboratory
Central Laboratory for Biological Hazardous Microbes
Library

Head,Professor Masato Okada Ph.D.
Head,Professor Masato Okada Ph.D.
Head, Professor Tatsuo Shioda Ph.D.
Head, Professor Toshihiro Horii Ph.D.

Administration

Head, Katsumi Uedono

Professor Toshihiro Horii Ph.D.
Professor Kazuyoshi Ikuta Ph.D.
Head, Professor Teruo Yasunaga Ph.D.
Professor Masaru Okabe Ph.D.
Professor Teruo Yasunaga Ph.D.
Head, Professor Toshihiro Horii Ph.D.
SA Professor Yukako Fujinaga Ph.D.
SA Associate Professor Eiji Morita Ph.D.
SA Professor Tetsuya Iida Ph.D.
Associate Professor Hiroki Nagai Ph.D.

SA Professor Shigeyuki Hamada D.D.S., Ph.D.
SA Professor Naokazu Tokeda Ph.D.

＊SA : Specially Appointed
ĵFaculty & Students

2012,04,01
Professor

16

Endowed Chair Professor

1

SA Professor

4

Associate Professor

14

Endowed Chair Associate Professor

0

SA Associate Professor

4

SA Lecturer

4

Assistant Professor

30

Endowed Chair Assistant Professor

1

SA Assistant Professor

10

Educational Support Staff

3

Technical Staff

3

Administrative Staff

21

SA Researcher

47

Part-time General & Technical staff

48

Department Heads, Faculty & Students

Staff

206

Total
SA：Specially Appointed

Graduate Students
2012,04,01

Doctor Course

Master Course

Graduate School of Medicine

213

Graduate School of Science3

11

Graduate School of Pharmaceutical Science01

Graduate School of Dentistry00

Graduate School of Frontier Biosciences

8

4

32

19

Total

Research Fellows & Research Students
2012,04,01

Special research students0

Research Students

7

Visiting Research Scholars

1

JSPS Research Fellows

3

Total

11
Ķ

Research & Activities

Division of Infectious Diseases

Department of Molecular Bacteriology
Research Group

Professor
Assistant Professor
Assistant Professor
SA Researcher
SA Researcher

Yasuhiko Horiguchi, D.V.M., D.Agr.Sci.
Shigeki Kamitani, D.M.Sc.
Hiroyuki Abe, Ph.D.
Aya Fukui, Ph.D.
Hirono Toshima, Ph.D.

Research Projects
The objective of this department is to understand the molecular mechanisms by which pathogenic bacterial virulence factors
affect host cell functions. Our present research interests include:(1) Analysis of the structure and function of bacterial protein toxins
Bacterial protein toxins, which are the most poisonous substances on the earth, act specifically on a particular cell or
biomolecule. To understand how bacterial toxins act so powerfully and specifically, we are analyzing their effects on the hostat the systemic, tissue, cellular, and molecular levels. The toxins currently under investigation include Bordetella
dermonecrotic toxin, Pasteurella toxin, Clostridium perfringens enterotoxin, and Escherichia coli cytotoxic necrotizing factor.
We are also analyzing the steric structure and molecular localization of the functional domains of these toxins. Together, theseapproaches will help to clarify the structure and function of these bacterial toxins.
(2) Analysis of the pathogenesis of whooping coughBordetella pertussis is a bacterial pathogen that infects the human respiratory tract and causes whooping cough, which ischaracterized by paroxysmal cough. Two important questions remain in terms of the pathogenesis of B. pertussis infection:
Why does B. pertussis infect humans but not other mammals? How does the bacteria cause the paroxysmal cough? We are
examining the pathology of the disease and function of bacterial virulence factors using an animal model of the infection.

Fig. 1: Bacterial protein toxins with various activities that influence particular cellular functions.
Many bacterial protein toxins exert their toxicity by modifying important functions of the host cells.
The relevant physiological functions of the cells can be determined by dissecting the actions of thebacterial toxins.

ķResearch & Activities

Fig. 2: Overall structure of Clostidium perfringens enterotoxin

Fig. 3: B. pertussis, B. parapertussis, and B. bronchiseptica areclosely related pathogenic bacteria. B. bronchiseptica with thelargest genome shows the broadest host range, whereas B. pertussis
with the smallest genome shows the narrowest range. B. pertussis
is believed to have evolved from a lineage of B. bronchiseptica
through deletion and/or translocation of a large number of genes.

Recent Publications
1. Horiguchi, Y. "Swine Atrophic Rhinitis Caused by Pasteurella Multocida Toxin and Bordetella Dermonecrotic
Toxin." Curr Top Microbiol Immunol (2012).
2. Kitadokoro, K., K. Nishimura, S. Kamitani, A. Fukui-Miyazaki, H. Toshima, H. Abe, Y. Kamata, Y. Sugita-Konishi,
S. Yamamoto, H. Karatani, and Y. Horiguchi. "Crystal Structure of Clostridium Perfringens Enterotoxin Displays
Features of {Beta}-Pore-Forming Toxins." J Biol Chem 3;286(22) (2011):19549-55.
3. Kamitani, S., S. Ao, H. Toshima, T. Tachibana, M. Hashimoto, K. Kitadokoro, A. Fukui-Miyazaki, H. Abe, and Y.
Horiguchi. "Enzymatic Actions of Pasteurella Multocida Toxin Detected by Monoclonal Antibodies Recognizing the
Deamidated Alpha Subunit of the Heterotrimeric Gtpase Gq." Febs J 278, no. 15 (2011): 2702-12.
4. Kimura, J. , H. Abe, S. Kamitani, H. Toshima, A. Fukui, M. Miyake, Y. Kamata, Y. Sugita-Konishi, S.
Yamamoto, and Y. Horiguchi. "Clostridium Perfringens Enterotoxin Interacts with Claudins Via Electrostatic
Attraction." J Biol Chem 285, no. 1 (2010): 401-08.
5. Kamitani, S, K. Kitadokoro, M. Miyazawa, H. Toshima, A. Fukui, H. Abe, M. Miyake, and Y. Horiguchi.
"Characterization of the Membrane-Targeting C1 Domain in Pasteurella Multocida Toxin." J Biol Chem 285, no. 33(2010): 25467-75.
ĸResearch & Activities

Division of Infectious Diseases

Department of Viral Infections
Research Group

Professor
Assistant Professor
Assistant Professor

Tatsuo Shioda, D.Med.Sc.
Jun-ichi Sakuragi, D.Med.Sc.
Emi E. Nakayama, M.D., D.Med.Sc.

The main focus of this department is to elucidate the molecular mechanisms of viral diseases, including humanimmunodeficiency virus (HIV). The following projects are currently underway.
(1) Antiretroviral factors
HIV does not establish a productive infection in any monkey other than the chimpanzee. This observation is thought to be due
WRWKHIDFWWKDWLQKLELWRUVLQVLPLDQO\PSKRF\WHVDFWDWWKHHDUO\VWDJH UHYHUVHWUDQVFULSWLRQ RIYLUDOLQIHFWLRQ7RGDWH75,0Įand cyclophilin A have been identified as such restriction factors. We have shown that differences in the amino acid sequencesin the C-terminal domain of TRIM5Į of different monkey species affect the species-specific restriction of retrovirus infection(Fig. 1, left). We also have found that sequence variations in the N-terminal half of the viral capsid protein (Fig. 1, right)
determine viral sensitivity to TRIM5Į-mediated restriction, which indicates that there is an interaction between TRIM5Įandthe virus capsid. We found that HIV-2 replication levels in infected individuals were associated with capsid variations andsuggested that viral sequence analysis can predict AIDS progression. Furthermore, we succeeded in improving thesimian-tropic HIV-1 virus and the methods of monkey genome analysis. These new developments will greatly facilitate thegeneration of an HIV-1 animal model, which would be a highly useful tool in research aiming to understand AIDS
pathogenesis and to develop an effective vaccine. We are seeking to identify the binding surface between the viral capsid
protein and TRIM5Į because this information may be useful for the development of new antiretroviral drugs.

Fig. 1. Structural models of TRIM5Į (left) and the viral capsid (right).

ĹResearch & Activities(2) Host factors that participate in antiretroviral drug side-effects
Human genetic variations affect differences in human phenotypes. In collaboration with Thai groups, we are analyzing therelationship between human genomic variation and antiretroviral therapy side-effects, with the aim of establishing “tailor-madetherapies” that will improve the quality of life of HIV-infected patients.
(3) Analysis of HIV-1 genome RNA dimerization
The genome of a retrovirus such as HIV-1 is a single-stranded, positive-sense RNA. The viral genome always exists as a dimerin virions. Genome dimerization plays important roles in various stages of the viral life cycle, including genome packaging andreverse transcription, and genome recombination processes involved in viral diversification. We have constructed HIV-1 Gag
cleavage site mutants to enable the steady-state observation of virion maturation steps to study Gag processing, RNAdimerization, virion morphology, and infectivity precisely. We revealed that the maturation of the viral RNA/protein plays
critical roles for the acquisition of viral infectivity (Fig. 2A). We previously identified the region that is necessary andsufficient for HIV-1 genome dimerization in the virion (DLS: Dimer Linkage Structure). By further detailing the fine mappingof DLS, we discovered the possibility of a long-range interaction, which had never been reported previously. We performed
computer-assisted structural modeling and obtained new 3D models of the HIV-1 DLS that revealed a unique pseudoknot-like
conformation (Fig. 2B). Because this conformation appears to be thermodynamically stable, forms a foundational skeleton for
the DLS, and sterically restricts the spontaneous diversification of DLS conformations, its unique shape may potentially serve
as a novel target for anti-HIV-1 therapies.

Fig. 2 HIV-1 genome RNA dimerization.

Recent Publications
1. A proposal for a new HIV-1 DLS structural model. Sakuragi JI, Ode H, Sakuragi S, Shioda T, Sato H. Nucleic Acids
Res. In press.
2. Role of human TRIM5Į in intrinsic immunity. Nakayama EE, Shioda T. Front Microbiol.2012;3:97.
3. TRIM5Į and Species Tropism of HIV/SIV. Nakayama EE, Shioda T. Front Microbiol. 2012; 3:13.
4. The relationship between HIV-1 genome RNA dimerization, virion maturation and infectivity. Ohishi M, Nakano T,
Sakuragi S, Shioda T, Sano K, Sakuragi JI. Nucleic Acids Res. 2011 Apr;39(8):3404-17.
5. HLA-Cw*04 allele associated with nevirapine-indued rash in HIV-infected Thai patients. Likanonsakul S,
Rattanatham T, Feangvad S, Uttayamakul S, Prosithsirikul W, Tunthanathip P, Nakayama EE, Shioda T. AIDS Res
Ther. 2009 Oct 21;6:22.
ĺResearch & Activities

Division of Infectious Diseases

Department of Molecular Virology
Research Group

Professor
Assistant Professor
Assistant Professor
SA Assistant Professor
SA Researcher
SA Researcher
Postdoctoral Fellow

Yoshiharu Matsuura
Toru Okamoto
Takasuke Fukuhara
Hiroto Kambara
Chikako Ono
Takashi Motomura
Hiroshi Kato

DVM, PhD
PhD
MD, PhD
PhD
PhD
MD, PhD
DVM, PhD

We are working towards understanding the molecular mechanisms of entry, replication, immune escape, and pathogenesis ofhepatitis C virus (HCV) and the development of a novel viral vector for gene delivery.
1. Studies on the molecular biology of HCV
replication and pathogenesis
More than 3% of the worldwide population is
infected with HCV, and 80% of these individuals
will develop persistent infection. Persistent HCV
infection often leads to chronic hepatitis, cirrhosis,
and hepatocellular carcinoma. The incidence ofhepatitis C has significantly decreased since theintroduction of a screening system for anti-HCV
antibodies in 1999; however, more than two
million people have already been infected with
HCV in Japan. Use of combination therapy withSHJ\ODWHG,)1ĮULEDYLULQDQG+&9SURWHDVH
inhibitor has greatly increased the proportion ofpatients achieving a sustained virological response.
Nevertheless, 30% of patients with HCV genotype
1 exhibit no response to the combination therapy. Although IL28B genetic polymorphism, viral RNA mutations, andexpression of ISGs, including IP-10, are associated with the response to IFN-based antiviral therapy, the mechanism has notbeen elucidated.
HCV exhibits quasispecies heterogeneity. As such, its infection mechanisms are difficult to assess by surrogate systems, suchas pseudotype and recombinant viruses derived from a single HCV clone. In vitro replication of genotype 2a HCV (HCVcc)
was recently established. Several receptor candidates, including hCD81, SR-BI, Claudin1, and Occludin, for HCV entry wereidentified by using the surrogate viruses and HCVcc. However, sera from persistently infected individuals show a high level ofneutralization antibodies to the artificial viruses, suggesting that these antibodies do not play a crucial role in HCV clearance.
HCV particles are internalized into cells throughendocytosis. HCV NS3/4A protease was shown tocleave adaptor molecules involved in the TLR- and
RIG-I-dependent signaling pathways. After uncoating, aviral RNA is translated into a large precursor
polyprotein composed of 3,000 amino acids. This viral
polyprotein is cleaved by signal peptidase (SP), signal
peptide peptidase (SPP), and viral-encoded proteases,
resulting in at least 10 viral proteins. The liver-specificmicroRNA miR-122 enhances translation through agenetic interaction with the 5’ UTR of HCV-RNA.
However, the molecular mechanism of the enhancement
has not been clarified. The formation of amultichaperone complex with NS5A is essential for the
replication of viral RNA. Viral RNA and core proteins
Ĳı

Research & Activities

are recruited around a lipid droplet, and viral capsids are formed.
Although novel innovative agents have shown significant antiviral activity in patients with HCV, drug-resistant viruses easily
emerge. Therefore, host factors indispensable for HCV replication are ideal targets for the development of new therapeutics forchronic hepatitis C with a broad spectrum and a low possibility of emergence of breakthrough viruses against antiviral drugs.
HCV belongs to the family of Flaviviridae, which includes flavivirus such as Japanese encephalitis virus (JEV) that has arobust cell culture system and a small animal model. We are investigating the replication and pathogenesis of JEV as asurrogate model for HCV.
2. Development of baculoviral vectors
Viral vectors are essential tools for studies of replication-deficient viral infectious diseases, such as HCV. Furthermore, thedevelopment of novel viral vectors is essential for future gene therapy. We are studying the baculovirus Autographacalifornica nucleopolyhedrovirus (AcNPV) as a versatile viral vector for gene delivery in vitro and in vivo. AcNPV is an
insect virus possessing a 134 kb double-stranded circular DNA genome. Due to its strong promoters, baculovirus is commonlyused as a tool for the large-scale production of recombinant protein in insect cells. Baculovirus is capable of entering various
mammalian cells to facilitate the expression of foreign genes under the control of mammalian promoters without replication of
the viral genome. Therefore, baculovirus is a useful viral vector, not only for the abundant expression of foreign genes in insect
cells, but also for efficient gene delivery to mammalian cells.
AcNPV has a number of unique beneficial properties as a viral vector, including a large capacity for foreign geneincorporation, easy manipulation, and replicationcompetence in insect cells combined with incompetence inmammalian cells. Therefore, the possibility of generating
replication-competent revertants expressing baculoviralgene products, which can often lead to harmful immune
responses against mammalian cells, is significantly lower
than that with other viral vectors currently in use.
Furthermore, intranasal inoculation with AcNPV induces astrong innate immune response, protecting mice fromlethal challenges of influenza viruses.
We demonstrated that the internalization of viral DNA via
membrane fusion by the envelope glycoprotein in theendosome is required for the induction of innate immune
response by AcNPV through a TLR9/MyD88-dependent
pathway. This finding raises the possibility that AcNPV
may be harnessed therapeutically to induce the hostimmune response against various infectious diseases,
especially those caused by pathogens invading from therespiratory tract.
Recent Publications
1. Abe T, Fukuhara T, Wen X, Ninomiya A, Moriishi K, Maehara Y, Takeuchi O, Kawai T, Akira S, Matsuura Y.
CD44 participates in the IP-10 induction in cells replicating HCV RNA through an interaction with TLR2 andhyaluronan. J Virol. 2012.
2. Kataoka C, Kaname Y, Taguwa S, Abe T, Fukuhara T, Tani H, Moriishi K, Matsuura Y. Baculovirus
GP64-mediated entry into mammalian cells. J Virol. 2012 Mar;86(5):2610-20.
3. Kambara H, Fukuhara T, Shiokawa M, Ono C, Ohara Y, Kamitani W, Matsuura Y. Establishment of a novel
permissive cell line for the propagation of hepatitis C virus by expression of microRNA miR122. J Virol. 2012
Feb;86(3):1382-93.
4. Taguwa S, Kambara H, Fujita N, Noda T, Yoshimori T, Koike K, Moriishi K, Matsuura Y. Dysfunction of
autophagy participates in vacuole formation and cell death in cells replicating hepatitis C virus. J Virol. 2011 Dec;85(24):13185-94.
5. Katoh H, Mori Y, Kambara H, Abe T, Fukuhara T, Morita E, Moriishi K, Matsuura Y. Heterogeneous nuclear
ribonucleoprotein A2 participates in the replication of Japanese encephalitis through an interaction with viralproteins and RNA. J Virol. 2011 Nov;85(21):10976-88.

ĲĲ

Research & Activities

Division of Infectious Diseases

Department of Immunoparasitology
Research Group

Associate Professor
Assistant Professor

Ph.D. Masahiro Yamamoto
Ph.D. Miwa Sasai

Our laboratory studies immmunoparasitology using an apicomplexan protozoan parasite called Toxoplasma gondii as a
research model to analyze the “host-to-pathogen” interaction.
1) What is Toxoplasma?
Toxoplasma gondii (T. gondii) is a eukaryotic pathogen that causes life-threatening toxoplasmosis, a condition that includes
encephalitis in immunocompromised individuals, such as those suffering from AIDS or being treated by chemotherapy, andcongenital diseases following primary infection during pregnancy in humans and animals. T. gondii is an obligate intracellular
eukaryotic parasite capable of proliferating exclusively inside a parasitophorous vacuole, which is formed during host cell
invasion.
Taxonomically, T. gondii belongs to the Apicomplexa phylum that is defined by the presence of an apical complex including
secretory organelles, such as conoids, micronemes, and rhoptries (Figure 1A). Among them, the large bulb-shaped organelles
called rhoptries contain various proteins that are secreted into the host cytoplasm or in the forming parasitophorous vacuole
during parasite entry to co-opt the host cell for growth and survival (Figure 1B and C).
2) Downregulation of host immune responses by T. gondii.
T. gondii is divided into three major lineages (types I, II, and III). Stimulation of innate immune cells, such as macrophages
and dendritic cells, by T. gondii-derived components results in strong production of proinflammatory cytokines. However,“live” infection with type I or type III, but not type II, parasites reduces cytokine production. We revealed that a single aminoacid substitution of the kinase domain of a rhoptry protein ROP16 is one molecular mechanism by which type II parasites failto suppress the host innate immune response (Figure 2). In terms of virulence, in mice, the type III parasite is avirulent
compared with the type I and type II parasites. Another ROP16 family member, ROP18, was identified as the gene responsible
for the genotype-dependent virulence mechanism. We revealed that ROP18 targets a host transcription factor, ATF6b, anddegrades the protein in a proteasome-dependent fashion (Figure 3).
Effector proteins, such as ROP16 and ROP18, constitute a large family of proteins. However, the functions of most of theeffectors released by this parasite in host cells remain uncertain. Moreover, the host target molecules are also unknown.
Therefore, we are studying the role of the T. gondii-derived effector molecules by generating and analyzing gene-targetedparasite lines. We are trying to uncover the strategy used by the parasite to suppress the host immune response, which may
result in the identification of novel immunological concepts.

Figure 1 Toxoplasma gondii and the rhoptry protein(A) A schematic figure of T. gondii. The apical complex includes rhoptries (red). (B) Electron micrograph of T. gondii residing ina parasitophorous vacuole (blue) in an infected host cell. (C) Immunofluorescent image of T. gondii showing detection of therhoptry protein ROP18.

Ĳĳ

Research & Activities

Figure 2. Single amino acid substitution of ROP16 determines
T. gondii infection-induced suppression of innate immune response.

))LJXUH$7)ȕ.2PLFHDUHKLJKO\VHQVLWLYHWRURS.2SDUDVLWHV
Luciferase-expressing T. gondii lacking ROP18 were infected in
ZLOGW\SHRU$7)ȕ.2PLFH/XFLIHUDVHHPLVVLRQZDVPRQLWRUHG
with a live imaging analyzer.

Recent Publications
1. Yamamoto M7DNHGD.,QKLELWLRQRI$7)ȕGHSHQGHQWKRVWDGDSWLYHLPPXQHUHVSRQVHE\D7R[RSODVPD
virulence factor ROP18. Virulence. (2012) In press.
2. Yamamoto M0D-60XHOOHU&.DPL\DPD16DLJD+.XER(.LPXUD72NDPRWR72NX\DPD0.D\DPD+
1DJDPXQH.7DNDVKLPD60DWVXXUD<6ROGDWL)DYUH'7DNHGD.$7)ȕLVDKRVWFHOOXODUWDUJHWRIWKH
Toxoplasma gondii virulence factor ROP18. J Exp Med. (2011) 208:1533-1546.
 6D\DPD. Yamamoto M* (* Equal contribution), Shirakata Y, Hanakawa Y, Hirakawa S, Dai X, Tohyama M,
7RNXPDUX66KLQ066DNXUDL+$NLUD6+DVKLPRWR.(SRO\XELTXLWLQFRQMXJDWLQJHQ]\PH8EFLQ
keratinocytes is essential for epidermal integrity. J Biol Chem. (2010) 285:30042-9.
4. Yamamoto M6WDQGOH\'07DNDVKLPD66DLJD+2NX\DPD0.D\DPD+.XER(,WR+7DNDXUD00DWVXGD
76ROGDWL)DYUH'7DNHGD.$VLQJOHSRO\PRUSKLFDPLQRDFLGRQ7R[RSODVPDJRQGLLNLQDVH523GHWHUPLQHV
the direct and strain-specific activation of Stat3. J Exp Med. (2009) 206: 2747-2760.
5. Yamamoto M8HPDWVX62NDPRWR70DWVXXUD<6DWR6.XPDU+6DWRK76DLWRK77DNHGD.,VKLL.-
7DNHXFKL2.DZDL7$NLUD6(QKDQFHG7/5PHGLDWHG1),/GHSHQGHQWJHQHH[SUHVVLRQE\7ULEGHILFLHQF\J Exp Med. (2007) 204:2233-9.
ĲĴDivision of Host Defense

Research & Activities

Department of Molecular Immunology
Research Group

Professor
Associate Professor
Assistant Professor
SA Researcher
SA Researcher
SA Researcher
SA Researcher

Hitoshi Kikutani, M.D., Ph.D.
Teruhito Yasui, Ph.D.
Shuhei Sakakibara, Ph.D.
Olivia Simma, Ph.D.
Kentaro Morita, Ph.D.
Takeharu Minamitani, Ph.D.
Chiau-Yuang Tsai, Ph.D.

Molecular mechanisms involved in the regulation of immune responses
T-cells are activated by antigenic peptides bound to the MHC complex on antigen-presenting cells. Once activated, the T-cells
differentiate into functional helper or effector T-cells. Antigen-stimulated B-cells differentiate into antibody-forming cells or
memory B-cells with the help of antigen-specific T-cells. Thus, T- and B-cell differentiation processes require physiologicalinteractions between T-cells and antigen-presenting cells and between T-cells and B-cells, respectively. Such cell–cellinteractions are mediated by various costimulatory molecules, including CD40, CD40 ligand, B-7, and CD28. In addition, it
was recently revealed that several members of the Semaphorin family are crucially involved in immune cell interactions. We
are currently studying the roles of these molecules in the regulation of immune responses.
A) Mechanisms of immune regulation by Semaphorin molecules
The Semaphorin family contains axonal guidance factors that function during neuronal development. However, studies by ourlaboratory have shown that several Semaphorin molecules play crucial roles at various stages of immune responses (Figure 1).
For instance, Sema4D/CD100 is involved in the activation of B-cells and dendritic cells, whereas Sema4A plays roles in T-cellpriming and Th1 differentiation. The interaction between Sema6D and its receptor Plexin-A1 mediates cellular immune
responses by activating dendritic cells and bone homeostasis by inducing osteoclastgenesis. Furthermore, we recently
demonstrated that Sema7A on activated T-cells activates macrophages to produce inflammatory cytokines and triggers
LQIODPPDWRU\UHVSRQVHVWKURXJKĮȕLQWHJULQ )LJXUH B) Elucidation of the molecular mechanism by which B-cells survive and differentiate into effecter cells
B-cells differentiate into antibody-secreting cells and memory B-cells to respond to nonself antigens. Signals through the B-cell antigen
receptor (BCR) and members of the TNF receptor family, such as CD40 and BAFF-R, on the B-cell surface are required for B-cell
activation, survival, and differentiation. Our group has been studying the roles of the molecules that are involved in the signaling
pathways downstream of BCR and CD40. We recently identified a PKC-related serine/threonine kinase, PKN1, which is associatedwith the TRAF family, as a negative regulator of Akt in BCR signaling. Our findings showed that the PKN1-mediated regulation of
Akt is critical for the selection of high-affinity B-cells.

Fig.1 Representative immune Semaphorins
Semaphorins and their receptors have been shown to play roles
in immune regulation by our research group

Fig.2 $FFXPXODWLRQRI6HPD$DQGĮ,QWHJULQLQ
immunological synapse by the interaction between T
cells and macrophages.

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infectious mononucleosis in healthy donors and proliferative disorders in immune-suppressive conditions induced by aging,
immunesuppresant therapy, and HIV infection. EBV infection is sometimes associated with B-cell malignancies, such as
Burkitt’ s lymphoma and Hodgkin’ s lymphoma, and with some autoimmune diseases, such as systemic lupus erythematosus(SLE) and multiple sclerosis (MS). By studying EBV biology, we hope to determine how EBV leads to autoimmune diseases
and human carcinogenesis. The outcomes from this study may contribute to the development of novel therapies for
EBV-associated disorders (Figure 3).
ĲĵFig.3 EBV and host immune system
The mechanism by which EBV induces human B cell growthtransformation is closely associated with the vulnerability of thehost immune system.

Research & Activities

A) Signaling pathways by which EBV latent membrane proteins and downstream molecules induce the cell growth andsurvival of infected cells
EBV infection elicits growth transformation and the immortalization of B-cells and epithelial cells. Recently, it was alsosuggested that EBV infects T-cells and NK-cells, leading to nasal lymphoma and chronic active EBV infection (CAEBV)
associated with hemophagocytic syndrome. How does EBV transform human cells? Of the EBV latent gene products, ourgroup is focusing on two latent membrane proteins, LMP1 and LMP2a, which are involved in EBV-induced transformation by
mimicking CD40 and BCR signals, respectively. We are currently studying the mechanisms by which these molecules modify
B-cell function and induce EBV-related disorders by using knock-in mice in which either or both proteins can be expressed ina cell-specific manner (Figure 4).

Fig.4 Schematic structure of EBV LMP1 and
LMP2a.

% (OXFLGDWLRQRIWKHYLUXVKRVWLQWHUDFWLRQXVLQJDPRXVHPRGHORIȖKHUSHVYLUXVLQIHFWLRQZLWK0+9
Intranasal inoculation of mice with MHV68, which is closely related to EBV,
results in an acute productive infection of the lung and latent infection inseveral cell types, including B-cells and macrophages. Because of the limited
susceptibility of experimental animals to EBV infection, MHV68 infection ofmice should be a useful animal model for understanding EBV pathogenesis. In
fact, MHV68 infection induces emerging autoreactive antibody that can trigger
a complement-dependent inflammation. We have been investigating various
immune responses against MHV68 to determine the molecular mechanisms by
ZKLFKȖKHUSHVYLUXVLQGXFHVDXWRDQWLERG\SURGXFWLRQDQGO\PSKRPD
development. Currently, we are investigating the relationship between infection
and the generation of autoreactive B-cells in MHV68-infected mice. In
addition, we have generated recombinant strains of MHV68, in which desired
gene cassettes, such as genes for chicken ovalbumin (OVA) and/or modified
fluorescent proteins (mVenus and mCherry), can be inserted. Together with
Fig.5 0RXVHPRGHORIȖKHUSHVYLUXVLQIHFWLRQ
knockout or transgenic mice, this technique would be of help to achieve ourwith recombinant MHV68 and gene-modified miceresearch goals.

Recent Publications
1. Takamatsu H, Takegahara N, Nakagawa Y, Tomura M, Taniguchi M, Friedel RH, Rayburn H, Tessier-Lavigne M,
Yoshida Y, Okuno T, Mizui M, Kang S, Nojima S, Tsujimura T, Nakatsuji Y, Katayama I, Toyofuku T, Kikutani H,
Kumanogoh A. Semaphorins guide the entry of dendritic cells into the lymphatics by activating myosin II. Nat
Immunol. 2010 Jul;11(7):594-600.
2. Mizui M, Kumanogoh A, Kikutani H. Immune semaphorins: novel features of neural guidance molecules. J Clin
Immunol. 2009 Jan;29(1):1-11.
3. Mizui M, Shikina T, Arase H, Suzuki K, Yasui T, Rennert PD, Kumanogoh A, Kikutani H. Bimodal regulation of T
cell-mediated immune responses by TIM-4.Int Immunol. 2008 May;20(5):695-708.
4. Suzuki K, Okuno T, Yamamoto M, Pasterkamp RJ, Takegahara N, Takamatsu H, Kitao T, Takagi J, Rennert PD,
Kolodkin AL, Kumanogoh A, Kikutani H. Semaphorin 7A initiates T-cell-mediated inflammatory responses throughalpha1beta1 integrin. Nature. 2007 Apr 5;446(7136):680-4.
5. Takegahara N, Takamatsu H, Toyofuku T, Tsujimura T, Okuno T, Yukawa K, Mizui M, Yamamoto M, Prasad DV,
Suzuki K, Ishii M, Terai K, Moriya M, Nakatsuji Y, Sakoda S, Sato S, Akira S, Takeda K, Inui M, Takai T, Ikawa
M, Okabe M, Kumanogoh A, Kikutani H. Plexin-A1 and its interaction with DAP12 in immune responses and bonehomeostasis. Nat Cell Biol. 2006 Jun;8(6):615-22.
ĲĶDivision of Host Defense

Research & Activities

Department of Immunoregulation
Research Group

Professor (SUP)
Associate Professor
Associate Professor (SUP)
Assistant Professor
SA Assistant Professor
Postdoctoral Fellow (SUP)

Taroh Kinoshita, D. Med. Sc.
Yusuke Maeda, M.D., D. Med. Sc.
Yoshiko Murakami, M.D., D. Med. Sc.
Morihisa Fujita, D. Sc.
Noriyuki Kanzawa, D. Med. Sc.
Yetao Wang, M.D., D. Med. Sc.

Research Projects
We mainly focus on two research areas involved in disease, immune defense, and homeostasis. The first research area
concerns how glycosylphosphatidylinositol (GPI)-anchored proteins are synthesized, processed, transported, and secreted, and
how GPI defects lead to the onset of diseases. The second research area concerns how the pH levels of the intracellularorganelles are regulated, and how these pH levels regulate many biological functions.
1) Biogenesis, transport, and remodeling of GPI-anchored proteins (GPI-APs).
GPI is a glycolipid that consists of phosphatidylinositol, glucosamine, mannoses, and phosphoethanolamines. It acts as a lipid
anchor for various plasma-membrane proteins. GPI-APs play important roles in host self-defense, intercellular signaltransduction, and other important processes. In addition, some GPI-APs function as receptors for certain viruses and toxins.
The GPI-anchor is widely distributed and conserved in various eukaryotes. It is essential for the development of higher animals
and for the growth of yeasts and protozoan parasites. Protein modifications due to attachment of the GPI-anchor serve asprotein localization and sorting signals. Our current project is to identify and clarify the functions of all of the genes involvedin the biosynthesis of the GPI-anchor in the ER (PIG genes; phosphatidylinositol glycan) and in the sorting and localization of
GPI-APs after their anchorage with GPI (PGAP genes; post GPI-attachment to proteins) (Fig. 1). We expect that these studies
will reveal why many proteins are modified with the GPI-anchor.

Fig. 1 GPI-anchor biosynthesis and transport/ remodeling of
GPI-APs. PIG genes are involved in the biosynthesis of the
GPI-anchor in the ER. During biosynthesis, an alkyl-lipid
generated in the peroxisome is utilized. Thereafter, GPI-APs
are transported to the plasma membrane and enriched in rafts.
PGAP genes are involved in these later processes. PGAP1
and PGAP5, which localize in the ER, and PGAP2 andPGAP3, which localize in the Golgi, are involved in the lipid
or glycan remodeling of the GPI-anchor. Remodeling affects
the sorting of GPI-APs because it alters the physicalcharacteristics of the GPI-anchor.

2) Molecular genetics of acquired paroxysmal nocturnal hemoglobinuria (PNH) and inherited GPI deficiencies.
PNH is an acquired hematopoietic stem cell disorder in which clonal cells that are defective in GPI biosynthesis are expanded.
As a result, abnormal erythrocytes that lack CD59 and DAF/CD55 predominate. CD59 and DAF/CD55 are widely distributed
GPI-anchored proteins that inhibit the activation of complement on the host cell surface. Their absence on erythrocytes makesthese cells very sensitive to complement and lysis during infections and other events. We propose a three-step model of PNH
pathogenesis. Step 1 involves the generation of GPI-deficient hematopoietic stem cells due to the somatic mutation of the
PIG-A gene. Step 2 involves the immunological selection of GPI-deficient hematopoietic stem cells. In this step, GPI-deficient
cells not only survive, but they also proliferate much more frequently than usual to compensate for anemia. This elevatedproliferation rate may increase the chance that additional genetic mutations are acquired, which leads to Step 3, in which a
subclone bearing the growth phenotype is generated (Fig. 2). We identified HMGA2 as the candidate gene for Step 3.
ĲķResearch & Activities

Together with our colleagues in England, we identified inherited GPI deficiency (IGD) caused by the PIGM mutation, amannosyltransferase-encoding gene that plays an essential role in GPI biosynthesis. Recently, through an analysis of patients’genomes using a second-generation sequencer, other IGDs, the responsible genes of which were PIGV, PIGN, and PIGA, werereported. Among these, PIGV was shown to be one of the genes responsible for Mabry syndrome, a hyperphosphatasia mental
retardation syndrome. Over 25 genes are responsible for the biosynthesis and transport of GPI-anchored proteins (Fig. 1). It is
likely that new IGDs with similar symptoms will be found. We will analyze these patients and establish a new disease categoryof “IGD” .

Fig. 2 Pathogenesis of PNH. Step 1 involves the generation of GPI-deficient hematopoietic stem cells due to the somatic mutation of the PIG-A gene.
Step 2 involves the immunological selection of GPI-deficient hematopoietic stem cells. In this step, GPI-deficient cells survive and proliferate much
more frequently than usual to compensate for anemia. This elevated proliferation may increase the chance that additional genetic mutations occur.
Step 3 involves the generation of a subclone bearing the growth phenotype.

Each intracellular organelle is compartmentalized by lipid bilayers andpossesses unique environment, proteins, and lipids in which it fully
exercises its functions. One of the major factors influencing theenvironment is the pH. Organelles in the secretory and endocytosis
pathways have acidic environments in their lumens. This acidic
environment is believed to be critical in many biological processes, such astransport, processing and glycosylation of both proteins and lipids, as wellas in the onset and pathology of diseases that affect pH regulation. Despite
their significance, the mechanisms by which abnormal phenotypes arecaused are largely unknown. We recently reported the first establishmentof mutant cells in which the Golgi pH was dysregulated and identified agene responsible for the mutants and its functions (Ref. 5). We aim toclarify the mechanisms and biological significance of acidic pH regulation
through establishing and analyzing mutant cells defective in organelle pH
regulation, and to understand the disease pathologies to allow drug
discovery.

Fig. 3 pH regulation of the intracellular organelles.
Luminal pH levels of organelles in secretory andendocytosis pathways are acidic, as indicated in theboxes (values are approximate).

Recent Publications
1. Murakami Y, Kanzawa N, Saito K, Krawitz PM, Mundlos S, Robinson PN, Karadimitris A, Maeda Y, Kinoshita T.
Mechanism for release of alkaline phosphatase caused by glycosylphosphatidylinositol deficiency in patients withhyperphosphatasia mental retardation syndrome. J Biol Chem. 2012 Feb 24;287(9):6318-25.
2. Fujita M, Watanabe R, Jaensch N, Romanova-Michaelides M, Satoh T, Kato M, Riezman H, Yamaguchi Y, Maeda
Y, Kinoshita T. Sorting of GPI-anchored proteins into ER exit sites by p24 proteins is dependent on remodeled GPI.
J Cell Biol. 2011 Jul 11;194(1):61-75.
3. Kanzawa N, Maeda Y, Ogiso H, Murakami Y, Taguchi R, Kinoshita T. Peroxisome dependency of alkyl-containing
GPI-anchor biosynthesis in the endoplasmic reticulum. Proc Natl Acad Sci U S A. 2009 Oct 20;106(42):17711-6.
4. Fujita M, Maeda Y, Ra M, Yamaguchi Y, Taguchi R, Kinoshita T. GPI glycan remodeling by PGAP5 regulates
transport of GPI-anchored proteins from the ER to the Golgi. Cell. 2009 Oct 16;139(2):352-65.
5. Maeda Y, Ide T, Koike M, Uchiyama Y, Kinoshita T. GPHR is a novel anion channel critical for acidification andfunctions of the Golgi apparatus. Nat Cell Biol. 2008 Oct;10(10):1135-45.
ĲĸDivision of Host Defense

Research & Activities

Department of Host Defense
Research Group

Professor (SUP)
Associate Professor
SA Associate Professor (SUP)
Assistant Professor
SA Assistant Professor
SA Assistant Professor (SUP)

Shizuo Akira
Taro Kawai
Satoshi Uematsu
Tatsuya Saitoh
Yutaro Kumagai
Takashi SatohThe innate immune system senses invading microbial pathogens, such as bacteria, viruses, and parasites, and plays an essential
role in inducing inflammatory responses and assisting adaptive immune responses. Pattern-recognition receptors (PRRs)
expressed on innate immune cells, such as macrophages and dendritic cells, recognize pathogen-associated molecular patterns,
which are conserved molecular features of microbial pathogens. Upon recognition of pathogen-associated molecular patterns,
PRRs initiate signaling pathways responsible for innate immune responses. We are seeking to clarify the complex regulatory
mechanisms of the innate immune system.
1) Understanding signaling pathways via PRRs and their roles in immune responses
So far, Toll-like receptors (TLRs), RIG-I-like receptors, and Nod-like receptors have been identified as PRRs. We are using
molecular biology and bioinformatics tools to identify molecules involved in PRR signaling. We are also generating mice
deficient in relevant genes to understand their in vivo function with respect to immune responses. In particular, we focus on therecognition mechanism of virus- or bacteria-derived nucleic acids by PRRs and their signaling pathways (Fig. 1).
Fig. 1: PRRs that mediate the sensing of foreign nucleic
acids and their signaling pathways
TLR3, TLR7, and TLR9 are exclusively localized to
endosomes and recognize viral double-stranded (ds)RNA, single-stranded (ss) RNA of RNA viruses, andDNA derived from viruses or bacteria, respectively.
Whereas TLR3 signals through an adapter TRIF, TLR7
and TLR9 use MyD88 as an adapter to induce type I
interferon and inflammatory cytokines. In thecytoplasm, RIG-I-like receptors, such as RIG-I and
MDA5, recognize different RNA species derived fromRNA viruses as indicated and signal through an adapter
IPS-1. Double-stranded DNA is recognized by AIM2 inthe cytoplasm, which promotes caspase-1–dependent
IL-1 production through recruitment of an adapter
ASC. Although STING appears to mediate a type I
interferon-inducing pathway in response to dsDNA,
cytosolic dsDNA sensors upstream of STING are notwell understood.

2) Regulation of innate immune responses by membrane trafficking
Dynamic membrane traffic plays a crucial role in the regulation of innate immune responses. We found that the autophagy
proteins ATG16L1 and ATG9a and the interferon-inducible protein Viperin regulate innate immune responses, such as the
production of inflammatory cytokines and type I interferon, in vivo. We seek to find new mechanisms of membrane trafficking
involved in the regulation of innate immune responses.
3) Analysis of transcriptional networks in innate immune cells
We aim to understand the transcriptional networks in innate immune cells by comprehensively comparing gene expression
profiles in dendritic cells derived from mice deficient for molecules involved in PRR signaling. We are comprehensively
analyzing promoter sequences of various genes to identify transcription factors that regulate innate immune responses.
Furthermore, we are investigating histone modifications to understand the role of histones in innate immune responses andmacrophage polarization (M1 vs. M2 macrophages) (Fig. 2).

ĲĹResearch & Activities

Fig. 2: Regulation of M2 macrophage polarization by Jmjd3
Macrophages are functionally polarized into M1 and M2 cells. M1
macrophages are critical for clearing bacterial, viral, and fungal infections,whereas M2 macrophages play an important role in responses to parasite
infection, tissue remodeling, angiogenesis, and tumor progression. Jmjd3 isa H3K27 demethylase catalyzing H3K27me3 (trimethylated) to HeK27me1(monomethylated). It is induced in macrophages by TLR stimuli. Jmjd3 isessential for M2 macrophage polarization in response to helminth infection
and chitin, although Jmjd3 is dispensable for M1 responses. Jmjd3
deficiency affects H3K27 trimethylation in only a limited number of genes.
Among them, IRF4 is a key transcription factor that controls M2
macrophage polarization. Thus, Jmjd3 plays a critical role in regulating M2
macrophage development, leading to antihelminth host responses.

4) Regulation of the stability of mRNA encoding inflammatory cytokines
The expression of genes encoding inflammatory cytokines is controlled by transcription factors and molecules that mediate themRNA stability. We identified a TLR-inducible CCCH-type zinc finger-containing protein, Regnase-1 (Zc3h12a), as amolecule that degrades IL-6 mRNA. Mice lacking this gene spontaneously exhibit severe autoimmune symptoms. Currently,
we are investigating the roles of other proteins belonging to this protein family in the regulation of inflammatory responses(Fig. 3).
Fig. 3: Regulation of inflammatory responses by Regnase-1
Regnase-1 is an RNase that regulates the decay of mRNA encoding
inflammatory cytokines (such as IL-6) and suppresses inflammation.
Regnase-1 constitutively degrades IL-6 mRNA under the unstimulated
condition (1). Activation of the IKK complex by TLR engagement
leads to the phosphorylation and degradation of I B and Regnase-1(2). Transcription of IL-6 mRNA is induced (3) and IL-6 is produced(4). Regnase-1 mRNA is also transcribed and Regnase-1 protein isexpressed (5). Regnase-1 promotes the degradation of IL-6 mRNA to
suppress IL-6 production (6).

Recent Publications
1. Iwasaki H, Takeuchi O, Teraguchi S, Matsushita K, Uehata T, Kuniyoshi K, Satoh T, Saitoh T, Matsushita M,
6WDQGOH\'0$NLUD67KH,ț%NLQDVHFRPSOH[UHJXODWHVWKHVWDELOLW\RIF\WRNLQHHQFRGLQJP51$LQGXFHGE\TLR-IL-1R by controlling degradation of regnase-1. Nat Immunol. 2011 Oct 30;12(12):1167-75.
2. Saitoh T, Satoh T, Yamamoto N, Uematsu S, Takeuchi O, Kawai T, Akira S. Antiviral protein Viperin promotes
Toll-like Receptor 7- and Toll-like Receptor 9-mediated type I interferon production in plasmacytoid dendritic cells.
Immunity. 2011 Mar 25;34(3):352-63.
3. Kawai T, Akira S. Toll-like Receptors and their crosstalk with other innate receptors in infection and immunity.
Immunity. 2011 May 27;34(5):637-50.
4. Tsuchida T, Zou J, Saitoh T, Kumar H, Abe T, Matsuura Y, Kawai T, Akira S. The ubiquitin ligase TRIM56
regulates innate immune responses to intracellular double-stranded DNA. Immunity. 2010 Nov 24;33(5):765-76.
5. Satoh T, Takeuchi O, Vandenbon A, Yasuda K, Tanaka Y, Kumagai Y, Miyake T, Matsushita K, Okazaki T, Saitoh
T, Honma K, Matsuyama T, Yui K, Tsujimura T, Standley DM, Nakanishi K, Nakai K, Akira S. The Jmjd3-Irf4 axis
regulates M2 macrophage polarization and host responses against helminth infection. Nat Immunol. 2010 Oct;11(10):936-44.
ĲĺResearch & Activities

Division of Host Defense

Department of Cell Biology
Research Group

Professor
Associate Professor
Assistant Professor
SA Researcher
SA Researcher
SA Researcher

Eisuke Mekada, Ph.D.
Ryo Iwamoto, Ph.D.
Hiroto Mizushima, Ph.D.
Takashi Nakamura, Ph.D.
Tomoya Hikita, Ph.D.
Mizuho Sato, Ph.D.

We are studying the mechanisms of cell growth and differentiation that involve growth factors and adhesion molecules
present at the cell-cell contact sites. In particular, we are focusing on the mode of action of HB-EGF, a membrane-anchoredprotein from the EGF family of growth factors, and the molecules that bind to HG-EGF, namely the tetraspanin family.
These proteins function in morphogenesis and tissue maintenance and repair by regulating cell proliferation, migration, andadhesion. They are also involved in the growth, invasion, and metastasis of cancer cells.
1) Mode of action of HB-EGF
HB-EGF is a member of the EGF family of growth factors and binds to and activates EGFR and ErbB4. It is synthesized as
proHB-EGF, a membrane-anchored precursor protein, and is cleaved on the cell surface to yield the soluble growth factor(sHB-EGF). The conversion of proHB-EGF into the soluble form is critical for its activity; therefore, this process is tightly
regulated. HB-EGF is secreted by various tissues and cells, and it functions in several physiological processes. For example,
it maintains heart muscle function, suppresses cell proliferation in the heart valve and lung alveolar development, promotes
cell migration in wound healing and eyelid closure, supports blastocyst attachment to the uterus during implantation, andpromotes cell proliferation in skin hyperplasia. ProHB-EGF is not only a precursor of the soluble form, but is also abiologically active molecule that regulates the growth of neighboring cells in a juxtacrine fashion.
How is the conversion of the membrane-anchored form into the soluble form regulated? How does HB-EGF function in the
manifold physiological processes that depend on this molecule? What roles do sHB-EGF and proHB-EGF play in suchphysiological processes? Do they participate in pathological processes? These questions are currently being analyzed at themolecular level.
2) Role of HB-EGF in cancer malignancy
HB-EGF is expressed in cancer cells and/or cancer-derived stroma (Ichise et al. 2010; Murata et al. 2011), and it is involved

Fig. 1 The role of HB-EGF in heart valve formation.
In heart valve development, HB-EGF is expressed and secreted fromendocardial cells. Secreted HB-EGF associates with heparan sulfate
proteoglycans (HSPGs) in the valve mesenchyme (upper right)(Fig. 1).
When EGFR on the surface of mesenchymal cells is activated by the
HB-EGF-HSPG complex, this receptor transduces inhibitory signals formesenchymal cell proliferation (lower left). Knock-in mice expressing a
PXWDQW+%(*)WKDWFDQQRWELQGWR+63*V ǻ+% DVZHOODV.2PLFH
develop enlarged cardiac valves with hyperproliferation of mesenchymal
cells (lower right) (Iwamoto et al., 2010).

ĳı

Research & Activities

in tumor growth, invasion, and metastasis (Fig. 2, 3). To develop a novel strategy for cancer therapy, we are analyzing therole of HB-EGF in cancer malignancy.

Fig. 3. Cancer-derived fibroblasts enhance tumorigenesis of
cervical cancer cells in nude mice (Murata et al. 2011).
Fluorescence imaging of tumors injected with cervical cancer
cells with fibroblasts (a) or with cervical cancer cells alone (b).
Fig. 2. Cancer cell–stromal fibroblast interactions in the cervix (Murata et al. 2011).

3) Development of anticancer drugs targeting HB-EGF
We are developing new anticancer drugs targeting HB-EGF. Preclinical and clinical studies of an anti-HB-EGF monoclonal
antibody (Miyamoto et al. 2011) and a nontoxic mutant protein of diphtheria toxin CRM197 (BK-UM) are in progress (Fig. 4).
4) CD9 and tetraspanin function
CD9, a member of the tetraspanin superfamily, is a membrane protein with four transmembrane domains. It associates with
proHB-EGF and upregulates proHB-EGF function. CD9 is also involved in cell signaling, growth, motility, and adhesion,and in tumor cell metastasis and sperm-egg fusion. In addition, the Caenorhabditis elegans tetraspanin TSP-15 is essential
for the epidermal integrity of the worm. We are analyzing the physiological activity of CD9 and other tetraspanins by using
genetically engineered mice or worms (C. elegans) lacking CD9 or other tetraspanins.

Fig. 4. Enhanced expression of HB-EGF in ovarian cancer tissue (right) and BK-UM,a therapeutic for ovarian cancer under development by our laboratory.

Recent Publications
1. Miyamoto S, Iwamoto R, Furuya A, Takahashi K, Sasaki Y, Ando H, Yotsumoto F, Yoneda T, Hamaoka M, Yagi H,
Murakami T, Hori S, Shitara K, Mekada E. A novel anti-human HB-EGF monoclonal antibody with multiple
anti-tumor mechanisms against ovarian cancer cells. Clin Cancer Res. 2011 Nov 1;17(21):6733-41.
2. Murata T, Mizushima H, Chinen I, Moribe H, Yagi S, Hoffman RM, Kimura T, Yoshino K, Ueda Y, Enomoto T,
Mekada E. HB-EGF and PDGF mediate reciprocal interactions of carcinoma cells with cancer-associated fibroblasts
to support progression of uterine cervical cancers. Cancer Res. 2011 Nov 1;71(21):6633-42.
3. Iwamoto R, Mine N, Kawaguchi T, Minami S, Saeki K, Mekada E. HB-EGF function in cardiac valve development
requires interaction with heparan sulfate proteoglycans. Development. 2010 Jul;137(13):2205-14.
 ,FKLVH7$GDFKL62KLVKL0,NDZD02NDEH0,ZDPRWR50HNDGD(+XPDQL]HGJHQHUHSODFHPHQWLQPLFH
reveals the contribution of cancer stroma-derived HB-EGF to tumor growth. Cell Struct Funct. 2010;35(1):3-13.
5. Mizushima H, Wang X, Miyamoto S, Mekada E. Integrin signal masks growth-promotion activity of HB-EGF in
monolayer cell cultures. J Cell Sci. 2009 Dec 1;122(Pt 23):4277-86.
ĳĲ

Research & Activities

Division of Host Defense

Department of Immunochemistry
Research Group

Professor
Assistant Professor
Assistant Professor
SA Assistant Professor
SA Researcher
SA Researcher

Hisashi Arase, M.D., Ph.D., SUP
Tadahiro Suenaga, M.D., Ph.D., SUP
Masako Kohyama, Ph.D., SUP
Kouyuki Hirayasu,Ph.D.
Fumiji Saito, Ph.D.
Jing Wang, Ph.D.

Research Projects
Our department analyzes how pathogens, such as viruses, have acquired the ability to evade the immune system and how hostimmune systems have acquired resistance to various pathogens. In particular, we seek to elucidate a fundamental mechanismof host defense against various pathogens by analyzing the immune regulatory receptors. Of particular interest are “paired
receptors,” which consist of activating and inhibitory receptors expressed on some immune cells (Figure 1). Results from theidentification of host ligands and their corresponding viral ligands have led us to suggest that these “paired receptors”
coevolved with the pathogens. We have also found that these receptors mediate viral entry into cells. Our studies will help to
explain the fundamental mechanisms of immune evasion by pathogens and the host factors that influence resistance to various
infections. This research will help to build the foundation required for the development of new vaccines and therapies for
infectious diseases.
(1) Analysis of recognition by “paired receptors”
Immune cells express various receptors comprised of highly homologous activating and inhibitory receptors. The
inhibitory receptors recognize self-antigens, such as MHC molecules. In contrast, the activating receptors generally do notrecognize self-antigens, and the ligands for activating receptors remain unclear. We have found that one of these “paired
receptors” recognizes cytomegalovirus protein and showed that “paired receptors” play an important role in determining
host resistance to pathogens (Figure 2). We are continuing to analyze the functions of these receptors using various
pathogens, including viruses, parasites, and bacteria, to elucidate the interactions between pathogens and the host immune
system.

Figure 1. Paired receptors
Paired receptors consist of inhibitory and activating
receptors that are highly homologous to each other. The
inhibitory receptors transduce inhibitory signals via
immunoreceptor tyrosine-based inhibition motif (ITIM)in their cytoplasmic domain, whereas the activating
receptors transduce activating signals by association
with immunoreceptor tyrosine-based activation motif(ITAM)-bearing adaptor molecules.

ĳĳ

Figure 2. Recognition of cytomegalovirus-infected cells by inhibitory andactivating paired receptors
Cytomegalovirus have MHC-like molecules that serve as ligands forinhibitory receptors expressed on the natural killer (NK) cells of susceptible
mouse strains. As a result, virus-infected cells are not killed by NK cells,
despite their low MHC expression (left). In contrast, NK cells from resistantmouse strains express activating rather than inhibitory receptors thatrecognize the viral MHC-like molecules. Therefore, these cells canefficiently eliminate virus-infected cells (right) (Arase et al. Science 2002).

Research & Activities(2) Entry mechanism of virus into cells
Several viruses that show persistent infection downregulate the immune response by expressing ligands for inhibitory
receptors. Interestingly, we have found that some viruses exploit inhibitory receptors to enter the cells. Interactions
between immune receptors and viral proteins play an important role in the entry mechanism of herpes simplex virus (HSV)(Figure 3). Because there is a possibility that other viruses may use similar receptors to enter cells, we will further
investigate the molecular mechanisms involved in viral entry into cells.

Figure 3. Entry mechanism of virus into cells
6RPHYLUXVHVH[SUHVVOLJDQGVIRULQKLELWRU\UHFHSWRUVDQGGRZQUHJXODWHWKHLPPXQHUHVSRQVH3,/5ĮRQHRIWKH
inhibitory paired receptor, was found to recognize herpes simplex virus (HSV)-infected cells. Molecular cloning of the
OLJDQGVIRU3,/5ĮUHYHDOHGWKDW3,/5ĮUHFRJQL]HV+69JO\FRSURWHLQ%ZKLFKSOD\VDQHVVHQWLDOUROHLQ+69LQIHFWLRQ,QWHUDFWLRQVEHWZHHQ3,/5ĮDQGJO\FRSURWHLQ%ZHUHLQYROYHGLQYLUDOHQWU\LQWRFHOOV2QWKHRWKHUKDQGJO\FRSURWHLQ%
of varicella-zoster virus associated with myelin-associated glycoprotein (MAG, Siglec-4), another component of a paired
receptor, which mediated VZV entry into cells. In this way, paired receptors were found to play important roles not onlyin immune regulation but also in viral entry into cells.

Recent Publications
 .RJXUH$6KLUDWRUL,:DQJ-//DQLHU/$UDVH+3$13LVDQRYHO2JO\FRV\ODWHG3,/5ĮOLJDQGH[SUHVVHGLQ
neural tissues. Biochem Biophys Res Commun 2011 Feb 18; 405(3): 428-433.
2. Suenaga T, Satoh T. Somboonthum P, Kawaguchi Y, Mori Y, and Arase H. Myelin-associated glycoprotein
mediates membrane fusion and entry of neurotropic herpesviruses. Proc Natl Acad Sci USA. 2010 Jan 12; 107(2):866-71.
3. Wang J, Fan Q, Satoh T, Arii J, Lanier LL, Spear PG, Kawaguchi Y, Arase H.Binding of herpes simplex virus
JO\FRSURWHLQ% J% WR3,/5ĮGHSHQGVRQVSHFLILFVLDO\ODWHG2OLQNHGJO\FDQVRQJ%J Virol. 2009 Dec ; 83(24):13042-5.
4. Satoh T, Arii J, Suenaga T, Wang J, Kogure A, Uehori J, Arase N, Shiratori I, Tanaka S, Kawaguchi Y, Spear PG,
Lanier LL, Arase H. PILRĮ is a herpes simplex virus-1 entry co-receptor that associates with glycoprotein B. Cell.
2008 Mar 21; 132(6): 935-44
5. Wang J, Shiratori I, Satoh T, Lanier LL, Arase H. An essential role of sialylated O-linked sugar chains in therecognition of mouse CD99 by paired immunoglobulin-like type 2 receptor (PILR). J Immunol. 2008 Feb 1; 180(3):1686-93ĳĴResearch & Activities

Division of Cellular and Molecular Biology

Department of Molecular Genetics
Research Group

Professor
Associate Professor
Assistant Professor (SUP)SA Researcher

Hiroshi Nojima, Ph.D.
Norikazu Yabuta, Ph.D.
Daisuke Okuzaki, Ph.D.
Yoko Naito, Ph.D.

Research Projects
We are studying the eukaryotic cell cycle to understand the mechanisms underlying the observed chromosomal instability incancer cells. Many human cancer cells exhibit mitotic defects (such as centrosome aberrations, abnormal spindle formation,and chromosome missegregation). The resulting chromosomal instability is a major cause of malignant tumor progression. Our
research focuses on functional analyses of the Ser/Thr kinases Lats (large tumor suppressor) and GAK (cyclin G-associatedkinase). These kinases localize to the centrosome, regulate mitotic progression in response to DNA damage, and cause
chromosome instability when their functions are disrupted. Both Lats (Lats1 and Lats2) and GAK form complexes with
Mdm2. In turn, Mdm2 controls the stability of p53, which is a transcriptional regulator of the Lats2, cyclin G1, and Mdm2
genes. Thus, the Lats and GAK complexes have intimate correlations in their function (Fig. 1).
Our research subjects are as follows:(1) Lats Group
Lats1 and Lats2, which belong to the Lats kinase family, are highly
conserved across species and localize to the centrosome during the cell
cycle. Two miRNAs, miRNA-372 and -373, function as potential novel
oncogenes in testicular germ cell tumors by inhibiting LATS2 expression.
This observation suggests that Lats2 is an important tumor suppressor(Voorhoeve et al., Cell, 2006). Lats1/2 plays an important role in the cell
cycle checkpoint and the Hippo pathway that mainly regulates organ sizeand cell growth. In particular, Lats2 binds to Mdm2, thereby inhibiting
its E3 ligase activity and activating p53. Then, p53 rapidly andselectively upregulates Lats2 expression in G2/M cells. This positive
feedback loop constitutes a novel checkpoint that plays a critical role inthe maintenance of proper chromosome numbers (Aylon et al., Gene
Dev., 2006).
We have discovered the following: (A) LATS2 knockout mice areembryonic lethal, which indicates the essential role of Lats2 in thedevelopment and differentiation of mammalian germ cells. (B) Lats2-/mouse embryonic fibroblasts (MEF) display an enhanced growth rate,
centrosome fragmentation (Fig. 2), misalignment of the chromosome at
M phase, abnormal chromosome segregation, and aberrant cytokinesis.
These results indicate the essential role that Lats2 plays in proper M
phase progression (Yabuta et al., J. Biol. Chem., 2007). (C) Lats2
mediates two novel signaling pathways, CLP (Chk1/2—Lats2—P-body)and ALB (Aurora-A—Lats1/2—Aurora-B). In the CLP pathway, Lats2
phosphorylates 14-3-3 for processing body (P-body) formation
downstream of Chk1 in response to DNA damage (Fig. 3; GW182 is acomponent of P-body) (Ref. 4). In the ALB pathway, Lats2 is
phosphorylated by Aurora-A. Phosphorylated Lats2 localizes to thechromosomes and the central spindle, and regulates Lats1 and Aurora-B,
thereby executing proper chromosomal segregation during mitosis (Fig.
4). (D) Lats2 phosphorylates and regulates ASPP1/p53 complexes,
thereby undergoing apoptosis in malignant tumor cells with aneuploidy
and polyploidy (Ref. 5). Moreover, Lats2 also phosphorylates thetranscription factor Snail, thereby regulating the epithelial-mesenchymaltransition (EMT) (Ref. 2).
(2) GAK（cyclin G-associated kinase）/cyclin G GroupGAK is an association partner of clathrin heavy chain (CHC) and isessential for clathrin-mediated membrane trafficking. Unlike
neuron-specific auxilin, which plays a similar role in neural cells, GAKĳĵFig. 1: The functions of the Lats and GAK complexes
correlate closely.

Fig. 2: Centrosome fragmentation in Lats2-/- MEFs.

Fig. 3: Lats2 is required for P-body formation after UV irradiation.

Fig. 4: Lats2 co-localizes with Aurora-B at the central spindle.

Research & Activities

has a kinase domain (Fig. 5) whose function has remained unclear. We have discovered the following: (a) GAK forms acomplex with PP2A B’ȖDQGF\FOLQ* F\FOLQ*DQGF\FOLQ* ZKLFKUHJXODWHWKHGHSKRVSKRU\ODWLRQDFWLYLW\RI33$ )LJ
6; Naito et al., Cell Cycle, 2012). (b) GAK localizes to the cytoplasm and the nucleus, where it has two additional functions(i.e., maintenance of proper centrosome maturation and mitotic chromosome congression). (c) GAK knockdown by siRNAcauses cell cycle arrest at metaphase, which indicates that GAK is required for proper mitotic progression. This impaired
mitotic progression was found to be due to the activation of the spindle assembly checkpoint (SAC), which senses protruding,misaligned, or abnormally condensed chromosomes in knockdown cells (Fig. 7; Shimizu et al., J. Cell Sci., 2009). (d)GAK-kd-/- mice exhibit neonatal lethality with pulmonary dysfunction (Fig. 8). Because an EGFR inhibitor, Gefitinib (Iressa),
also efficiently inhibits the kinase activity of GAK, the side effects of Gefitinib, including interstitial pneumonia, may be
responsible for the inhibition of GAK (Ref. 1).

Fig. 5: GAK is similar to auxilin except for bearing a kinase domain.

)LJ7RI33$%¶ȖE\*$.LVDQLPSRUWDQWSKRVSKRU\ODWLRQ
site for the activity of PP2A complex.

Fig. 7: GAK knockdown by siRNA (Ki9) generates
abnormal chromosomes.

Fig. 8: Severe defects of fetal lung maturity in GAK-kd-/- mice.

Recent Publications
1. Tabara H, Naito Y, Ito A, Katsuma A, Sakurai MA, Ohno S, Shimizu H, Yabuta N, Nojima H. Neonatal lethality inknockout mice expressing the kinase-dead form of the gefitinib target GAK is caused by pulmonary dysfunction.
PLoS ONE. 2011, 6(10): e26034.
2. Zhang K, Rodriguez-Aznar E, Yabuta N, Owen RJ, Mingot JM, Nojima H, Nieto MA, Longmore GD. Lats2 kinasepotentiates Snail1 activity by promoting nuclear retention upon phosphorylation. EMBO J. 2011 Sep 27;31(1):29-43.
3. Yabuta N, Mukai S, Okada N, Aylon Y, Nojima H. The tumor suppressor Lats2 is pivotal in Aurora A and Aurora B
signaling during mitosis. Cell Cycle. 2011, 10(16):2724-36.
4. Okada N, Yabuta N, Suzuki H, Aylon Y, Oren M, Nojima H. A novel Chk1/2-Lats2-14-3-3 signaling pathway
regulates P-body formation in response to UV damage. J. Cell Sci. 2011 Jan 1;124(Pt 1):57-67.
5. Aylon Y, Ofir-Rosenfeld, Y., Yabuta N, Lap, E. Nojima H, Lu, X. and Oren M. The Lats2 tumor suppressor
augments p53-mediated apoptosis by promoting the nuclear proapoptotic function of ASPP1. Genes Dev. 2010 Nov
1;24(21):2420-9.
ĳĶResearch & Activities

Division of Cellular and Molecular Biology

Department of Oncogene Research
Research Group

Professor
Associate Professor
Associate Professor
SA Researcher
SA Researcher

Masato Okada, D.Sc.
Shigeyuki Nada, D.Sc.
Chitose ONEYAMA, D.Sc.
Kentaro KAJIWARA, D.Sc.
Kyoko KAKUMOTO, D.Sc.

Research Projects
Cancers arise, evolve, and develop progressively due to the accumulation of mutations and/or modifications in the genomic
DNA. Loss-of-function mutations in “tumor suppressor genes” induce cell immortalization, whereas gain-of-function
mutations in “proto-oncogenes” induce cell transformation. Cell immortalization prevents the induction of apoptosis and/or
senescence, which is a defense mechanism against cancer development. Cell transformation involves the gain of autonomous
cell growth, loss of cell communication, morphological changes, and increased production of matrix proteases and growth
factors that participate in invasion, metastasis, and angiogenesis. These cellular events lead to the malignant conversion ofcancer cells.
The primary focus of this department is to understand the molecular basis of the cell transformation that is induced bygain-of-function mutations of proto-oncogenes. As a representative proto-oncogene, we have focused on the c-Src
proto-oncogene, which encodes a nonreceptor tyrosine kinase. To date, we have analyzed its physiological roles indevelopment and the mechanisms by which it is modulated by its specific regulators, such as Csk and Cbp. To obtain a full
picture of the cell signaling pathways that lead to c-Src–mediated cell transformation and to search for new therapeutic targets
that will block c-Src–mediated cancer progression, the following projects are currently in progress:
I. Function and regulation of c-Src
In normal cells, c-Src is present as an inactive form that is
phosphorylated by its negative regulatory kinase Csk. Extracellular
stimuli transiently activate it, after which it activates downstream
components, such as the MAPK pathway and Rho family GTPases,
thereby inducing gene expression and cytoskeletal rearrangements,
respectively. Consequently, promotion of cell growth andphenotypic changes that are involved in cell transformation are
induced (Fig. 1). Although the c-src gene is rarely mutated inhuman cancers, its protein is frequently hyperactivated andoverexpressed. This aberrant activation of c-Src may contribute tocancer malignancy. We recently showed that the oncogenic
potential of c-Src is regulated by a membrane adaptor protein, Cbp,
which is exclusively localized to the membrane microdomain. We
are currently analyzing the relationship between the disruption ofthis regulatory system and cancer progression.
II. Mechanism of cell transformation induced by c-Src activation.
To define the signaling pathway that leads to c-Src–mediated celltransformation, we performed a comprehensive study of target
molecules using a newly developed c-Src inducible system.
Proteomic analysis identified Arhgef5, which is a Dbl family
member GEF for Rho GTPase, as a critical c-Src target. A miRNAarray analysis revealed that c-Src activation altered the gene
expression of a limited number of miRNAs. We identified several
target genes for these miRNAs and analyzed their functions. Manyof the target molecules (e.g., FGFR3, mTOR, and ILK) werecomponents of the c-Src signaling pathway; therefore, we proposed aĳķFig. 1. Function and regulation of c-Src.

Fig. 2. A mechanism of c-Src–induced cell transformation
regulated by miRNA.

III. Regulation of cell growth signaling via late endosomes/lysosomes.
During the search for c-Src targets, we identified themembrane adaptor protein p18, which is exclusively localized
to late endosomes/lysosomes, as a new regulator of cell growthsignaling. The p18 protein forms a ternary complex withscaffolding p14/MP1 proteins and plays an essential role toactivate mTORC1 on late endosomes/lysosomes. We foundthat p18 KO mice are embryonic lethal and exhibit severe
defects in lysosome biogenesis (Fig. 3). To address thefunction of p18 in lysosome biogenesis, we are currently
searching for target molecules for the p18-mTORC1 pathway
in late endosomes/lysosomes. The p18-dependent pathway is
crucial for controlling cell growth and oncogene-mediated celltransformation. Elucidation of the underlying mechanism isanother important subject of our project.

Research & Activities

new tuning system of c-Src–mediated cell transformation via miRNA. Analysis of the regulatory mechanism forc-Src–mediated miRNA expression is now underway.

Fig. 3. Roles played by the late endosome/lysosome-anchoredp18-mTORC1 pathway.

Recent Publications
1 : Takahashi Y, Nada S, Mori S, Soma-Nagae T, Oneyama C, Okada M. The late endosome/lysosome-anchoredp18-mTORC1 pathway controls terminal maturation of lysosomes. Biochem Biophys Res Commun. 2012 Jan
27;417(4):1151-7.
2 : Oneyama C, Morii E, Okuzaki D, Takahashi Y, Ikeda J, Wakabayashi N, Akamatsu H, Tsujimoto M, Nishida T,
Aozasa K, Okada M. MicroRNA-mediated upregulation of integrin-linked kinase promotes Src-induced tumor
progression. Oncogene. 2012 Mar 29;31(13):1623-35.
3 : Kuroiwa M, Oneyama C, Nada S, Okada M. The guanine nucleotide exchange factor Arhgef5 plays crucial roles in
Src-induced podosome formation. J Cell Sci. 2011 May 15;124(Pt 10):1726-38.
4 : Oneyama C, Ikeda J, Okuzaki D, Suzuki K, Kanou T, Shintani Y, Morii E, Okumura M, Aozasa K, Okada M.
MicroRNA-mediated downregulation of mTOR/FGFR3 controls tumor growth induced by Src-related oncogenic
pathways. Oncogene. 2011 Aug 11;30(32):3489-501.
5 : Oneyama C, Hikita T, Enya K, Dobenecker MW, Saito K, Nada S, Tarakhovsky A, Okada M. The lipid raft-anchoredadaptor protein Cbp controls the oncogenic potential of c-Src. Mol Cell. 2008 May 23;30(4):426-36.
ĳĸResearch & Activities

Division of Cellular and Molecular Biology

Department of Signal Transduction
Research Group

Professor
Assistant Professor
Assistant Professor
SA Researcher
SA Researcher
Postdoctoral Fellow
Postdoctoral Fellow

Nobuyuki Takakura, M.D., Ph.D.
Hiroyasu Kidoya, Ph.D.
Hisamichi Naito, M.D., Ph.D.
Yinglu Han, Ph.D.
Daishi Yamakawa, Ph.D.
Yumi Kinugasa, Ph.D.
Tomomi Mohri, Ph.D.

Research Projects
The development of normal tissues and organs requires the generation of tissue-specific cells from stem cells. Maintenance of
this stem cell system also requires the generation of an appropriate microenvironment. Blood vessels are the most essential
structures in tissues and organs. With a few exceptions, the development of all tissues requires blood vessel formation. Our
research group is analyzing the molecular mechanisms by which blood vessels form under physiological and pathologicalconditions, including in cancers and inflammation. We are also elucidating the mechanisms that cause stem cells to associate
closely with blood vessels. Our ultimate goal is to establish strategies to inhibit the malignant progression of various diseases.
Our specific research projects are as follows:
I. Analysis of the molecular mechanisms of blood vessel formation
1) Molecular analysis of angiogenesis, with a particular focus on the Tie1 and Tie2 receptors
2) Physiologic and pathologic function of endothelial stem cells (ESCs)
3) Molecular characterization of arterio-venous patterning, with a particular focus on the apelin/APJ system
4) Development of a system that delivers drugs into blood vessels
II. Molecular analysis of self-renewal in normal and cancer stem cells (CSCs)
1) Mechanism of stem cell reprogramming2) Analysis of cell cycle regulation in stem cells, with a special focus on Galectin-3 and the GINS complex
3) Establishment of a strategy that can inhibit the formation of the vascular niche inhabited by CSCs

Figure 1. ESCs and angiogenesis. During angiogenesis, tip cells in the front of new vessels determine the migration
direction of new vessels; stalk cells produce greater numbers of endothelial cells (ECs) for elongation; and the phalanx
cells induce maturation of new blood vessels. Recently, ESCs were identified in the preexisting blood vessels. These
ESCs may produce heterogeneous ECs and critically regulate the processes of pathological and physiologicalangiogenesis.

ĳĹResearch & Activities

Figure 2. Vascular niche of CSCs. The vascular niche was foundto contain CSCs by using PSF1, a member of DNA replication
factors GINS. Molecular analysis of vascular niche formation
may be highly useful for the development of new therapies for
CSCs.

Figure 3. Implication of the Tie2/Angiopoietin-1 and APJ/apelin axis for blood vessel
maturation. Angiopoietin-1 (Ang1) stimulates Tie2 on endothelial cells (ECs), which
results in EC migration and proliferation. Ang1 promotes the production of apelin by ECs,
thereby stimulating the apelin receptor APJ on ECs. The activation of APJ induces theproliferation and assembly of ECs, resulting in blood vessel enlargement. Apelin also
induces firm cell–cell adhesion between ECs regulating VE-cadherin and claudin-5.
Although VEGF induces narrow and leaky blood vessels, Ang1 and apelin regulate mature
blood vessels.

Recent Publications
1. Naito H, Kidoya H, Sakimoto S, Wakabayashi T, Takakura N. Identification and characterization of a resident
vascular stem/progenitor cell population in preexisting blood vessels. EMBO J. 2011 Dec 16;31(4):842-55.
2. Kidoya H, Naito H, Takakura N. Apelin induces enlarged and non-leaky blood vessels for functional recovery fromischemia. Blood. 2010 Apr 15;115(15):3166-74.
3. Nagahama Y, Ueno M, Miyamoto S, Morii E, Minami T, Mochizuki N, Saya H, Takakura N. (2010). PSF1, a DNA
replication factor expressed widely in stem and progenitor cells, drives tumorigenic and metastatic properties.
Cancer Res. 2010 Feb 1;70(3):1215-24.
4. Ueno M, Itoh M, Sugihara K, Asano M, and Takakura N. (2009). Both Alleles of PSF1 are Required for
Maintenance of Pool Size of Immature Hematopoietic Cells and Acute Bone Marrow Regeneration. Blood. 2009 Jan
15;113(3):555-62.
5. Kidoya H, Ueno M, Yamada Y, Mochizuki N, Nakata M, Yano T, Fujii R, Takakura N. (2008). Spatial and temporalrole of the apelin/APJ system in the caliber size regulation of blood vessels during angiogenesis. EMBO J. 2008 Feb
6;27(3):522-34.
ĳĺResearch & Activities

Division of Cellular and Molecular Biology

Department of Cellular Regulation
Research Group

Professor
Assistant Professor
Assistant Professor
SA Researcher
SA Researcher

Hiroaki Miki, Ph.D.
Daisuke Yamazaki, Ph.D.
Yosuke Funato, Ph.D.
Kanami Uesugi, Ph.D.
Yusuke Hirata, Ph.D.

Cells are equipped with a signal transduction system that enables them to respond appropriately to the surrounding
environment, such as stimulation with various hormones/growth factors and physical interaction with other cells or the
extracellular matrix. Malfunctions in the signaling system are responsible for various human diseases, such as in the case of theabnormal proliferation of cancer cells. In our laboratory, we are investigating the mechanism of intracellular signaling that
regulates the proliferation, differentiation, and motility of cells at levels ranging from molecules to organisms using nematodes
and mice. Our present major research interests are (1) oxidative stress signaling by reversible oxidation of proteins, and (2)cancer metastasis and Mg2+-regulation by redox-responsive proteins.
(1) Oxidative stress signaling by reversible oxidation of proteins
We discovered thioredoxin-related protein nucleoredoxin (NRX) as a novel regulator of Wnt signaling, which plays importantroles in early development and oncogenesis. We observed that NRX directly bound to the Wnt signal transducer Dishevelled(Dvl) and inhibited its function. Interestingly, the NRX-Dvl interaction was negatively regulated by the formation of
intramolecular S-S bonds in NRX. Thus, NRX regulates Wnt signaling in a redox-dependent manner (Fig. 1).
We also developed a novel method to search for proteins forming S-S bonds in cells by using thioredoxin mutants. With thismethod, we identified several proteins such as CRMP2, which functions in Semaphorin signaling. Semaphorin treatment
stimulated H2O2 generation and CRMP2 oxidation, resulting in the formation of homodimers linked with S-S bonds. This
oxidation of CRMP2 mediated the repulsive axon guidance induced by Semaphorin stimulation (Fig. 2).

Fig. 1: Redox regulation of Wnt signaling by reversible oxidation of NRX(A) Developmental abnormality of bones in NRX-/- mice. (B) Schematic illustration of the mechanism of redox regulation of Wnt signalingE\15;15;LQKLELWVWKHIXQFWLRQRI'YO7KHSURWHLQFRPSOH[FRPSRVHGRI$3&$[LQDQG*6.LQGXFHVWKHGHJUDGDWLRQRIȕFDWHQLQ:KHQ15;LVR[LGL]HG'YOEHFRPHVIUHHIURP15;DQGLQKLELWVWKHGHJUDGDWLRQRIȕFDWHQLQZKLFKLQGXFHVWKHH[SUHVVLRQRILWVWDUJHW
genes.

Ĵı

Research & Activities

Fig. 2: CRMP2 oxidation in Semaphorin signaling(A) Repulsive guidance of axons by Semaphorin 3A (Sema3A). (B) Increase of H2O2 in neuronal growth cones by Sema3A. (C) Schematic
illustration of the mechanism of Semaphorin signaling via CRMP2 oxidation. Sema3A stimulation generates H2O2 and oxidizes CRMP2.
Oxidized CRMP2 transiently forms protein complexes with TRX, which induces CRMP2 phosphorylation by GSK3.

(2) Cancer metastasis and Mg2+-regulation by redox-responsive protein
We identified PRL, a tyrosine phosphatase with unknown function, as a novel S-S bond-containing protein. PRL is
consistently overexpressed in various human metastatic cancers and can promote experimental metastasis. We searched fornovel binding proteins for PRL and identified magnesium-exporting protein (MagEx), an integral membrane protein that
regulates the intracellular Mg2+ level by exporting Mg2+. The activity of MagEx was inhibited by its interaction with PRL.

Recent Publications
1. Morinaka A, Funato Y, Uesugi K, Miki H. Oligomeric peroxiredoxin-I is an essential intermediate for p53 toactivate MST1 kinase and apoptosis. Oncogene. 2011 Oct 6;30(40):4208-18.
2. Morinaka A, Yamada M, Itofusa R, Funato Y, Yoshimura Y, Nakamura F, Yoshimura T, Kaibuchi K, Goshima Y,
Hoshino M, Kamiguchi H, Miki H. Thioredoxin mediates oxidation-dependent phosphorylation of CRMP2 andgrowth cone collapse. Sci Signal. 2011 Apr 26;4(170):ra26.
3. Funato Y, Terabayashi T, Sakamoto R, Okuzaki D, Ichise H, Nojima H, Yoshida N, Miki H. Nucleoredoxin sustains:QWȕFDWHQLQVLJQDOLQJE\UHWDLQLQJDSRRORILQDFWLYHGLVKHYHOOHGSURWHLQCurr Biol. 2010 Nov 9;20(21):1945-52.
4. Hayashi T, Funato Y, Terabayashi T, Morinaka A, Sakamoto R, Ichise H, Fukuda H, Yoshida N, Miki H.
Nucleoredoxin negatively regulates Toll-like receptor 4 signaling via recruitment of flightless-I to myeloid
differentiation primary response gene (88). J Biol Chem. 2010 Jun 11;285(24):18586-93.
5. Yoshimura Y, Terabayashi T, Miki H. Par1b/MARK2 phosphorylates kinesin-like motor protein GAKIN/KIF13B toregulate axon formation. Mol Cell Biol. 2010 May;30(9):2206-19.
ĴĲResearch & Activities

Research Center for Infectious Disease Control

Department of Molecular Protozoology
Research Group

Professor
Invited Professor
Assistant Professor
Assistant Professor
SA Assistant Professor
SA Researcher

Toshihiro Horii, Ph. D.
Kazuyuki Tanabe, Ph. D.
Nobuko Arisue, Ph. D.
Takahiro Tougan, Ph. D.
Nirianne M. Q. Palacpac, Ph. D.
Masanori Yagi, Ph. D

Research Projects
Malaria is a serious threat to global human health. More than 40% of the world’ s population lives in malaria-endemic areas,
and two million people succumb to the disease annually. Controlling malaria has become more challenging since theemergence of drug-resistant malaria parasites. This development has intensified the need for novel drug targeting strategies
and an effective malaria vaccine. Our department focuses on both malaria vaccine and anti-malarial drug development. We
also seek to understand the mechanisms used by the malaria parasite to survive in the host.
(1) Development of a recombinant vaccine based on the malaria protein SERA
We are developing a malaria vaccine based on SE36, a recombinant protein that corresponds to an amino acid sequence
present in the serine repeat antigen (SERA) of malaria parasites. We and coresearchers in malaria-endemic areas havedemonstrated that naturally acquired immunity against malaria exclusively correlates with the development of anti-SERA IgG3
antibodies. We have also shown that many types of animals, including chimpanzees, develop antibodies upon vaccination with
SE36 and that these antibodies inhibit the growth of malaria parasites.
Together with the Kanonji Institute of The Research Foundation of Osaka University, we constructed a system by which theSE36 malaria vaccine can be mass produced. In 2005, we conducted a phase I clinical trial in Japan with SE36 to assess its
safety and immunogenicity. All vaccine-administered volunteers sero-converted and showed no serious adverse events. We aremoving towards further phase Ib clinical trials in an endemic region in Uganda. This project is being conducted incollaboration with The Research Foundation for Microbial Diseases of Osaka University.
We are also studying the function of the SERA molecule in the parasite and characterizing the host immune response against
SERA. In addition, we have started a new research project for P. vivax vaccine development by collaborating with colleagues
in Uganda, Thailand, Indonesia, and the Solomon Islands.

Fig. 1. Patients at Apac hospital (Uganda). Of
the patients waiting at the Out-patient
Department of Apac Hospital in Northern
Uganda, majority of victims are children under
5 years old.
Fig. 2. Processed fragments of P. falciparum SERA and the structure ofrecombinant SE36

Ĵĳ

Research & Activities

Fig. 4. The SE36 malaria vaccine for clinical
trials is produced under Good Manufacturing
Practices (GMP) conditions at the Kanonji
Institute of The Research Foundation forMicrobial Diseases of Osaka University.
Fig. 3. P. falciparum life cycle and vaccine targets(2) Approaches for understanding theevolution of malaria parasites
Malaria parasites have many unique features, such as a highly
restricted host range, complex life cycle, and host immune evasion
systems. To understand the molecular basis of those features, we
utilized several genetic approaches, such as nuclear and organelle
genome analyses, as well as population genetic analyses of theantigen coding genes. The findings from these studies can beeffectively utilized in vaccine development.

Fig. 5. Evolutionary tree of malaria parasites inferred from 30 protein-encoding
genes from the apicoplast genome. Four human parasites do not show amonophyletic relationship revealing host-switching during parasite evolution.

Recent Publications
1. Arisue N, Hashimoto T, Mitsui H, Palacpac NM, Kaneko A, Kawai S, Hasegawa M, Tanabe K, Horii T. The
Plasmodium apicoplast genome: conserved structure and close relationship of P. ovale to rodent malaria parasites.
Mol Biol Evol. 2012 Mar 6. (in press)
2. Tanabe K, Arisue N, Palacpac NM, Yagi M, Tougan T, Honma H, Ferreira MU, Färnert A, Björkman A, Kaneko A,
Nakamura M, Hirayama K, Mita T, Horii T. Geographic differentiation of polymorphism in the Plasmodium
falciparum malaria vaccine candidate gene SERA5. Vaccine. 2012 Feb 21;30(9):1583-93.
3. Palacpac NM, Arisue N, Tougan T, Ishii KJ, Horii T. Plasmodium falciparum serine repeat antigen 5 (SE36) as amalaria vaccine candidate. Vaccine. 2011 Aug 11;29(35):5837-45.
4. Arisue N, Kawai S, Hirai M, Palacpac NM, Jia M, Kaneko A, Tanabe K, Horii T. Clues to evolution of the SERA
multigene family in 18 Plasmodium species. PLoS One. 2011 Mar 15;6(3):e17775.
5. Horii T, Shirai H, Li J, Ishii KJ, Palacpac NM, Tougan T, Hato M, Ohta N, Bobogare A, Arakaki N, Matsumoto Y,
Namazue J, Ishikawa T, Ueda S, Takahashi M. Evidences of protection against blood-stage infection of Plasmodium
falciparum by the novel protein vaccine SE36. Parasitol Int. 2010 Sep;59(3):380-6.
ĴĴResearch & Activities

Research Center for Infectious Disease Control

Department of Virology
Research Group

Professor
Assistant Professor
Assistant Professor
SA Assistant Professor
SA Assistant Professor
SA Researcher
SA Researcher
SA Researcher

Kazuyoshi Ikuta, Ph.D.
Takeshi Kurosu, D.V.M., Ph.D.
Yohei Watanabe, D.V.M., Ph.D.
Tadahiro Sasaki, Ph.D.
Azusa Asai, Ph.D.
Yuji Inoue, Ph.D.
Yuta Kanai, D.V.M., Ph.D.
Mitsuhiro Nishimura, Ph.D.

Research in our department focuses on several viruses that target immune and respiratory regions, as well as prions that target
the central nervous system. We seek to understand the mechanisms by which these pathogens replicate and induce
pathogenesis, and to apply our understanding to the development of methods to control these agents, remove them from blood
products, and diagnosis their infection rapidly.
(1) Infections to the immune system
Dengue illness, which is transmitted to humans by mosquitoes, is among the most important viral infectious diseases intropical and subtropical regions around the world. There are four antigenically distinct dengue virus serotypes. Severe dengue
cases, such as dengue hemorrhagic fever and dengue shock syndrome, mostly occur by secondary infection with a serotype
different from the primary infected serotype. Although individuals infected with dengue virus produce high titers ofneutralizing antibodies, some of these antibodies may play a role in the antibody-dependent enhancement of the secondarily
infected serotype, which could make vaccine development difficult.
We seek to characterize the mechanism by which dengue virus derived from South Asian countries induces dengue illness inorder to develop an antidengue viral drug. For its in vivo evaluation, we are also developing animal models for dengue viral
infection. In addition, we are preparing human monoclonal antibodies to understand the viral-induced pathogenicity and todevelop therapeutic antibodies.
(2) Infections to the respiratory system
Influenza virus induces typical acute infection in the respiratory region. In addition to seasonal influenza viruses, aswine-origin pandemic virus appeared in 2009. Currently, the possible global emergence of a pandemic virus from highly
pathogenic avian influenza virus, such as H5N1, is a public concern worldwide. Together with Alexandria University, we are
studying H5N1 circulating in Egypt. Recently, we found that some of the new H5 sublineages in Egypt have acquired anenhanced binding capacity to human-type receptors. The purpose of our search is to ascertain the pandemic potential of these
H5N1 viruses in Egypt. We have prepared several human neutralizing monoclonal antibodies against influenza virus. Because
the epitope region recognized by one of the monoclonal antibodies is highly conserved and forms a conformational structure,
we are working on the possible development of a new vaccine with this conformation in the synthetic peptide in collaboration
with several companies.
(3) Blood-borne infections
We are working to establish how to remove infectious agents, such as parvovirus B19, SARS-corona virus, hepatitis E virus,
and prions, from blood products in collaboration with a company.
(4) Rapid diagnosis kits
There are many techniques for the diagnosis of viral infections, including immunofluorescence, ELISA, Western blot, and
PCR assays. We are currently working with several companies towards the development of rapid diagnosis kits against several
infectious diseases.

ĴĵResearch & Activities

Dengue virus is produced as a single polyprotein and is cleaved into 10 proteins for viral replication. The dengue virus protease complex, which is essential forcleavage, is composed of the NS3 protease and NS2B cofactor. Small molecules that interfere with the interaction between NS2B and NS3 impair proteaseactivity and inhibit viral replication. NS2B (frame structure) interacts with the pocket site of NS3 (green). The key residues of NS3 that interact with the newly
developed small compound are indicated in pink.

Dengue viral infection induces strong humoral immunity and antibody production. These
antibodies include neutralizing antibodies and negatively regulating antibodies by
antibody-dependent enhancement. Human monoclonal antibodies are being prepared by using
immune cells from patients to investigate dengue virus-induced pathogenicity and to develop
antibody therapeutics.

Some of the new H5 sublineages in Egypt have acquired an increased attachment to and infectivity in the human lower respiratory tract (H5N1 virus attachedto human tracheal epithelia is stained red in the panel). We are currently investigating the mechanism(s) underlying the possible emergence of pandemic H5N1
viruses in Egypt and its risk in the field, in collaboration with Alexandria University. (a) Pathology of the classical virus lineage (H5N1) in humans. The virus
OLQHDJHRQO\KDVELQGLQJDIILQLW\IRUĮVLDO\OJO\FDQDQGQHHGVWRUHDFKWKHORZHUUHVSLUDWRU\WUDFW E 3DWKRORJ\RIWKHQHZ+VXEOLQHDJHWKDWKDVHPHUJHG
LQ(J\SW([SDQVLRQRIUHFHSWRUXVDJH LQFUHDVHGĮVLDO\OJO\FDQELQGLQJ HQDEOHVWKHQHZVXEOLQHDJHWRELQGPRUHHIILFLHQWO\WRHSLWKHOLDLQWKHORZHU
respiratory tract.

Recent Publications
1. Yasugi M, Nakamura S, Daidoji T, Ramadhany R, Yang C-S, Yasunaga T, Iida T, Horii T, Ikuta K, Takahashi K,
Nakaya T. Frequency of D222G and Q223R hemagglutinin mutants of pandemic (H1N1) 2009 influenza virus in
Japan between 2009 and 2010. PLoS One  H(SXE)HE
2. Watanabe Y, Ibrahim MS, Suzuki Y, Ikuta K. The changing nature of avian influenza A virus (H5N1). Trends
Microbiol. 2012 Jan;20(1):11-20. Epub 2011 Dec 5.
3. Watanabe Y, Ibrahim MS, Ellakany HF, Kawashita N, Mizuike R, Hiramatsu H, Sriwilaijaroen N, Takagi T, Suzuki
Y, Ikuta K. Acquisition of human-type receptor binding specificity by new H5N1 influenza virus sublineages during
their emergence in birds in Egypt. PLoS Pathog0D\  H(SXE0D\4. Mizuike R, Sasaki T, Baba K, Iwamoto H, Shibai Y, Kosaka M, Kubota-Koketsu R, Yang CS, Du A, Sakudo A,
Tsujikawa M, Yunoki M, Ikuta K. Development of two types of rapid diagnostic test kits to detect the hemagglutinin
or nucleoprotein of the swine-origin pandemic influenza A virus H1N1. Clin. Vaccine Immunol0DU
(3):494-9. Epub 2011 Jan 12.
5. Kurosu T, Khamlert C, Phanthanawiboon S, Ikuta K, Anantapreecha S. Highly efficient rescue of dengue virus using
a co-culture system with mosquito/mammalian cells. Biochem Biophys Res Commun$SU  
Epub 2010 Mar 7.
ĴĶResearch & Activities

Genome Information Research Center

Department of Experimental Genome Research
Research Group

Professor (SUP)
Associate Professor
Associate Professor (SUP)
Assistant Professor
Assistant Professor
Assistant Professor (SUP)SA Assistant Professor (SUP)

Masaru Okabe, Ph.D.
Takeshi Miwa, Ph.D.
Masahito Ikawa, Ph.D.
Hidetoshi Hasuwa, Ph.D.
Naokazu Inoue, Ph.D.
Ayako Isotani, Ph.D.
Yuhkoh Satouh, Ph.D.

In the past, naturally mutated animals have been used to elucidate the mechanisms of various diseases. However, in the"postgenome project era", genetically manipulated animals play a key role in such investigations and provide important diseasemodels. Our laboratory, in collaboration with the Animal Resource Center for Infectious Diseases, elucidates the mechanismsof mammalian reproductive systems through gene manipulation in mice (http://www.egr.biken.osaka-u.ac.jp/).
Research Projects
We have successfully produced the first genetically altered “green mouse” in the world. These green fluorescent protein(GFP)-expressing mice are highly useful for many types of research projects, such as those involving stem cell transplantation
and regeneration studies. Utilizing one of these animals, we demonstrated that it was possible to label sperm with fluorescentproteins and visualize the fertilization process (Figures 1 and 2).
Using gene knockout technology, we showed that testis-specific chaperones (CLGN, CALR3, and PDILT) are required for thequality control of the sperm membrane protein ADAM3 in the endoplasmic reticulum. Mutant mice were male infertile
because their spermatozoa lacking ADAM3 could not migrate through the utero-tubal junction (Figure 2) (1). However, mutant
spermatozoa were able to fertilize eggs when they were directly deposited into the oviduct.
Peculiarly, spermatozoa with defective zona pellucida (ZP)-binding ability were able to fertilize cumulus-covered, ZP-intact
eggs effectively in vivo (1). Acrosome-reacted mouse spermatozoa that were recovered from the perivitelline space could be
used to fertilize other eggs (Figure 3) (2). From these results, we propose that the observation of numerous sperm binding to
the ZP surface of cumulus-free eggs could have been a “red herring” that was less important than was previously supposed.

Fig. 1. Transgenic mouse spermatozoa
labeled with GFP in the acrosome and red
fluorescent protein (RFP) in the
mitochondria. Spermatozoa lose green
fluorescence after acrosome reaction.

Fig. 2. Pdilt -/- spermatozoa lacking ADAM3 are unable to migrate through the utero-tubal junction.

Fig. 3. Acrosome-reacted mouse spermatozoa recovered from the perivitelline space can fertilize other eggs.

ĴķFig. 4. Accumulation of many Izumo1 -/spermatozoa in the perivitelline space,because Izumo1-/- spermatozoa are unable
to fuse with egg.

Research & Activities

We have also reported that the sperm protein IZUMO1 is essential for fusion with eggs, and that SPESP1 is necessary to
produce fully “fusion-competent” sperm (Figure 4).
In addition to studying the mechanism of the sperm-egg interaction, we are trying to invent new technologies to perform novel
investigations in the field of reproduction. By transducing blastocysts with lentiviral vectors, we developed theplacenta-specific gene manipulation method. Expression of soluble FLT1 in the placenta allowed us to prepare preeclampsia
model mice (3).
:HUHFHQWO\HVWDEOLVKHGUDWHPEU\RQLFVWHP (6 FHOOVDQGJHQHUDWHG0RXVHļ5DWFKLPHULFDQLPDOV )LJXUH :HZRXOGOLNH
to use this animal model to study body/organ size control in vivo. The chimeric animals may provide a method for thederivation of various organs from ES or iPS cells (4). We are also examining the role(s) of miRNA in live animals using
miRNA knockout technique.

)LJ0RXVHļ5DWFKLPHULFDQLPDOVDUHJHQHUDWHGE\LQMHFWLQJUDW(6FHOOVLQWRPRXVHEODVWRF\VWV OHIW &RQWULEXWLRQRIPRXVH ZKLWH DQG
rat (black) cells are visualized by coat color.

Research Projects (Miwa’ s group)
We investigate the molecular biological mechanisms involved in human diseases, especially cardiovascular diseases, by using
animal models. We have established a diastolic heart failure model using Dahl salt-sensitive rats. Left ventricular (LV) fibrosis
and stiffening play crucial roles in the development of heart failure with preserved ejection fraction (HFpEF). Digitalis-likefactors and subsequently activated Na+ /Ca 2+ exchanger entry mode may play an important role in the development of
K\SHUWHQVLYH+)S()  :HKDYHDOVRDGPLQLVWHUHGFDUQLWLQHWRWKH+)S()PRGHO )LJXUH (a)Control

HFpEF

Carnitine)LJ(IIHFWVRIFDUQLWLQHRQFDUGLDFILEURVLVLQ/9RI'DKO
salt-sensitive rats at age 20 weeks. The typical Azan Mallory
staining of the LV in the control, HFpEF, and carnitine treatmentgroups. Carnitine administration significantly attenuated LV
fibrosis.

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Recent Publications
1. Tokuhiro K, Ikawa M, Benham AM, Okabe M. Protein disulfide isomerase homolog PDILT is required for quality
control of sperm membrane protein ADAM3 and male infertility. Proc Natl Acad Sci U S A0DUFK 
2. Inoue N, Satouh Y, Ikawa M, Okabe M, Yanagimachi R. Acrosome-reacted mouse spermatozoa recovered from theperivitelline space can fertilize other eggs. Proc Natl Acad Sci U S A'HF  
3. Kumasawa K, Ikawa M, Kidoya H, Hasuwa H, Saito-Fujita T, Morioka Y, Takakura N, Kimura T, Okabe M.
Pravastatin induces placental growth factor (PGF) and ameliorates preeclampsia in a mouse model. Proc Natl Acad
Sci U S A-DQ  
4. Isotani A, Hatayama H, Kaseda K, Ikawa M, Okabe M. Formation of a thymus from rat ES cells in xenogeneic nude
PRXVHļUDW(6FKLPHUDVGenes Cells$SU  
 .DPLPXUD'2KWDQL70LZD7HWDO&D(QWU\PRGHRI1D&DH[FKDQJHUDVDQHZWKHUDSHXWLFWDUJHWIRU
heart failure with preserved ejection fraction. European Heart J. 2011 Apr 13.
ĴĸGenome Information Research Center

Research & Activities

Department of Genome Informatics
Research Group

Professor
Professor (SUP)
Assistant Professor
Assistant Professor
Assistant Professor (SUP)SA Researcher
SA Researcher

Teruo Yasunaga, Ph.D.
Tatsuya Takagi, Ph.D.
Naohisa Goto, Ph.D.
Shota Nakamura, Ph.D.
Norihito Kawashita, Ph.D.
Akifumi Yamashita, Ph.D.
U. Chandimal de Silva, Ph.D.

Research Projects
Our group studies the genome information of various organisms by using high-performance computers, to identify new
biological phenomena and to understand how organisms evolve. In addition, we develop software tools for bioinformatics andmolecular biology. Our laboratory operates a computer system for genome sequence data analyses that is made available toresearchers in our university, and holds training courses for genome analysis at least once every year.
(1) Large-scale genome analysis
The complete genome sequences of more than 1,000 organisms are currently available. By using bioinformatics and molecular
evolution techniques, we intend to analyze this enormous body of genome data. We are developing software and algorithms forlarge-scale genome analysis. In particular, we developed CONSERV, a conserved sequence finder. By analyzing the complete
genome sequences of 266 organisms, we identified invariant sequences that may have been present in the last common
ancestor of all extant life forms (Goto et al, 2007; Fig. 1). Through a comprehensive analysis of its genome, we also intend to
elucidate the evolutionary pathways of the influenza virus (Fig. 2).

Figure 1. Large-scale genome analysis of 266 organisms. Identification ofsequences conserved in almost all known genomes.

ĴĹ

Figure 2. Comprehensive analysis of the influenza virus genome.

Research & Activities(2) Next generation sequencer data analysis
The recently developed “next generation sequencing” technology has enabled sequencing of the whole genome of any
microorganism in one sequencer run, producing massive amounts of nucleotide sequence data in each run. We have beendeveloping software to handle these data and have set up an analysis system for collaborative sequencing projects of
microorganisms with other laboratories (Fig. 3)(3) Operation of a computer system for genome information analysis at Osaka University.
We provide computer resources for researchers in our university. We also provide mirrored access to major nucleotide, protein,
and genome databases through our servers (Fig.4) which are fully synchronized with the mother servers and kept up to date at
all times.

Figure 3. Next generation sequencing enables
sequencing of a whole genome in one run.

Figure 4. Genome Information Research Center Computer System:
Analysis servers (left) and 1.8PB storage system (right).

Recent Publications
1. de Silva UC, Tanaka H, Nakamura S, Goto N, Yasunaga T. A comprehensive analysis of reassortment in influenza A
virus. Biol Open. 2012 Feb 23;1(4):385-90.
2. Yamashita A, Kawashita N, Kubota-Koketsu R, Inoue Y, Watanabe Y, Ibrahim MS, Ideno S, Yunoki M, Okuno Y,
Takagi T, Yasunaga T, Ikuta K. Highly conserved sequences for human neutralization epitope on hemagglutinin of
influenza A viruses H3N2, H1N1 and H5N1: Implication for human monoclonal antibody recognition. Biochem
Biophys Res Commun. 2010 Mar 19;393(4):614-8.
3. Nakamura S, Yang CS, Sakon N, Ueda M, Tougan T, Yamashita A, Goto N, Takahashi K, Yasunaga T, Ikuta K,
Mizutani T, Okamoto Y, Tagami M, Morita R, Maeda N, Kawai J, Hayashizaki Y, Nagai Y, Horii T, Iida T, Nakaya
T. Direct metagenomic detection of viral pathogens in nasal and fecal specimens using an unbiased high-throughput
sequencing approach. PLoS One. 2009;4(1):e4219.
4. Yamashita A, Goto N, Nishiguchi S, Shimada K, Yamanishi H, Yasunaga T. Computational search forover-represented 8-mers within the 5'-regulatory regions of 634 mouse testis-specific genes. Gene. 2008 Dec 31;427(1-2):93-8.
5. Goto N, Kurokawa K, Yasunaga T. Analysis of invariant sequences in 266 complete genomes. Gene. 2007 Oct
15;401(1-2):172-80.
ĴĺResearch & Activities

Genome Information Research Center

Department of Infection Metagenomics
Research Group

Professor (SUP)
Professor (SUP)SA Professor (SUP)
Assistant Professor (SUP)
Assistant Professor (SUP)

Toshihiro Horii, Ph.D.
Teruo Yasunaga, Ph.D.
Tetsuya Iida, Ph.D.
Naohisa Goto, Ph.D.
Shota Nakamura, Ph.D.

Research Projects
1.RAPID (Robotics Assisted Pathogen IDentification)
Under the aegis of the Program of Research Centers for Emerging and Re-emerging Infectious Diseases of MEXT, Japan, andin collaboration with the Omics Science Center, RIKEN, we are constructing a framework called “RAPID” that will facilitate
the emergency diagnosis of infectious diseases. We are also cooperating with the research centers of eight countries in Asia
and Africa in an effort to identify the causative agents in naturally occurring outbreaks.
2.Metagenomic Diagnosis of Infectious Diseases
Metagenomic analysis allows us to diagnose many of the major human infectious diseases (including respiratory tract
infections, enteric infections, and blood-borne infections) by using a single common protocol. In addition, to pre-empt zoonotic
disease outbreaks, we are seeking to identify new pathogenic microorganisms in animal-derived samples that may have
zoonotic potential.
3.Metagenomic Analysis of the Intestinal Microbiome
The intestinal microbiome plays an important role in protecting the host from pathogen invasion. We are currently analyzing
the intestinal microbiome of patients with diarrheal diseases to elucidate how the human host, intestinal microbiome, andpathogenic microorganisms interact. This analysis will help us to understand the changes that occur in the intestinal
microbiome during the course of infection.
4.Development of Novel Methods for Pathogen Detection
To develop more efficient and comprehensive methods of identifying pathogens, we are studying the efficacy of different
methods to amplify the genome of pathogenic microorganisms and subtract the host genome.

Extract DNA or RNA

Next-generation sequencer

DATA analysis

Fig. 1. Metagenomic diagnosis of infectious diseases using a next-generation sequencer.

ĵıResearch & Activities

Fig. 2. The next-generation sequencers installed in our department, Roche 454 GS Junior Bench Top System, illumina MiSeq Personal Sequencer,
Pacific Biosciences RS System.

Fig. 3. Distribution of detected organisms in viral infection cases
Fig. 4. Standard operating protocol for RAPID

Recent Publications
1. Yasugi M, Nakamura S, Daidoji T, Kawashita N, Ramadhany R, Yang CS, Yasunaga T, Iida T, Horii T, Ikuta K,
Takahashi K, Nakaya T. Frequency of D222G and Q223R Hemagglutinin Mutants of Pandemic (H1N1) 2009
Influenza Virus in Japan between 2009 and 2010. PLoS One. 2012;7(2):e30946.
2. Monira S, Nakamura S, Gotoh K, Izutsu K, Watanabe H, Alam NH, Endtz HP, Cravioto A, Ali SI, Nakaya T, Horii
T, Iida T, Alam M. Gut microbiota of healthy and malnourished children in bangladesh. Front Microbiol.
2011;2:228.
3. Nakamura S, Nakaya T, Iida T. Metagenomic analysis of bacterial infections by means of high-throughput DNA
sequencing. Exp Biol Med (Maywood). 2011;236(8):968-71.
4. Nakamura S, Yang CS, Sakon N, Ueda M, Tougan T, Yamashita A, Goto N, Takahashi K, Yasunaga T, Ikuta K,
Mizutani T, Okamoto Y, Tagami M, Morita R, Maeda N, Kawai J, Hayashizaki Y, Nagai Y, Horii T, Iida T, Nakaya
T. Direct metagenomic detection of viral pathogens in nasal and fecal specimens using an unbiased high-throughput
sequencing approach. PLoS One. 2009;4(1):e4219.
5. Nakamura S, Maeda N, Miron IM, Yoh M, Izutsu K, Kataoka C, Honda T, Yasunaga T, Nakaya T, Kawai J,
Hayashizaki Y, Horii T, Iida T. Metagenomic diagnosis of bacterial infections. Emerg Infect Dis. 2008 Nov;14(11):1784-6.
ĵĲResearch & Activities

International Research Center for Infectious Diseases
Department of Infectious Disease Control

Laboratory for Clinical Research on Infectious Diseases
Research Group

Invited Professor
SA Associate Professor
SA Researcher
SA Researcher
SA Researcher

Kazunori Oishi M.D., Ph.D.
Yukihiro Akeda Ph.D.
Tatsuya Nakayama Ph.D.
Zhenyu Piao Ph.D.
Dan Takeuchi M.D., Ph.D.

The research activities in our group include the diagnosis, pathogenesis, and prevention of invasive pneumococcal diseases(IPD). We also seek to investigate the epidemiology and pathogenesis of Streptococcus suis infection, the pathogenesis ofthrombocytopenia in dengue illness, and the mechanism of protein secretion systems of pathogenic bacteria. In addition, ourgroup is registered as a member of the World Health Organization (WHO)/Global Outbreak Alert & Response Network(GOARN) and will join the outbreak response team in the global effort to control emerging and re-emerging infectious
diseases.
1) Diagnosis, pathogenesis, and prevention of IPD
1. IPD and pneumococcal pneumonia in children and adults
We reported the effects of 23-valent pneumococcal polysaccharide vaccine (PPV) on the incidence and medical cost for allcauses of pneumonia among elderly people receiving a routine influenza vaccine. We are currently conducting a prospective
study of the measurement of IgG and opsonophagocytic activity to the infecting serotype in sera of pediatric patients with IPD.
In particular, we are analyzing the pathogenesis of IPD before and after vaccination with 7-valent conjugate vaccine. We arealso conducting a prospective, multicenter study to determine the infecting serotype by PCR using clinical specimens and theprotective threshold concentration of pneumococcal antibody in adult patients with IPD and pneumococcal pneumonia.
2. Development of a pneumococcal surface protein A (PspA) vaccine
PspA can elicit protective antibodies in animals. We reported the protective effects of PspA nasal vaccine in a mouse model ofsecondary pneumonia after influenza virus infection. We aim to develop a cross-protective PspA vaccine.
2) Research on Streptococcus suis infections prevalent in Thailand
Streptococcus suis, an important zoonotic pathogen, causes invasive infections, such as meningitis, in humans. In a
retrospective study of S. suis infection in 2006–2008 in Thailand, we reported that human cases were prevalent in the northern
region. The causative pathogens were serotype 2 for 165 cases (92.2%) and serotype 14 for 12 cases (6.7%) (Emerg Infect Dis
2011). One case each of previously unreported serotypes 5 and 24 were confirmed in patients with liver cirrhosis (Lancet2011). Analysis of serotype 2 infection demonstrated that 58.9% of 158 patients were associated with meningitis and the others
were associated with sepsis. The sequence type (ST)1 and ST104 strains were found to be capable of causing sepsis, but only
the ST1 strain commonly caused meningitis (Figure 1). In a population-based study in Phayao Province in 2010, 31 humancases were confirmed, with a case fatality of 16.1%. The estimated incidence rate was 6.2 per 100,000 in the general
population (PLoS ONE, 2012). Consumption of raw pork products was confirmed in 22 cases. The incubation period was 2days after the consumption of raw pork products. We are aiming at eliminating this disease from northern Thailand byeducating people about the risks of eating raw pork dishes.
3) Mechanisms by which dengue viral infections lead to thrombocytopenia
Dengue illness is a major public health concern, particularly in tropical countries, and the mechanism of thrombocytopenia
remains uncertain. Based on a clinical study in the Philippines, we reported a significant association between
thrombocytopenia and increased phagocytosis of platelets in an ex vivo setting during the acute phase of secondary dengue
virus infection. We recently reported increased phagocytosis of DV-induced apoptotic platelets by macrophages via a
PS-recognizing pathway in secondary DV infection (Figure 2). Accelerated platelet clearance was overcome bythrombopoietin-induced enhanced thrombopoiesis in these patients (J Infect Dis, 2012).
4) Protein secretion systems of pathogenic bacteria
The development of bacterial infections requires many virulence factors. Because most such factors are proteins secreted fromĵĳFigure 1. Streptococcus suis infection in Thailand: distribution
and pathogenesis (Emerg Infect Dis 2011).

Research & Activities

pathogenic bacteria, it is essential to study the mechanisms of protein secretion systems in pathogenic bacteria during theinfection process. In our laboratory, a food-borne pathogen, Vibrio parahaemolyticus, and a causative agent of pneumonia,
Streptococcus pneumoniae, are used to study their protein secretion systems and secreted virulence factors. We recently
identified a novel chaperone responsible for virulence factor secretion (FEMS Microbiol 2011). We have also been
investigating the involvement of secretion system-associated ATPase on the protein secretion mechanism.

Figure 2. Thrombocytopenia and apoptotic platelet
clearance in dengue (J Infect Dis, 2012).

Recent Publications
1. Alonzo MTG, Lacuesta TLV, Dimaano EM, Kurosu T, Suarez LC, Mapua CA, Akeda Y, Matias RR, Kuter DJ,
Nagata S, Natividad FF, Oishi K. Platelet apoptosis and apoptotic platelet clearance by macrophages in secondary
dengue virus Infections. J Infect Dis 2012 Apr;205(8):1321-9.
2. Takeuchi D, Kerdsin A, Pienpringam A, Loetthong P, Samerchea S, Pakkinee Loetthong P, Khamisra K, Wongwan
N, Areeratana P, Chiranairadul P, Lertchayanti S, Petcharat S, Yowang A, Chaiwongsaen P, Nakayama T, Yukihiro
Akeda Y, Hamada S, Sawanpanyalert P, Dejsirilert S, Oishi K. Population-based Study of Streptococcus suis
Infection in Humans in Phayao Province in Northern Thailand. PLoS ONE. 2012; 7(2):e31265.
3. Kerdsin A, Dejsirilert S, Sawanpanyalert P, Boonnark A, Noithachang W, Sriyakun D, Simkum S, Chokngam S,
Gottschalk M, Akeda Y, Oishi K. Sepsis and spontaneous bacterial peritonitis in Thailand. Lancet 2011 Sep 3; 378(9794):960.
4. Kerdsin A, Dejsirilert S, Puangpatra P, Sripakdee S, Chumla K, Boonkerd N, Polwichai P, Tanimura S, Takeuchi D,
Nakayama T, Nakamura S, Akeda Y, Gottschalk M, Sawanpanyalert P, Oishi K. Genotypic profile of Streptococcus
suis serotype 2 and clinical features of infection in humans, Thailand. Emerg Infect Dis. 2011 May 17(5):835-42.
5. Akeda, Y, Kodama T, Saito K, Iida T, Oishi, K, Honda T. Identification of the Vibrio parahaemolyticus Type III
secretion system 2-associated chaperone VocC for the T3SS2-specific effector VopC. FEMS Microbiol Lett 2011
Nov; 324(2):156-64.
ĵĴResearch & Activities

International Research Center for Infectious Diseases
Department of Special Pathogens

Laboratory for Infection Cell Biology
Research GroupSA Professor
SA Assistant Professor
SA Assistant Professor
SA Researcher

Yukako Fujinaga, Ph.D.
Yo Sugawara, Ph.D.
Takuhiro Matsumura, Ph.D.
Masahiro Yutani, Ph.D.

Research Projects: Many bacterial toxins are able to damage the host severely, even at very low concentrations. Most bacterial
toxins are enzymes that act catalytically and with high specificity on the functional host cell molecules, thereby markedly
modulating host homeostasis. The toxins are also often highly efficient in accessing their target molecule in the host. The
ingenious transport systems involved often exploit the fundamental machinery of membrane trafficking and the functions of
intracellular organelles. Studies that elucidate toxin trafficking could provide valuable information about basic cellularfunction, aid our understanding of the pathology induced by these toxins, and help in the development of effective therapeuticstrategies against them. Our laboratory group is studying the transport mechanisms of the botulinum neurotoxin complex,
which must pass through the digestive tract and cross the epithelial barrier lining the intestine to cause food-borne botulism.

Hypothetical model for intestinal absorption of botuliunum neurotoxin complex (type B 16S toxin). We propose a three-stepmechanism by which the botulinum neurotoxin complex breaches the intestinal epithelial barrier. ① Apically located 16S toxin istranscytosed. ② Once located in the basolateral compartment, HA of the 16S toxin binds to E-cadherin, and thereby disrupts theparacellular barrier. ③ A large amount of toxin accumulates in the basolateral area via paracellular movement.

Recent Publications
1. Sugawara Y, Fujinaga Y. The botulinum toxin complex meets E-cadherin on the way to its destination. Cell Adh
Migr. 2011; 5(1): 34-36.Ȯ௙୰ȯ
2. Sugawara Y, Matsumura T, Takegahara Y, Jin Y, Tsukasaki Y, Takeichi M, Fujinaga Y. Botulinum HA disrupts theintercellular epithelial barrier by directly binding E-cadherin. J Cell Biol. 2010; 189 (4), 691-700
3. Jin Y1, Takegahara Y1, Sugawara Y, Matsumura T, Fujinaga Y. Disruption of the epithelial barrier by botulinum
hemagglutinin (HA) proteins - Differences in cell tropism and the mechanism of action between HA proteins oftypes A or B, and HA proteins of type C. Microbiology. 2009; 155(Pt 1): 35-45. 1 These authors are contributed
equally.
4. Matsumura T, Jin Y, Kabumoto Y, Takegahara Y, Oguma K, Lencer WI, Fujinaga Y. The HA proteins of botulinum
toxin disrupt intestinal epithelial intercellular junctions to increase toxin absorption. Cell Microbiol. 2008; 10(2):355-364.
5. Matsumura T1, Fujinaga Y1, Jin Y, Kabumoto Y, Oguma K. Human milk SIgA binds to botulinum type B 16S toxin
and limits toxin adherence on T84 cells. Biochem Biophys Res Commun. 2007; 352(4), 867-872.
ĵĵResearch & Activities

International Research Center for Infectious Diseases
Department of Special Pathogens

Research GroupSA Associate Professor

Eiji Morita, Ph.D.

Research & Activities

Laboratory of Viral Infection

Positive-strand RNA viruses can dramatically rearrange the intracellular membranes of the host cell and produce unusualorganelle-like structures called “replication complexes” or “membranous webs” . These membrane structures appear in close
proximity to the endoplasmic reticulum and likely serve as a scaffold for the assembly of replication machinery by providing
an organization and environment facilitating viral propagation. These structures also serve as shells that protect the viruses
against various cellular stress responses and allow persistent viral replication in the cytoplasm.
Our group has focused on the molecular mechanisms involved in the formation of these replication complexes to determine thedynamic state of viral and/or host factors during the viral propagation cycle. Recently, we successfully purified replication
complexes from cells infected with hepatitis C virus or flavivirus, and performed quantitative mass spectrometry analyses. In
these studies, we identified several cellular factors that were specifically recruited to viral replication complexes. We are
currently focusing on the molecular functions of these newly identified cellular factors on the presumption that they are
involved in the biosynthesis of the viral replication complexes. We are also studying the molecular mechanisms of viralparticle formation and the mechanisms of autophagy induction seen in virus-infected cells. These studies may contribute to thedevelopment of novel antiviral therapies.

Figure: Fluorescent (A-D) and electron (E and F) microscopic images of viral replication complexes in flavivirus-infected Vero cells. At 24
hours after infection, the cells were fixed and stained with antibodies against viral nonstructural (anti-NS2B, magenta), structural (anti-E,
green), and nuclear (DAPI, cyan) proteins. Unusual membrane structures, 5 to 10 nm in diameter and positive for viral antigens, weredetected adjacent to the nucleus in the virus-infected cells. The endoplasmic reticulum is observed at the periphery of this structure, andmany virus-like particles are detected inside the lumen.

Recent Publications
1. Morita, E., Sandrin, V., McCullough, J., Katsuyama, A., Hamilton, IB., Sundquist, WI. ESCRT-III protein
requirements for HIV-1 budding. Cell Host Microbe. 2011, 3:235-42.
2. Matsunaga, K., Morita, E., Saitoh, T., Akira, S., Ktistakis, NT., Izumi, T., Noda, T., Yoshimori, T. Autophagy
requires endoplasmic reticulum targeting of the PI3-kinase complex via Atg14L. J Cell Biol. 2010 190(4):511-21.
3. Morita, E., Colf, LA., Karren, MA., Sandrin, V., Rodesch, CK., Sundquist, WI. Human 5. ESCRT-III and VPS4
proteins are required for centrosome and spindle maintenance. Proc Natl Acad Sci U S A. 2010 107(29):12889-94.
4. Bajorek, M., Morita, E., Skalicky, JJ, Morham, SG., Babst, M., Sundquist, WI. Biochemical analyses of human IST1
and its function in cytokinesis. Mol Biol Cell 2009 20(5):1360-73.
5. Kieffer, C., Skalicky JJ., Morita, E., De Domenico, I., Ward, DM., Kaplan, J., Sundquist, WI. A New Functionally
Important ESCRT-III Recognition Mode for the VPS4 ATPases. Dev. Cell 2008, 15(1):62-73.
ĵĶResearch & Activities

International Research Center for Infectious Diseases
Department of Special Pathogens

Laboratory of Genomic Research on Pathogenic Bacteria
Research GroupSA Professor
SA Researcher
SA Researcher

Tetsuya Iida, Ph.D.
Shigeaki Matsuda, Ph.D.
Hirotaka Hiyoshi, Ph.D.

This research group is studying pathogenic bacteria from the genomic perspective. Our main research targets are as follows:
1. Characterization of the mechanism(s) used by bacterial pathogens to infect host organisms by identifying infection-relatedchanges in pathogen genome expression.
To understand the molecular mechanisms by which bacterial pathogens infect host organisms, we use DNA microarrays andother molecular methods to investigate changes in the genome expression pattern of various bacterial pathogens that occur
during their interactions with the target host.
2. Analysis of the mechanism(s) that lead to the emergence of new infectious diseases.
The unique features of various newly-emerged bacterial pathogens are studied by analyzing their genomes and comparing
them with those of other bacterial strains.
3. Investigation of the life cycles of bacterial pathogens in their natural environments
On the basis of what is currently understood about various bacterial pathogens, we seek to characterize their life cycles in theirnatural habitats.
4. Development of new methods for the rapid identification of bacterial pathogens based on genomic information.
To diagnose bacterial infections rapidly, we are developing a novel system for identifying bacterial pathogens by
high-throughput DNA sequencing.

Figure 1. Whole genome sequence of Vibrio parahaemolyticus

ĵķResearch & Activities

Figure 2. Characterization and comparison of the genomes of pathogenic bacteria by using DNA microarrays.

Recent Publications
1. Hiyoshi H, Kodama T, Saito K, Gotoh K, Matsuda S, Akeda Y, Honda T, Iida T. VopV, an F-actin-binding type III
secretion effector, is required for Vibrio parahaemolyticus-induced enterotoxicity. Cell Host Microbe. 2011 Oct
20;10(4):401-9.
2. Gotoh K, Kodama T, Hiyoshi H, Izutsu K, Park KS, Dryselius R, Akeda Y, Honda T, Iida T. Bile acid-induced
virulence gene expression of Vibrio parahaemolyticus reveals a novel therapeutic potential for bile acid sequestrants.
PLoS One. 2010 Oct 13;5(10):e13365.
3. Matsuda S, Kodama T, Okada N, Okayama K, Honda T, Iida T. Association of Vibrio parahaemolyticus
thermostable direct hemolysin with lipid rafts is essential for cytotoxicity but not hemolytic activity. Infect Immun.
2010 Feb;78(2):603-10.
4. Dryselius R, Izutsu K, Honda T, Iida T. Differential replication dynamics for large and small Vibrio chromosomes
affect gene dosage, expression and location. BMC Genomics. 2008 Nov 26;9:559.
5. Nakamura S, Maeda N, Miron IM, Yoh M, Izutsu K, Kataoka C, Honda T, Yasunaga T, Nakaya T, Kawai J,
Hayashizaki Y, Horii T, Iida T. Metagenomic diagnosis of bacterial infections. Emerg Infect Dis. 2008 Nov;14(11):1784-6.
ĵĸResearch & Activities

International Research Center for Infectious Diseases
Department of Infectious Disease Control

Laboratory of Combined Research on Microbiology and Immunology
Research Group

Associate Professor
SA Associate Professor
Postdoctoral Fellow
SA Researcher

Hiroki Nagai, Ph.D.
Tomoko Kubori, Ph.D.
Andree Marie Hubber, Ph.D.
Akiko Okura

Research Projects
Protein secretion is a process of fundamental importance for bacterial pathogenesis. Whether they deliver toxins or directly
inject effector proteins into the cytoplasm of host cells, bacterial protein secretion systems play a central role in modulating
eukaryotic cell functions. Legionella pneumophila are Gram-negative bacteria that are found ubiquitously in soil andfreshwater environments. Once inhaled by humans, Legionella infections can result in a severe form of pneumonia known as
Legionnaires' disease. Legionella use a type IV secretion system (T4SS) to deliver effector proteins, which mediates theestablishment of a replicative niche in host cells.
The goal of our research is to understand, at the molecular level, how Legionella
subvert host cellular functions to accomplish successful intracellular replication.
To this end, the following research projects are currently in progress.
(1) Analysis of the structure and function of the type IV secretion apparatus.
Essentially nothing is known about the substrate transfer across eukaryotic andFig. 1. T4SS core component DotD represents abacterial membranes that occurs via type IV secretion systems. Furthermore, theconserved structural motif of the periplasmic ring.
macromolecular structure of the type IV secretion apparatus is largely unknown.
To address these questions, we are analyzing the structure and function of thetype IV secretion apparatus from Legionella.
(2) Analysis of effector proteins that translocate from Legionella to host cells.
We previously demonstrated that RalF is translocated by the type IV secretion
system of Legionella into host cells and is required for the recruitment of hostARF proteins to Legionella-containing vacuoles. We also recently demonstrated
that the effector LubX acts as an E3 ligase and targets another effector forproteasomal degradation within host cells. LubX is the first effector protein that
Fig. 2 Discovery of metaeffector.
has been shown to target and regulate another effector within host cells.

Recent Publications
1. Tomoko Kubori and Hiroki Nagai. Bacterial effector-involved temporal and spatial regulation by hijack of the hostubiquitin pathway. Front. Microbiol., 2011:2,145.
2. Hiroki Nagai and Tomoko Kubori. Dot/Icm type IVB secretion systems of Legionella and other gram-negative
bacteria. Front. Microbiol.,2011: 2,136.
3. Kubori T, Shinzawa N, Kanuka H, Nagai H. Legionella metaeffector exploits host proteasome to temporally regulate
cognate effector. PLoS Pathog. 2010;6(12),e1001216.
4. Nakano N, Kubori T, Kinoshita M, Imada K, Nagai H. Crystal structure of Legionella DotD: insights into therelationship between type IVB and type II/III secretion systems. PLoS Pathog. 2010;6(10),e1001129.
5. Kubori, T., Hyakutake, A. and Nagai, H. Legionella translocates an E3 ubiquitin ligase that has multiple U-boxes
with distinct functions. Mol. Microbiol. 2008;67(6),1307-1319.
ĵĹOffice of Combined Program on
Microbiology and Immunology

Research & Activities

Research promotion group
Education promotion group

Associate Professor
Associate Professor

Yoshiko Murakami, M.D., Ph.D.
Hodaka Fujii, M.D., Ph.D

Research & Activities

Office activities
Our institute and the Immunology Frontier Research Center are world-premier institutes for microbiology and immunology,
respectively. Taking maximal advantage of the proximity of these centers, we propose research in the combined fields ofmicrobiology and immunology.

Research & Activities

Office of Combined Program on Microbiology and Immunology

Research promotion
To promote combined research in microbiology and immunology, we are carrying out the following plans:
1. The Awaji international forum on infection and immunology is organized and held annually each September.
2. A research progress report is organized and held every month within the institute.
3. Large research presentations and competitions are organized and held annually within the institute.
4. Collaborative projects are organized with Institut Pasteur in France, Chonnam University in Korea, and the Research
Collaboration Center on Emerging and Re-emerging Infections in Thailand. These activities facilitate research on
microbiology and immunology by promoting collaboration, information exchange, and personal exchange between
laboratories and by facilitating a research atmosphere.
Education promotion
To facilitate seamless research on microbiology and immunology, we direct a multidisciplinary graduate program on
microbiology and immunology, for which we design the curriculum and contents. We also organize an Open House of theinstitute and provide guidance to new students. We organize a combined program on clinical microbiology and immunology inclose cooperation with the Thailand-Japan Research Collaboration Center on Emerging and Re-emerging Infections.

Research group

Associate Professor

Yoshiko Murakami, MD. PhD.

Research projects
Dr. Murakami has an additional appointment in the Department of Immunoregulation as the leader of the paroxysmal nocturnal
hemoglobinuria (PNH) group (see details on the Department page). The research projects for this group include:
1. Investigation of the pathogenesis of PNH;
2. Investigation of the pathogeneses of acquired and inherited glycosylphosphatidylinositol (GPI) deficiencies; and3. Investigation of the functional significance of GPI-anchored proteins using the Pgap3 KO mouse in which GPI anchoredproteins fail to localize to the raft due to defective GPI anchor remodeling.

Recent Publications
1. Murakami Y, Kanzawa N, Saito K, Krawitz PM, Mundlos S, Robinson PN, Karadimitris A, Maeda Y, Kinoshita T.
Mechanism for release of alkaline phosphatase caused by glycosylphosphatidylinositol deficiency in patients withhyperphosphatasia mental retardation syndrome. J Biol Chem. 2012 Feb 24;287(9):6318-25.
2. Murakami Y, Inoue N, Shichishima T, Ohta R, Noji H, Maeda Y, Nishimura J, Kanakura Y, Kinoshita T.
Deregulated expression of HMGA2 is implicated in clonal expansion of PIGA deficient cells in paroxysmal
nocturnal haemoglobinuria. Br J Haematol. 2012 Feb;156(3):383-7.
3. Murakami H, Wang Y, Hasuwa H, Maeda Y, Kinoshita T, Murakami Y. Enhanced response of T lymphocytes from
Pgap3 knockout mouse: Insight into roles of fatty acid remodeling of GPI anchored proteins. Biochem Biophys Res
Commun. 2012 Jan 27;417(4):1235-41.
4. Krawitz PM, Schweiger MR, Rödelsperger C, Marcelis C, Kölsch U, Meisel C, Stephani F, Kinoshita T, Murakami
Y, Bauer S, Isau M, Fischer A, Dahl A, Kerick M, Hecht J, Köhler S, Jäger M, Grünhagen J, de Condor BJ, Doelken
S, Brunner HG, Meinecke P, Passarge E, Thompson MD, Cole DE, Horn D, Roscioli T, Mundlos S, Robinson PN.
Identity-by-descent filtering of exome sequence data identifies PIGV mutations in hyperphosphatasia mental
retardation syndrome. Nat Genet. 2010 Oct;42(10):827-9.
5. Almeida AM*, Murakami Y*, Baker A, Maeda Y, Roberts IA, Kinoshita T, Layton DM, Karadimitris A. Targeted
therapy for inherited GPI deficiency. N Engl J Med. 2007 Apr 19;356(16):1641-7(* equally contributed).
ĵĺOffice of Combined Program on
Microbiology and Immunology

Research & Activities

Fujii Group
Research Group

Associate Professor
Assistant Professor

Hodaka Fujii, M.D., Ph.D.
Toshitsugu Fujita, Ph.D.

Locus-specific biochemical epigenetics
Comprehensive understanding of mechanisms of epigenetic regulation requires identification of molecules bound to genomic
regions of interest in vivo. However, non-biased methods to identify molecules bound to specific genomic loci in vivo arelimited. To perform biochemical and molecular biological analyses of specific genomic regions, we developed the insertional
chromatin immunoprecipitation (iChIP) technology to purify the genomic regions of interest.
The scheme of iChIP is as follows:(i) A repeat of the recognition sequence of an exogenous
DNA-binding protein such as LexA is inserted into the genomic
region of interest in the cell to be analyzed.
(ii) The DNA-binding domain (DNA DB) of the exogenous
DNA-binding protein is fused with a tag(s) and a nuclear
localization signal(s) to be expressed in the cell to be analyzed.
(iii) The resultant cell is stimulated, if necessary, and crosslinked
with formaldehyde or other crosslinkers.
(iv) The cell is lysed, and crosslinked DNA is fragmented bysonication.
(v) The complexes including the exogenous DNA DB are
immunoprecipitated with an antibody against the tag.
(vi) The isolated complexes retain molecules interacting with the
genomic region of interest. Reverse crosslinking and purificationof DNA, RNA, proteins, or other molecules allows theiridentification and characterization.
We applied iChIP to direct identification of components ofinsulator complexes, which function as boundaries of thechromatin domain. We showed that it is feasible to directly
identify proteins and RNA bound to a specific genomic region invivo by using iChIP. We will apply iChIP to elucidation ofmolecular mechanisms of important epigenetic phenomena
including:(a) Lineage commitment of lymphocytes,(b) Expression regulation of odorant receptors,(c) Insulator function,(d) Detection and repair of DNA double-strand breaks, and(e) Epigenetic suppression of expression of tumor suppressor genes.

Figure. Scheme of insertional chromatin immunoprecipitation (iChIP)

Recent Publications
1. Fujita T, Fujii H. Direct identification of insulator components by insertional chromatin immunoprecipitation. PLoS
One. 2011;6(10):e26109.
2. Hoshino A, Fujii H. Insertional chromatin immunoprecipitation: a method for isolating specific genomic regions. J
Biosci Bioeng. 2009 Nov;108(5):446-9.
3. Saint Fleur S, Hoshino A, Kondo K, Egawa T, Fujii H. Regulation of Fas-mediated immune homeostasis by anactivation-induced protein, Cyclon. Blood. 2009 Aug 13;114(7):1355-65.
4. Singh AP, Buscaglia CA, Wang Q, Levay A, Nussenzweig DR, Walker J, Winzeler EA, Fujii H, Fonoura BMA,
Nussenzweig, V. Plasmodium circumsporozoite protein promotes the development of the liver stages of the parasite.
Cell. 2007 Nov 2;131(3):492-504.
5. Hoshino A, Hirst JA, Fujii H. Regulation of cell proliferation by interleukin-3-induced nuclear translocation of
pyruvate kinase. J Biol Chem. 2007 Jun 15;282(24):17706-11.
ĶıResearch Collaboration Center in Overseas
Research & Activities

Thailand-Japan Research Collaboration Center on
Emerging and Re-emerging Infections
Director

SA Professor Naokazu Takeda, Ph.D.

Research Collaboration Center in Overseas

Until recently, it was believed that infectious diseases could be conquered through the development of chemotherapies andvaccines; however, the recent worldwide emergence of new infectious diseases and re-emergence of infectious diseases thatwere thought to be controlled has seriously challenged this notion. Under these circumstances, intensive research that closely
monitors and rapidly analyzes emerging and re-emerging infections is urgently required. Because many infectious diseases canspread rapidly across national borders, these diseases clearly cannot be controlled by the independent efforts of individual
nations.

Campus of Ministry of Public HealthTo this end, Osaka University founded the Research Collaboration Center on Emerging and Re-emerging Infections(RCC-ERI) within the Thai National Institute of Health (NIH), Department of Medical Sciences of the Ministry of Public
Health of Thailand in 2005. The second phase of the program (2010-2014), which is named the Japan Initiative for Global
Research Network on Infectious Diseases (J-GRID), is on-going.
The facility consists of P2 and P3 biohazard containment laboratories and various other equipment and facilities in 600 m2
floor space. Previously, most research projects conducted abroad were short-term, with the researchers only staying for, at
most, a few months to complete their experiments. Due to the installation of the RCC-ERI, researchers are now able to stay for
longer periods of time. The RCC-ERI aims to carry out research projects on both emerging and re-emerging infections in close
collaboration with researchers at the NIH, while simultaneously developing the talents of young scientists from Japan andSoutheast Asian countries in the field of infection.
To conduct basic and applied research and to develop human resources, the RCC-ERI consists of two sections that are devoted
to bacterial and viral infection research. We aim to establish an effective system that would (i) provide information to preventthe emergence of emerging and re-emerging infections, and (ii) promptly activate various countermeasures, should such a
disease emerge, including developing therapeutics or vaccines. Finally, we wish to begin collaborations with laboratories fromnations neighboring Thailand so that we can be at the frontline with the capacity to respond quickly to any globally spreading
infectious disease.

BSL-2 Laboratory

BSL-3 LaboratoryĶĲResearch Collaboration Center in Overseas

Section of Bacterial Infections
Research GroupSA Professor
SA Associate Professor
SA Assistant Professor
SA Researcher
SA Researcher

Shigeyuki Hamada, D.D.S. Ph.D.
Yumi Kumagai, Ph.D.
Kazuhisa Okada, Ph.D.
Chetsada Boonthimat, M.Sc.
Wirongrong Natakuathung, M.Sc.

In collaboration with the National Institute of Health, Department of Medical Sciences of the Ministry of Public Health of Thailand, theSection of Bacterial Infections pays special attention to emerging and re-emerging bacterial diseases that are prevalent or break out in
Asian countries. We conduct molecular epidemiology studies on enteric and systemic bacterial infections, and develop detection andidentification techniques for the diagnosis of the bacterial diseases described below.
Pneumonia, tuberculosis, and acute diarrheal
diseases are associated with high morbidity andmortality rates in Thailand. Consequently, during
the 2010–2014 fiscal years, the Program for the
Promotion of Research Networks for Emerging andRe-emerging Infectious Diseases prioritizedresearch on enteric infectious diseases in Thailandthat are caused by Vibirio cholerae.
Streptopcoccus suis, which is occasionallypathogenic and frequently isolated from diseased
pigs, has been found to cause several systemic(zoonotic) infectious diseases in humans, namely
meningitis, infective endocarditis, and toxic
shock-like syndrome. These diseases have mainly
been observed in Asian countries, including
Northern Thailand. We will examine this emerging
zoonotic infection closely to elucidate the molecularpathogenesis of S. suis infections.

Figure 1. Geographical spread and temporal changes of Vibrio cholerae O1 during the
2007–2010 cholera outbreaks in Thailand. Yellow squares indicate the major MLVA type(s)in each outbreak site shown in the map.

Recent Publications
1. Okada K, Roobthaisong A, Nakagawa I, Hamada S, Chantaroj S. Genotypic and PFGE/MLVA analyses of Vibrio
cholerae O1: geographical spread and temporal changes of isolates during the 2007-2010 cholera outbreaks in
Thailand. PLoS One. 2012 7(1):e30863.
2. Takeuchi D, Kerdsin A, Pienpringam A, Loetthong P, Samerchea S, Luangsuk P, Khamisara K, Wongwan N,
Areeratana P, Chiranairadul P, Lertchayanti S, Petcharat S, Yowang A, Chaiwongsaen P, Nakayama T, Akeda Y,
Hamada S, Sawanpanyalert P, Dejsirilert S, Oishi K. Population-Based Study of Streptococcus suis Infection in
Humans in Phayao Province in Northern Thailand. PLoS One. 2012 7(2):e31265.
3. Okada K, Chantaroj S, Roobthaisong A, Hamada S, Sawanpanyalert P. A Cholera Outbreak of the Vibrio cholerae
O1 El Tor Variant Carrying Classical CtxB in Northeastern Thailand in 2007. Am J Trop Med Hyg. 2010 82(5):875-878.
4. Puiprom O, Chantaroj S, Gangnonngiw W, Okada K, Honda T, Taniguchi T, Sawanpanyalert P. Identification ofcolonization factors of enterotoxigenic Escherichia coli with PCR-based technique. Epidemiol Infect. 2010 138(4):519-524.
5. Okada K, Chantaroj S, Taniguchi T, Suzuki Y, Roobthaisong A, Puiprom O, Honda T, Sawanpanyalert P. A rapid,
simple, and sensitive loop-mediated isothermal amplification method to detect toxigenic Vibrio cholerae in rectal
swab samples. Diagn Microbiol Infect Dis. 2010 66(2):135-139.
ĶĳResearch Collaboration Center in Overseas

Research GroupSA Professor
Postdoctoral Fellow
Postdoctoral Fellow
Postdoctoral Fellow
Postdoctoral Fellow
Research Fellow
Research Fellow
Research Fellow

Naokazu Takeda, Ph.D.
Natsuko Kishishita, Ph.D.
Uamporn Siripanyaphinyo, Ph.D.
Sompong Sapsutthipas, Ph.D.
Nitchakarn Noranate, Ph.D.
Piraporn Utachee, M.Sc.
Chris Verathamjamras, M.Sc.
Uranan Tumkosit, M.Sc.

Research Collaboration Center in Overseas

Section of Viral Infections

The Section of Viral Infections focuses on two emerging and re-emerging viral diseases that are prevalent in Asian countries,
including Thailand, in collaboration with the National Institute of Health, Department of Medical Sciences of the Ministry of
Public Health of Thailand. One subset of these diseases includes mosquito-borne infectious diseases and chikungunya fever,which we study from their epidemiological, molecular biological, and immunological aspects. The other subset of diseases
includes blood-borne infectious diseases (HIV diseases/AIDS). We perform virological and immunological characterizationsof the HIV-1 CRF01_AE strains prevalent in Southeast Asian countries, including Thailand. In addition, studies to elucidate
the mechanism of anti-retroviral drug resistance are in progress.

Recent Publications
1. Boonchawalit S, Jullaksorn D, Uttiyoung J, Yowang A, Krathong N, Chautrakul S, Yamashita A, Ikuta K,
Roobsoong A, Kanitvittaya S, Sawanpanyalert P, Kameoka M. Molecular evolution of HIV-1 CRF01_AE Env inThai patients. PLoS One.2011 6: e27098.
2. Utachee P, Nakamura S, Isarangkura-Na-Ayuthaya P, Tokunaga K, Sawanpanyalert P, Ikuta K, Auwanit W,
Kameoka M. Two N-linked glycosylation sites in the V2 and C2 regions of human immunodeficiency virus type 1
CRF01_AE envelope glycoprotein gp120 regulate viral neutralization susceptibility to the human monoclonal
antibody specific for the CD4 binding domain. J Virol. 2010 84: 4311-20.
3. Soonthornsata B, Tian YS, Utachee P, Sapsutthipas S, Isarangkura-na-Ayuthaya P, Auwanit W, Takagi T, Ikuta K,
Sawanpanyalert P, Kawashita N, Kameoka M. Design and evaluation of antiretroviral peptides corresponding to the
C-terminal heptad repeat region (C-HR) of human immunodeficiency virus type 1 envelope glycoprotein gp41.
Virology 2010 405: 157-64.
4. Li YG, Chittaganpitch M, Waicharoen S, Kanai Y, Bai GR, Kameoka M, Takeda N, Ikuta K, Sawanpanyalert P.
Characterization of H5N1 influenza viruses isolated from humans in vitro. Virol J. 2010 7: 112.
5. Kanai Y, Boonsathorn N, Chittaganpitch M, Bai G, Li Y, Kase T, Takahashi K, Okuno Y, Jampangern W, Ikuta K,
Sawanpanyalert P. The impact of antigenic drift of influenza A virus on human herd immunity: Sero-epidemiologicalstudy of H1N1 in healthy Thai population in 2009. Vaccine 2010 28: 5437-44.
ĶĴResearch Collaboration Center in Overseas

Mahidol-Osaka Center for Infectious Diseases (MOCID)
Research Group

Director Professor (SUP)
Associate Professor
Postdoctoral Fellow
Postdoctoral Fellow
Research Fellow
Research Fellow

Yoshiharu Matsuura, DVM, Ph.D.
Tamaki Okabayashi, DVM, Ph.D.
Mikiko Sasayama, Ph.D.
Promsin Masrinoul, Ph.D.
Orapim Puiprom, M.Sc.
Panjaporn Chaichana, M.Sc.

The interest of MOCID focuses on several tropical infectious diseases that are of human health importance in Thailand,particularly mosquito-borne viral infectious diseases such as dengue fever/dengue hemorrhagic fever and chikungunya fever.
We focus on the development of rapid diagnosis kits for viral diseases. We also study the prevalence of viral infections inhuman and mosquitoes and the pathogeneses of the viruses. We would like to raise young scientists’ interests and researchskills on infectious diseases through clinical collaborations with Mahidol University.

Our main research projects include:
1) Epidemiology of mosquito-borne viral infections in humans and mosquitoes in Thailand;
2) Generation and characterization of human and mouse monoclonal antibodies against dengue virus and chikungunya virus; and
3) Development of diagnosis kits for viral diseases.

Recent Publications
1. Co-existence of major and minor viral populations from two different origins in patients secondarily infected withdengue virus serotype 2 in Bangkok. Puiprom O, Yamashita A, Sasayama M, Limkittikul K, Boonha K,
Jittmitraphap A, Leaungwutiwong P, Kurosu T, Ramasoota P, Ikuta K. Biochem Biopys Res Commun. 2011, 143:
136-142.
2. Fab MAbs specific to HA of influenza virus with H5N1 neutralizing activity selected from immunized chicken
phage library. Pitaksajjakul P, Lekcharoensuk P, Uparagarin N, Barbas C, Ibrahim MS, Ikuta K, and Ramasoota P.
Biochem Biopys Res Commun. 2010, 395: 496-501.
ĶĵResearch Collaboration Center in Overseas

Research Group

Endowed Chair Professor Ph.D.
Endowed Assistant Professor Ph.D.
Specialized Research Affiliate Ph.D.

Eiji Konishi
Atsushi Yamanaka
Miwa Kuwahara

BIKEN Endowed Department of Dengue Vaccine Development was established in Faculty of Tropical Medicine, Mahidol
University, Thailand, in 2011 by endowment from The Research Foundation for Microbial Diseases of Osaka University,
Osaka, Japan to Research Institute for Microbial Diseases, Osaka University, Osaka, Japan.

Research Collaboration Center in Overseas

BIKEN Endowed Department of Dengue Vaccine Development

Dengue fever is the most important mosquito-borne viral disease, which is distributed in tropical regions and producing an
estimated 300,000 patients daily. Dengue hemorrhagic fever is its severer form and has a mortality up to 20% if anappropriate treatment is not done. Unfortunately, no approved vaccines or specific antivirals have been developed.
Our department will carry out basic research studies on (1) mechanisms involved in pathogenesis of dengue fever anddengue hemorrhagic fever, (2) virulence, transmission and evolution of dengue viruses, and (3) dengue vaccine
development using several strategies.

Recent publications

Recent Publications
1. Eiji Konishi, Yuko Miyagawa: Balance of infection-enhancing and neutralizing antibodies induced by a dengue
tetravalent DNA vaccine in a mouse model. Microbes and Infection. 13(12-13):1091-8, 2011
2. Eiji Konishi, Mayu Konishi: Nonstructural Protein 1 Antibody-Based Epitope-Blocking Enzyme-Linked
Immunosorbent Assay to Differentiate Japanese Encephalitis Virus from Dengue Virus Infections in Humans. Jpn J
Infect Dis. 64(4):284-91, 2011
3. Atsushi Yamanaka, Kris C. Mulyatno, Helen Susilowati, Eryk Hendrianto, Amor P. Ginting, Dian D. Sary, Fedik A.
Rantam, Soegeng Soegijanto, Eiji Konishi: Displacement of the Predominant Dengue Virus from Type 2 to Type 1
with a Subsequent Genotype Shift from IV to I in Surabaya, Indonesia 2008-2010. PLoS One. 2011;6(11):e27322.
Epub 2011 Nov 7.
4. Kris C. Mulyatno, Atsushi Yamanaka, Ngadino, Eiji Konishi: Resistence of Aedes aegypti laevae to temephos
insecticide in Surabaya, Indonesia. Southeast Asian J Trop Med Public Health. 43:29-33. (2012).
5. Kris C. Mulyatno, Helen Susilowati, Atsushi Yamanaka, Soegeng Soegijanto, Eiji Konishi: First isolation andphylogeny of Chikungunya virus from Surabaya, Indonesia. Jpn J Infect Dis. 65, 92-94 (2012).
ĶĶOsaka University Global COE Program
Frontier Biomedical Science Underlying Organelle Network Biology
Research program
Our main goal is to create an interdisciplinary research center, which will coordinate work in cell biology,
microbiology/immunology, and glycobiology in order to converge on a greater knowledge of the organellar network.
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By combining these fundamental studies with clinical research, we will drive the creation and development of the new field oforganelle network medicine. To this end, we will conduct research focused on achieving an integrated understanding of humandisease, and on developing technological solutions to clinical problems.
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level of both the molecular and organelle networks.
Ultimately, our greater understanding of the organelle network, and of clinically important issues in organelle biology, willallow us to develop novel therapeutic strategies that accelerate the medicine of the 21st century.

Education program
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leadership roles in 21st century science.
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professor whose primary responsibility is the cultivation of our collaborative networks with other institutions.
Ķķ

Name
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Yoshihiro Yoneda*UDGXDWH6FKRRORI)URQWLHU%LRVFLHQFHV 'HSDUWPHQWRI)URQWLHU%LRVFLHQFHV 3URIHVVRU&HOO%LRORJ\0'3K'

Tatsushi Toda
*UDGXDWH6FKRRORI0HGLFLQH 'LYLVLRQRI0HGLFLQH 9LVLWLQJ3URIHVVRU0RO*HQHWLFV 1HXURORJ\0'3K'

Yoshihide Tsujimoto*UDGXDWH6FKRRORI0HGLFLQH 'LYLVLRQRI0HGLFLQH 3URIHVVRU0HGLFDO*HQHWLFV3K'

Kiyoshi Takeda*UDGXDWH6FKRRORI0HGLFLQH 'LYLVLRQRI0HGLFLQH 3URIHVVRU,PPXQRORJ\0'3K'

Kazunori Tomono*UDGXDWH6FKRRORI0HGLFLQH 'LYLVLRQRI0HGLFLQH 3URIHVVRU&OLQLFDO0LFURELRORJ\0'3K'

Taiichi Katayama
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Toshikazu Nakamura
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Tetsuo Takehara*UDGXDWH6FKRRORI0HGLFLQH 'LYLVLRQRI0HGLFLQH 3URIHVVRU*DVWURHQWHURORJ\ +HSDWRORJ\0'3K'

Eiji Miyoshi*UDGXDWH6FKRRORI0HGLFLQH 'LYLVLRQRI+HDOWK6FLHQFHV 3URIHVVRU&OLQLFDODQG/DERUDWRU\0HGLFLQH0'3K'

Yoshinao Wada

Division of roles

Global COE Program

Members&RRUGLQDWLRQRIDFHQWHUHVWDEOLVKPHQWDQG
elucidation of the organelle network
Sugar chain modifications andneurodegeneration diseases
Mechanism of cell death and organelle
Analysis of activity regulation mechanism of thenatural immunity system
Analysis of antibacterial activity anddevelopment of new antimicrobial agents
Neurological function abnormalities andorganella
Treatment strategies against neurodegenerative and
UHQDOGLVHDVHVZLWK+*)+HSDWLWLVRQVHWPHFKDQLVPDQGWUHDWPHQWstrategies
Development of biomarkers using sugar chain
technologies
Development of sugar chain analysis methods*UDGXDWH6FKRRORI0HGLFLQH 'LYLVLRQRI0HGLFLQH 9LVLWLQJ3URIHVVRU0DVV6SHFWURPHWU\0'3K'

Yukari Fujimoto*UDGXDWH6FKRRORI6FLHQFH 'HSDUWPHQWRI&KHPLVWU\ $VVRFLDWH3URIHVVRU2UJDQLF&KHPLVWU\3K'

Naoyuki Taniguchi

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glycoconjugate
Functional analyses of sugar chains/proteins

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Tamotsu Yoshimori*UDGXDWH6FKRRORI)URQWLHU%LRVFLHQFHV 'HSDUWPHQWRI)URQWLHU%LRVFLHQFHV 3URIHVVRU&HOO%LRORJ\3K'

Hitoshi Kikutani

Analyses of roles of membrane traffic in
infection/immunity
Study on the dynamics of the acquired immunity

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Eisuke Mekada
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Yoshiharu Matsuura
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Tatsuo Shioda$QDO\VHVRIIDFWRUVUHODWHGWRWR[LFLW\PDQLIHVWDWLRQRIWKHGLSKWKHULDWR[LQ,QIHFWLRQPHFKDQLVPRI+HSDWLWLV&YLUXVDQG
studies on the control methods
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Yasuhiko Horiguchi
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Kazuyoshi Ikuta
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Toshihiro Horii
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Shizuo Akira:3,,PPXQRORJ\)URQWLHU5HVHDUFK&HQWHU3URIHVVRU,PPXQRORJ\0'3K'

Taroh Kinoshita:3,,PPXQRORJ\)URQWLHU5HVHDUFK&HQWHU3URIHVVRU,PPXQRORJ\3K'

Hisashi Arase:3,,PPXQRORJ\)URQWLHU5HVHDUFK&HQWHU3URIHVVRU,PPXQRORJ\0'3K'

Atsushi Kumanogoh*UDGXDWH6FKRRORI0HGLFLQH 'LYLVLRQRI0HGLFLQH 3URIHVVRU,PPXQRORJ\0'3K'

Tadashi Suzuki
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Yoshiki Yamaguchi
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Analyses of functions and structures of bacterial
virulence factors
Studying emerging viral infections and theirpathogenesis
Development of malaria vaccines and analyses
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KRVWSDWKRJHQLQWHUDFWLRQVStudies on mechanism of immunoregulation bypathogensStudies of immunoregulation/regulatory
molecules of cell migration
Quality controls of free sugar chains andglycoproteins
6WUXFWXUDODQDO\VHVRIJO\FRFRQMXJDWHVE\105ĶĸResearch & Activities

Germ Cell Group
Research Group

Associate Professor

Masami Nozaki, Ph. D.

Research Projects(1) Epigenetics in germ cells
Many testicular germ cell-specific genes are retroposons, most ofwhich contain a CpG-rich region within their ORFs. We discovered
that, in somatic cells, methylation of CpG dinucleotides within the
ORF represses its promoter, and that demethylation is necessary forgene expression in spermatogenic cells. We are currently
examining the molecular basis of epigenetic modifications,
including DNA methylation and histone methylation, which occur
within a distinct genomic region in germ cells.

Figure legend. Neural crest cells derived from ES cells
differentiated into osteoblasts.

(2) Unique structure of sperm chromatin
The haploid genome in the mammalian sperm nucleus is packaged into a highly compact structure containing protamines andsome remaining histones. We are currently analyzing the physiological importance of the somatic-like, histone-containing
regions of sperm chromatin.
(3) Establishment of neural crest stem cell lines and their application to regenerative medicine
Recent Publications
Inoue H, Ohnishi Y, Nakajima M, Kakudo M, Nozaki M. A novel function of EpCAM in oral squamous cell carcinoma
cells under anchorage-independent conditions. Int J Oncol. 2011 Dec; 39 (6): 1401-5.

ĶĹSpecial Research Facilities
Common Research Facilities

Research Facilities

Research & Activities

Animal Resource Center for Infectious Diseases
Research Group

Masaru OKABE，Ph.D.
Masahito IKAWA，Ph.D.
Ayako ISOTANI，Ph.D.
Hidetoshi HASUWA，Ph.D.
Naokazu INOUE，Ph.D.
Yuhkoh SATOUH，Ph.D.

To study microbial diseases, it is important to analyze the interactions between the host and pathogenic organisms. Animalmodels are indispensable in current microbiological and immunological research. Through the use of molecular biology andbiotechnology, we can generate gene-manipulated mice that can aid our understanding of the mechanisms of infection. For
these purposes, experimentally infected animals should be managed in a suitable, safe, and controlled manner.
The Animal Resource Center for Infectious Diseases is a unique facility that was established in 1967 to meet theserequirements. The center is separated into two major areas: one for animal experiments for microbial disease models(Biosafety Level 2 and 3: BSL2 and BSL3), and another is for SPF (Specific Pathogen Free) area. The section for animal
experimentation for the disease models is completely air-conditioned and maintained at a negative air pressure to minimize therisk of contamination. Each subarea has an individual pass through-type autoclave to sterilize all materials before theirremoval. The exhaust air is filtered to avoid exterior dissemination of pathogenic microbes. These measures ensure that diseasemodel animals can be handled safely without accidental cross-contamination.
Before gaining access to this restricted facility, researchers must take an official orientation and submit a research plan forcommittee review. The condition of the animals is regularly inspected.

Research Facilities

Head, Professor
Associate Professor
Assistant Professor
Assistant Professor (SUP)
Assistant Professor (SUP)SA Assistant Professor (SUP)

Services available at our facility include the generation of gene-manipulated animals, in vitro fertilization, andcryopreservation of mouse strains (Table 1).

Table 1. No. of mouse lines produced/preserved

Period1995-2000
2001-2003
2004-2006
2007-20092010,2011
total
Figure 1: Biosafety level 3 room (Building A).
The room for disease model animals andexperimentation at biosafety level 3. Hemorrhageic
fever with renal syndrome-causing virus (HFRSV)
was isolated in this area. Animal experiments for
Creutzfeldt-Jakob Disease (CJD), severe acuterespiratory syndrome (SARS), and Acquired
Immune Deficiency Syndrome (AIDS) also can behandled in this facility.

Tg mice
228
104
43
22
31
428

KO mice
50
57
69
74
79
329

Cryopreservation
261
443
331
216
220
1471

Tg, transgenic; KO, knock-outĶĺSpecial Research Facilities
Common Research Facilities

Research Facilities

DNA-chip Development Center for Infectious Diseases
Research Group

Head, Professor
Assistant Professor
Associate Professor (SUP)

Hiroshi Nojima, Ph.D.
Daisuke Okuzaki, Ph.D.
Norikazu Yabuta, Ph.D.

Facility Management: The establishment of infectious diseases is driven by the gene expression of pathogenic organisms
within the infected host cells. To understand parasite pathogenesis and pathophysiology and to develop new methods toprevent and treat infectious diseases, it is necessary to identify the pathogenic genes that are expressed in the infected host cells
and to determine how they induce disease at the genetic level. This process requires an analysis of the transcriptional patternsof both the genes of the pathogenic organism and the responsive genes of the host genome.
The DNA-chip Development Center for Infectious Diseases is a unique facility that was established in 2004 to analyze thetranscriptional dynamics and variations involved in infectious diseases. Two research approaches are employed in this facility:(1) Transcriptome analysis using DNA-chip analyzers
The high-density DNA microarray system in this facility permits
comprehensive transcriptional analysis of gene expression in the human ormouse host, and in various pathogenic organisms. Two DNA microarraysystems (Agilent- and Affymetrix-type) are available in this center. Our
real-time PCR analysis system (ABI, PRISM7900HT-2) and Nano-counterare useful for more accurate quantitative analysis of the transcriptional levelsof particular genes. In addition, a novel DNA microarray system (Genopal of
Mitsubishi Rayon Co. Ltd.) is currently being used in this center. An example
of its practical use is the establishment of a blood RNA-based system for the
diagnosis of autoimmune diseases such as vasculitis. This system employs a“focused microarray” that examines the expression of ~200 bloodcell-specific and disease-related genes.
(2) Proteome analysis using mass spectrometry
Comprehensive translational analyses are also very important in furthering
our understanding of infectious diseases. The MS/MS spectrometer installedin this facility allows the analysis of the expression, interactions, andmodifications of proteins from humans, mice, and pathogenic organisms. Thiscenter is also capable of recent technical innovations, such as the mass
spectrometric detection of pathogenic organisms to facilitate the developmentof novel diagnostic systems for infectious diseases.

Fig. 1: High-density DNA microarray system.

Fig. 2: MS/MS spectrometer.

Recent Publications
1. Okuzaki D, Kimura S, Yabuta N, Oomine T, Nojima H. LeukoCatch, a quick and efficient tool for the preparation ofleukocyte extracts from blood. BMC Clin Pathol. 2011 Aug 17;11(1):9.
2. Okuzaki D, Fukushima T, Tougan T, Ishii T., Kobayashi S, Yoshizaki K, Akita T, and Nojima H. GenopalTM: anovel hollow fiber array for focused microarray analysis. DNA Res., 2010 Dec;17(6):369-79.
3. Tougan T, Okuzaki D, Nojima H. Chum-RNA allows preparation of a high-quality cDNA library from a single-cell
quantity of mRNA without PCR amplification. Nucleic Acids Res. 2008 Sep;36(15):e92.
4. Kobayashi S, Ito A, Okuzaki D, Onda H, Yabuta N, Nagamori I, Suzuki K, Hashimoto H, Nojima H. Expression
profiling of PBMC-based diagnostic gene markers isolated from vasculitis patients. DNA Res. 2008 Aug;15(4):253-65.
5. Tougan T, Onda H, Okuzaki D, Kobayashi S, Hashimoto H, Nojima H. Focused microarray analysis of peripheralmononuclear blood cells from Churg-Strauss syndrome patients. DNA Res. 2008 Apr 30;15(2):103-14.
ķıSpecial Research Facilities
Common Research Facilities

Research Facilities

Research Group

Research Facilities

Center for genetic analysis of biological responses<Production laboratory for genetically manipulated animals>Head, Professor
Masaru Okabe, Ph.D.
SA Associate Professor
Kazuo Yamagata, Ph.D.
Assistant Professor (SUP)
Hidetoshi Hasuwa, Ph.D.
SA Assistant Professor
Jun Ueda, Ph.D.

<Laboratory for analysis of genetically manipulated animals>
Professor (SUP)
Shizuo Akira, M.D., Ph.D.
Professor (SUP)
Taroh Kinoshita, Ph.D.
Professor (SUP)
Hisashi Arase, M.D., Ph.D.
Professor (SUP)
Hitoshi Kikutani, M.D., Ph.D.
Professor (SUP)
Masato Okada, Ph.D.
<Resource laboratory for genetically manipulated animals>
Professor (SUP)
Nobuyuki Takakura, M.D., Ph.D.
Visiting Professor
Kenichi Yamamura, M.D., Ph.D. Professor (SUP)
Hiroshi Nojima, Ph.D.
Associate Professor (SUP)
Masahito Ikawa, Ph.D.
Assistant Professor (SUP) Naokazu Inoue, Ph.D.
Assistant Professor (SUP)
Ayako Isotani, Ph.D.
<Laboratory for promotion of collaborative research>
Visiting Professor
Yoichiro Iwakura, Ph.D.
Visiting Professor
Nobuaki Yoshida, M.D., Ph.D.
Assistant Professor (SUP)
Naohisa Goto, Ph.D.
SA Assistant Professor (SUP) Yuhkoh Satouh, Ph.D.
Our bodies are kept homeostatically stable through functions of proteins produced by many genes. In other words, our health isbasically maintained in accordance with the balance of our gene products. Many diseases can be traced to a defect ormalfunction in specific genes. To identify and develop new drugs or therapies, it is very important to identify the function ofeach gene in vivo. However, at present, we do not have enough information about the function of genes to clarify theirrelationships to each other, or to analyze the relationships to specific diseases systematically.
Gene-disrupted animals can be a powerful tool in helping us understand the role of certain genes in vivo. Such animals can beproduced and used as a model for various human diseases and to screen new drugs. With this in mind, the preparation ofgene-disrupted mouse lines of entire genes is planned and is progressing in the form of research projects on a national level in many
countries. Gene-manipulated animals can also be valuable resources that are patent-protected and offer potential for thedevelopment of new drugs and therapeutic methods. It is extremely important for Japan to make a strong contribution in this area.
Under these circumstances, we created a consortium of three research institutes from three universities (The Research Institutefor Microbial Diseases, Osaka University; The Institute ofMedical Science, University of Tokyo; and The Center for
Animal Resources and Development, Kumamoto
University), with headquarters at Kumamoto University.
Within this collaborative consortium, we aim to produce
many gene-manipulated animals that are closely focused onhuman diseases. At our center, we are mainly focused on
genes related to reproduction, infection, and allergy, taking
advantage of an existing disease-screening system in our
university, including features such as fluorescent sperm andeggs (Figure 1). Through these gene-manipulated animals,
we will perform translational research for the establishmentof new therapeutic systems and pharmaceuticals.
Figure 1. Strategy for elucidating sterility

Recent Publications
1. Tokuhiro K, Ikawa M, Benham AM, Okabe M. Testis specific PDILT is required for quality control of sperm
membrane protein ADAM3 and male infertility. Proc Natl Acad Sci U S A. 2012 March 6; 109 (10): 3850-3855.
2. Inoue N, Satouh Y, Ikawa M, Okabe M, Yanagimachi R. Acrosome-reacted mouse spermatozoa recovered from theperivitelline space can fertilize other eggs. Proc Natl Acad Sci U S A. 2011 Dec 13;108(50):20008-11.
3. Kumasawa K, Ikawa M, Kidoya H, Hasuwa H, Saito-Fujita T, Morioka Y, Takakura N, Kimura T, Okabe M.
Pravastatin induces placental growth factor (PGF) and ameliorates preeclampsia in a mouse model. Proc Natl Acad
Sci U S A. 2011 Jan 25;108(4):1451-5.
4. Isotani A, Hatayama H, Kaseda K, Ikawa M, Okabe M. Formation of a thymus from rat ES cells in xenogeneic nude
PRXVHļUDW(6FKLPHUDVGenes Cells. 2011 Apr;16(4): 397-405.
5. Ikawa M, Tokuhiro K, Yamaguchi R, Benham AM, Tamura T, Wada I, Satouh Y, Inoue N, Okabe M. Calsperin is atestis-specific chaperone required for sperm fertility. J Biol Chem. 2011 Feb 18;286(7):5639-46.
ķĲResearch Facilities

Special Research Facilities
Common Research Facilities

The Biken History Museum
Head, Professor

Professor

Hiroshi Nojima, Ph.D.

The Research Institute for Microbial Diseases (RIMD) was established in 1934 by combining two institutions, namely the
Takeo Tuberculosis Institute and the Osaka Leprosy Institute. To commemorate the 70th anniversary of its establishment, aplan of the Biken History Museum was proposed, and it was opened on December 17, 2010.
The Research Institute for Microbial Diseases (RIMD) originated from three institutions, namely Osaka Medical School, theTakeo Tuberculosis Institute (donated by Mr. Jiemon Takeo), and the Osaka Leprosy Institute (donated by an anonymous
benefactor). In 1929, Dr. Chozaburo Kusumoto (the president of Osaka Medical School) and Dr. Tenji Taniguchi (professor of
Bacterial Serology, Osaka Medical School) were concerned about thefrequent introduction of non-native infectious diseases like cholera andthe plague into the Kansai district via the Kobe port, a majorinternational port of Japan at that time; given its central location of the
Kansai district, it readily served as a gateway from which these diseases
could spread throughout the rest of Japan. The concerns of Dr.
Chozaburo Kusumoto and Dr. Tenji Taniguchi were heightened by theirexperiences with the Great Kanto Earthquake in 1923, which had
revealed to them how rapidly infectious diseases could spread.
These concerns led them to strongly encourage the Governor of the
Osaka Prefecture, Mr. Zenzaburo Shibata, and the Osaka business
community to establish public institutions on infectious diseases in theFig.1. A picture of Biken History Museum before theopening ceremony.
Kansai district, particularly in Osaka. In response to this campaign, thebusinessman Mr. Gendo Yamaguchi donated 200,000 yen (theequivalent now of several hundred million yen), which made it possible
to construct the main building of RIMD at the Nakanoshima campus of
Osaka Medical School (Dojima, Osaka City) in February 1934. After its
construction, the researchers of the three institutions mentioned above
moved into the RIMD building and continued their active research.
Both members and non-members of Osaka University can visit theMuseum free of charge from 9:00 a.m. to 5 p.m. on working days. A
gorgeous pamphlet will be presented by writing the visitors name at the
information desk of RIMD which is located near the Museum entrance.
Inside the museum, you will find the portrait bust of Mr. Jiemon
Takeo, 10th (left) and 11th (right), Koch’ s microscope donated by theGerman government, influenza and SARS virus model made by Kaiyodo
Fig. 2. The portrait bust of Mr. Jiemon Takeo, 10th (left)
Co. Ltd. (Kadoma, Osaka).
and 11th (right).

Fig. 3. Koch’ s microscope.

ķĳFig. 4. A plastic model of In fluenza virus
made by Kaiyodo Co. Ltd.

Fig. 5. A plastic model of Severe Acute Respiratory
Syndrome (SARS) virus made by Kaiyodo Co. Ltd.

Research Facilities

Special Research Facilities
Common Research Facilities

Head

Professor
Assistant Professor

Masato Okada, Ph.D.
Shinji Higashiyama, Ph.D.

Research Facilities

Central Instrumentation Laboratory

Since our laboratory was established in the late 1950s, it has grown topossess various high-performance instruments, including a laser
microdissection system and mass spectrum analyzer, as well as
ultracentrifuges, electron microscopes, cell sorters, automatic plasmid
purification systems, and DNA sequencing machines. Our laboratory is
installed with large liquid nitrogen tanks for the preservation of living
materials, such as cells and viruses, and contains a room for specified
chemical substances. Several technicians are employed to keep the
instruments in proper working condition and to provide advice tobeginners and ongoing support for researchers. In addition, thetechnicians execute cell sorting, nucleotide sequencing, electron
microscope observations, and mass spectrometric analyses on samples
upon request from Institute researchers. We anticipate that technical services will become increasingly more important in the
future because many new instruments are precise, complicated, and require extensive training to use. Plans to accommodate
such changes are currently in progress.

Radioisotope Laboratory
Head

Professor
Assistant Professor

Masato Okada, Ph.D.
Shinji Higashiyama, Ph.D.

The Radioisotope (RI) Laboratory was built adjacent to the main building of theInstitute in 1967. It was extended by branch laboratories, for a combined space of about
600 m2, during the establishment of the North building in 1979 and the Central
Laboratory for Biological Hazardous Microbes in 1983. In 1998, a radiation exposure
room was established on the first basement level of the South building. The Genome
Information Research Center Radioisotope Laboratory joined the RI Laboratory in
2007. The Main RI Laboratory, North Building RI Laboratory, and Genome
Information Research Center Radioisotope Laboratory were closed in 2010 and 2011,
and the new RI Laboratory was established in the Immunology Frontier ResearchCenter building in 2011.
The RI Laboratory is designed for biomedical experiments utilizing radioisotopes and it
plays an important role in the Institute. Its facilities include an RI stockroom,
distribution room, tissue culture room, and area for RI measuring equipment. Safety
requirements are met by a stringent security system that involves the use of ID cards
and the computerized management of radioisotopes. About 200 researchers use thislaboratory every year.

ķĴResearch Facilities

Special Research Facilities
Common Research Facilities

Central Laboratory for Biological Hazardous Microbes
Head

Professor

Tatsuo Shioda, D. Med. Sc.
This laboratory was set up in 1983 to ensure the safe handling of biologicallyhazardous microbes, such as hemorrhagic fever with renal syndrome (HFRS) virus.
Since then, all experimental studies using such microbes, including the humanimmunodeficiency virus (HIV), have been carried out in this laboratory. The
laboratory is a three-story building that is 550 m2 in area. The first floor is reservedfor experiments using radioisotopes. The facilities are designed 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 a negative pressure.
Air is released from the facility through high-quality outlet filters to minimize
contamination of the outside environment. Each room is equipped with safetycabinets and autoclaves for the sterilization of used materials before their disposal.
The entire laboratory was renovated from 2005 to 2007 to increase number ofpathogens simultaneously used in this laboratory.
In 2010 and 2011, 50 and 52 researchers, respectively, were approved by the
Biosafety Committee to use this laboratory. The microbes used included HIV,
human and avian influenza viruses, SARS corona virus, and scrapie agent.

Library
HeadProfessor

Toshihiro Horii, Ph.D.

The RIMD library collects academic books and journals on microbiology and immunology, as well as work on relatedscientific fields such as cell biology, genetics, histology, developmental biology, biochemistry, pharmacology pathology,
microbiology and oncology. In particular, we have collected rare books on parasitology that cannot be found at other institutes.
These books are frequently accessed by visitors to the RIMD library. Due to the construction of a new building for the
Integrated Life Science Building, the RIMD library was moved to a temporary library room set up on the 1st floor of the southbuilding of RIMD on December 2007. At July 2010, the RIMD library was moved to a new space located at the 1st floor of the
new RIMD main building. Since the temporary library space was quite small, we were forced to discard most of the old books
and journals. For this reason, we now only hold journals published after 1991, all of the back issues of Biken Journal, and13,000 books. We now purchase 34 and 25 journals published in English and Japanese, respectively. Most of the books are
kept in the stock room, but textbooks and newly arrived journals are arranged on the front shelf of the bureau. Most of thematerials in the RIMD library are registered on the online index at the main Library of Osaka University, which can also beaccessed by libraries throughout Japan via the Inter Library Loan (ILL) system. One librarian handles the RIMD library
together with two professors, two associate professors and one assistant professor who act as members of RIMD librarycommittee. These members also take care of the publication of the “Annual Reports of the Research Institute for Microbial
Diseases Osaka University” (online only from 2003).

ķĵRelated Facility
Related Facility

World Premier International Research Center

Immunology Frontier Research Center
ɜUniqueness and Objectives
Immunology has always been a scientific strength of Japan. In
particular, Osaka University has been historically known for its
leading immunology researchers including Prof. Shizuo Akira.
The Osaka University Immunology Frontier Research Center(IFReC), directed by Prof. Akira, is an example of this reputation.
IFReC was selected by the Japanese Ministry of Education, Culture,
Sports, Science and Technology (MEXT) as one of the nation's
exclusive World Premier International (WPI) Research Center
Initiative Programs. Established on October 1st of 2007, the centeris expected to engage in high level research that will make it aninternational leader in the field of immunology. Immunology
investigates the mechanisms that protect the body against microbial
infection. Because the immune system is essential for eliminating
infectious pathogens from the body, its malfunction gives rise tovarious disorders such as autoimmune diseases, allograft rejection
during transplantation, and allergies.
The scientific aim of IFReC is to unveil a comprehensive
understanding of the dynamic immune system by employing avariety of imaging technologies and Bioinformatics to immunology.

International Advisory Review Board
Director

Deputy Director
Management Committee
Board of Representatives
Laboratories
Immunology Groups
Host Defense（Shizuo Akira）

Experimental Immunology（Shimon Sakaguchi）

Immunoglycobiology（Taroh Kinoshita）

Cell Signaling（Takashi Saito）

Immunopathology（Atsushi Kumanogoh）

Lymphocyte Diﬀerentiation（Tomohiro Kurosaki）

Immunochemistry（Hisashi Arase）

Lymphocyte Development（Fritz Melchers）

Immune Regulation（Tadamitsu Kishimoto）

Gastrointestinal Immunology（Myoung Ho Jang）

Immune Regulation（Tsuneyasu KAisho）

Malaria Immunology（Cevayir Coban）

Developmental Immunology

Vaccine Science（Ken Ishii）

Mucosal Immunology（Kiyoshi Takeda）

Immune Network（Rikinari Hanayama）

Molecular Immunology（Hitoshi Kikutani）

Immunoparasitology（Masahiro Yamamoto）

Imaging Groups
Single Molecule Imaging（Toshio Yanagida）

Chemical Imaging Techniques（Kazuya Kikuchi）

Biofunctional Imaging（Yoshichika Yoshioka）

Biophotonics（Nicholas Isaac Smith）

Cellular Dynamics（Masaru Ishii）

Immune Response Dynamics（Kazuhiro Suzuki）

Nuclear Medicine（Jun Hatazawa）
Bioinformatics Groups
Information System（Yutaka Hata）

Systems Immunology（Daron M. Standley）

Bioinfomatics and Genomics（Diego Miranda-Saavedra）
Research Planning and Management Oﬃce
Administrative Director

General Aﬀairs Section

Accounting SectionķĶɜResearch fields / Expected Achievements
To date, research in immunology has either been carried out by isolating immune cells from the body and examining these cells
in vitro or by using in vitro cultured cell lines. Although such studies have provided many new insights on the immunology
system, we still do not understand the system well enough to predict how it will respond when a certain pathogen invades. One
approach to resolving this problem is to study immune responses in a spatiotemporal manner. Given the importance of thespatiotemporal organization of the lymphoid organs in an immune response and the importance of understanding how single
immune cells behave, combining immunology techniques with imaging techniques is vital for advancing immunology to a newfrontier.
To meet our needs and expectations, new imaging techniques will need to be developed by applying an interdisciplinary effort
that combines people from disparate fields like physics, computer science, and immunology.
By integrating the immunology and imaging fields, we will be able to understand the dynamic interactions of immune cells andtheir activation. This will lead to new and more efficient development strategies for vaccines and immune therapies when
combating infectious diseases, cancers and autoimmunity disorders.

ķķRelated FacilityķĸAccounts
Management Expenses Grants（unit : thousand yen）
Classification

2007

2008

2009

2010

2011

Personnel

917,415

905,437

859,673

887,150

863,168

Non-Personnel

495,488

513,073

548,947

704,408

567,143

Total

1,412,903

1,418,510

1,408,620

1,591,558

1,430,311

Other Grants（unit : thousand yen）
Classification

2007

2008

2009

2010

2011

Contract Research

1,175,396

1,022,353

1,040,180

908,861

711,772

Donations for Research

1,215,677

187,969

343,772

689,654765,777

4,591

3,406

2,090

4,506

4,082

2,395,664

1,213,728

1,386,042

1,603,021

1,481,631

Miscellaneous
Total

Grants-in-Aid for Scientific Research（unit : thousand yen）
Classification

2007

2008

2009

2010

2011

MEXT Research
Grants613,870863.592

688,999453,744466,212

Health and Labor
Sciences Research Grants

237,575

163,278

118,789

107,632

87,913

Health and Laborother Research Grants-

18,00013,9880

0

21st Century COE
Program Grants

196,900

0

1,048,3450

1,048,3450

1,048,345
0Global COE
Program Grants-

149,599

120,037
1,149,599

1,149,599
85,441

1,149,599
74,992

1,048,345

1,194,469

941,813646,817629,117Total
ķĹBuilding Area③⑦

Building Area …… 6,061㎡

Accounts, Building Area

Site Area …… 36,036㎡

Gross Floor Area …… 23,120㎡

⑫

⑪

⑥

⑨
⑨

⑧

④

⑤

① Main building（left）and⑩ Integrated Life Science building（right）

⑩

①

Under
reconstruction② South buildingⅡ②⑦ Central Laboratory for Biological Hazardous Microbes
and⑤⑥ Animal Resource Center

Building name

Total floor numbers①Main building

7

1,546

6,087

3（1basement）

409

945

②South building Ⅱ

Building area（㎡）Total floor area（㎡）DNA-chip Developmet Center and Genome Information Research Center are included③North 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 building⑪Animal Resource Center C（belonging to IFReC）⑫New IFReC building（tentative name）

10

1,072

9,258

3（ 1 basement）

738

2,482

9

770

6,585

ķĺ

Map & Access
LocationJapan

To Saito Nishi

Kyoto
Tokyo
Osaka
NagoyaTo MinohKayanoOnoharaImamiya

Route 171

Shimizu

National
Cardiovascular Center
Senri Kinran University

Shiga

Ibaraki-shi Stn.

HankyuSenri Line

Senri-Chuo
Stn.

Nara
To
Umeda

To
Kawaramachi
Stn.

Meishin
Expressway

Handai Byoin Mae Stn.
MonorailNational
Museumof Ethnology
Expo Park

Chugoku Expressway
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Kita-Osaka
Kyuko Line

Mie

Osaka
University
Suita
CampusKita-Senri
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Suita
I.C.

Banpaku Kinen
Koen Stn.
r Lo
op R oute

HankyuKyoto Line

Osaka Monorail
Unobe Stn.

Kinki
Expressway

Minami-IbarakiStn.
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Stn.

Wakayama

To Matsubara
To
Kadoma-shi Stn.

Paciﬁc Ocean

Transportation access
To Saito Nishi
Osaka University Suita Campus

Chugoku Expressway
To Takarazuka

Handai Byoin Mae Stn.
Banpaku Kinen
Koen Stn.

To
To
Kyoto Stn. Kyoto Stn.

Osaka (Itami) Airport

SubwayMidosuji Line

Meishin Expressway

HankyuSenri Line

Kita-Osaka
Kyuko LineTo Shin-Kobe Stn.

Yamada Stn.

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do

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nk

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JR Osaka
Loop Line

Osaka(Umeda)Stn.

Tsuruhashi Stn.

Namba Stn.

Kintetsu Line

To
Nara

JR Lines
Nankai Line

Tennoji Stn.
JR Hanwa Line

JR Shinkansen
Monorail

To Kansai International Airport and Wakayamaĸı

Ibaraki-shi
Stn.

JRTokaidoLine

Chuo Loop
Highway

To Awaji Stn.

To Umeda Stn.

Kita-Senri Stn.
Senri-Chuo Stn.

To Kyoto Stn.

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To Kyoto

YukoukaiGeneralHospital●Train : 12-minute walk from “Kita-Senri” Station on Hankyu Senri Line.
●Monorail : 20-minute walk from “Handai Byoin Mae” Station on Osaka Monorail Saito Line.
●Bus : From Senri-Chuo Station :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”.
●Bus : From Hankyu Ibaraki-shi Station:
12-minute walk from “Handai Honbu Mae” Bus Stop on buses heading to “Handai Honbu Mae” (via JR Ibaraki Station).

Map & Access

Suita Campus
Research Institute forMicrobial Diseases

Campus Map
30

NNorth Gate

27
28

25
13

22

24
12

14

18

26

19

29
East Gate

Icho Kaikan

8

5

21
Senri Gate

15

23

West Gate

3

16
1
6

KitasenriHankyuGymnasium

Main Gate

Suita Campus

StationKitasenri

92

Convention Center

4

120

20
11

17
10

119

Osaka Monorail
Handai-ByoinMae
StationExpo Park
7

Line

Expo Park

129
Expo Park West
Chu

gok

Yamada Station

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Osa
Ya

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Open Laboratories for Advanced Bioscience and Biotechnology

1 Administration Bureau

17

2 Graduate School/School of Human Scienses

18

3 Graduate School/Faculty of Medicine

19

4 Faculty of Medicine (Dept. of Allied Health Sciences)20

5 Osaka University Hospital21

6 OSaka University Dental Hospital

22

7 Graduate School/School of Pharmaceutical Scienses

23

8 Graduate School/ School of Engineering

24

9 Graduate School of Frontier Biosciences

25

10 Graduate School of Information Science and Technology

26

11 United Graduate School of Child Development

27

12 Research Institute for Microbial Diseases

28

13 Institute for Protein Research29

14 Low Temperature Center

30

International Student Center
International Center for Biotechnology
Center for Advanced Science and Innovation
The Center for Advanced Medical Engineering and Informatics
Global Collaboration Center
Sustainability Design Center
Institute of Laser Engineering
Immunology Frontier Research Center
Institute of Scientiﬁc and Industrual ResearchInstitute of Social and Economic Reseach
Joining and Welding Research InstituteResearch Center for Ultra-high Voltage
Cybermedia Center
Research Center for Nuclear Physics

15 Radioisotope Research Center
16 Research Center for Environmental Preservation

Published by : RIMD, Osaka University／Edited By : Hiroshi Nojima, the chair of RIMD Public Relations Committee
RIMD, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871, Japan
Tel: +81-6-6877-5111, Fax: +81-6-6879-8266 URL: http://www.biken. osaka-u. ac. jpĸĲ