search enter rontier mmunology World Premier International Research Center Osaka University I F Re C FY 2016 of IFReC Annual Report
WPIWPI ImmunologyImmunologymmunology FrontierFrrontierontier RReResearchessearchearch CCentereenternter FFY2016Y 20162016 Osaka University Osaka University IFReC), IFReC), IFReC Research Center ( Research Center Research Planning & Management Office, Copyright : Frontier Immunology Published in June, 2017 Edit : Message from the Director
As the Director of the Immunology Frontier Re- In FY2016, IFReC organized the “International search Center (WPI-IFReC) at Osaka University, I Symposium on Advanced Immunology” to cele- am very pleased to present the IFReC annual re- brate IFReC’s fi rst decade and Professor Tadamitsu port for fi scal 2016. Kishimoto’s 77th birthday. In the symposium, Since its inception in 2007, IFReC has established the researchers of IFReC recognized past achieve- Contents itself as a Visible International Research Center of ments, and shared the challenges in diverse re- Immunology with the support of many people search fi elds for the future. Message from the Director ・・・・・・1 Outreach Activities including the WPI Program Director and the Pro- We are committed to continuing contributions Looking back on IFReC’s activities over the years ・・・・・・2 Science Cafe ・・・・・・78 Super Science High School Students Fair / Students Visit・・・・・ 79 gram Offi cer. to scientifi c advances through research and edu- Organization WPI 10 years Memorial Symposium・・・・・・80 IFReC was created as part of the WPI program, a cation and evolvement into a world top immu- Organization Chart・・・・・・6 AAAS 2017 Annual Meeting・・・・・・81 national project led by the Japanese Ministry of nology research center. Committee and Advisory Board for IFReC ・・・・・・8 Education, Culture, Sports, Science and Technol- Administrative Office of IFReC 10 Research Projects ・・・・・・ogy. However, from FY2017, IFReC will mark a Support Programs for Fusion Researches・・・・・・84 new stage in its history with a novel agreement Laboratories Young Scientist Support Program for Research Abroad ・・・・・ 86 for academic-industry partnership. This gover- Immunology Groups ・・・・・・12 Imaging Groups ・・・・・・40 Data nance system is an ambitious program without Informatics Groups ・・・・・・54 Common Facilities・・・・・・88 precedent. Units for Combined Research Fields ・・・・・・58 Kishimoto Foundation Fellowship ・・・・・・89 Although the governing structure will change, IFReC's New PIs Major Awards ・・・・・・62 ・・・・・・90 our most important mission “Constructing a Chugai Lab 64 Composition 92 ・・・・・・world-top immunology research center” remains Finance ・・・・・・93 the same. We will make unceasing eff orts to de- Symposia & Seminars Selected Articles・・・・・・94 velop immunology research to ensure translation International Symposium on Advanced Immunology ・・・・・・66 Publications・・・・・・98 -Commemorating the 10 th anniversary of IFReC to medical science.
& the 77 th anniversary og Kishimoto Tadamitsu's birth- Access Map ・・・・・・104 IFReC Seminars ・・・・・・68 IFReC Colloquia・・・・・・70
Events The 6 th NIF Winter School on Advanced Immunology ・・・・・・74 Japanese Lessons ・・・・・・76
Shizuo Akira, MD/PhD Director WPI Immunology Frontier Research Center
1 Looking back on IFReC’s activities over the years Table 2. Performance Comparison between IFReC and Other Institutions Jun Sakanoue (Research Planning and Management Offi ce, IFReC)
Staf f (approx.) 200 250 1,000 Papers in IFReC’s Research Achievements over the past decade Immunology, Biochemistry & 619 715 1,247 Molecular Biology, Since the establishment of IFReC in 2007, there have been FY2017. However, the conclusion of contracts between Osaka Cell Biology 1270 scientific articles published by IFReC researchers. Over University and several pharmaceutical companies offered a Citation Number 36,325 22,720 40,447 10% of these articles have appeared in high impact journals bright outlook and people began to look toward the future Citation Impact 58.7 31.8 32.4 such as Science, Nature, Cell and their affi liates. again for the next generation of IFReC. For this reason, the The productivity of scientifi c articles by IFReC researchers is number of researchers at IFReC increased again in 2016, and h-index 77 71 94 shown in Figure 1. In 2015, the number of articles dipped which the number of papers recovered. Documents in Top 1% 6.6% 4.7% 5.4% is thought to be due to the end of the WPI grant by MEXT from Documents in Top 10% 29.8% 23.4% 27.3%
Web of Science ™ © 2017 Clarivate Analytics/Citation number and h-index as of March 10, 2017 Number of Papers Number of Citation 200 12000 Since IFReC has a lower number of researchers than LIAI or ably with the other two institutes, allowing us to conclude that WEHI, the total citation number or h-index of IFReC did not IFReC researchers have produced high-quality papers for the 10000 150 match these institutes. However, IFReC’s fi gures for Citation Im- past ten years. 8000 pact, and Documents in Top 1% and Top 10% compared favor- 100 6000
4000 50 2000 Internationalization and Interdisciplinary Researches 0 0 2008 2009 2010 2011 2012 2013 2014 2015 2016 2008 2009 2010 2011 2012 2013 2014 2015 2016 Promotion of internationalization and interdisciplinary re- researchers in the immunology fi eld are around 25% (data not search is an important mission of the WPI program and WPI shown). IFReC had been internationalized to some extent at an Web of Science™ © 2017 Clarivate Analytics as of March 10, 2017 institutions including IFReC. early stage of the WPI program, and generally continued to be The ratio of international co-authored papers is considered fl at thereafter. The ratio of international collaborative papers of Figure 1. Research Output in 2008-2016 to be one of the indexes for internationalization of the insti- LIAI and WEHI were 52% and 48%, respectively (2014-2016). tute. The ratio for IFReC remained around 41 to 43% after 2010 The ratio of papers in interdisciplinary researches areas in (Figure 2), whereas the average values by all Osaka University IFReC was 21% (2014-2016), and is still increasing.
Table 1 shows the extracted parameters for institution evaluation in all the research fi elds of IFReC. 41.7 41.3 Table 1. Number of Published Papers and their Citations in 35.6 International Co-authored (%) All the Research Fields Interdisciplinary Research (%)
Number of Papers 1,270 21.0
Number of Citations 54,840 14.7 13.1
Citation Impact 43.2 Figure 2. International Co-authored & Interdisciplinary Research h-index 96 2008-2010 2011-2013 2014-2016
Web of Science™ © 2017 Clarivate Analytics as of March 10, 2017 IFReC, which has created a large number of quality science research, is surely commensurable with institutes honored with papers, and promoted internationalization and interdisciplinary World Premier Status.
Table 2 shows the performance comparison between IFReC Since each of the three institutes has a diff erent composition of and two world top research institutions in the life science fi eld, research fi elds, to be fair, the parameters for comparison were References La Jolla Institute for Allergy and Immunology (LIAI), and Walter calculated only in the three research areas of Immunology, Bio- ・ Annual Reports of IFReC FY2010-2015 and Eliza Hall Institute of Medical Research (WEHI), Australia. chemistry & Molecular Biology, and Cell Biology. ・ Web of ScienceTM © 2017 CLARIVATE ANALYTICS ・ IncitesTM © 2017 CLARIVATE ANALYTICS
2 3 Organization As of March, 2017 Organization Chart Immunology Group
Host Defense ...... Shizuo Akira Immunoglycobiology ...... Taroh Kinoshita Immunopathology ...... Atsushi Kumanogoh Immunochemistry ...... Hisashi Arase Immune Regulation ...... Tadamitsu Kishimoto MEXT Mucosal Immunology ...... Kiyoshi Takeda Immune Regulation ...... Hitoshi Kikutani WPI Initiative Experimental Immunology ...... Shimon Sakaguchi ProgramProgram Committee JSPS Cell Signaling ...... Takashi Saito Lymphocyte Differentiation...... Tomohiro Kurosaki WPI Working Group Lymphocyte Development ...... Fritz Melchers Malaria Immunology ...... Cevayir Coban Vaccine Science ...... Ken J. Ishii Immunoparasitology ...... Masahiro Yamamoto Biochemistry and Immunology ...... Shigekazu Nagata International Scientific Advisory Board Imaging Group
Single Molecule Imaging ...... Toshio Yanagida Biofunctional Imaging ...... Yoshichika Yoshioka Director Research Groups Immunology and Cell Biology ...... Masaru Ishii Nuclear Medicine ...... Jun Hatazawa Chemical Imaging Techniques ...... Kazuya Kikuchi Biophotonics ...... Nicholas Isaac Smith Deputy Directors Management Committee Immune Response Dynamics ...... Kazuhiro Suzuki Brain-Immune Interaction ...... Ben Seymour
Board of Informatics Group Administrative Representatives Director Information Systems ...... Yutaka Hata Systems Immunology ...... Daron M Standley
General Affairs Section Section Accounting Section Units for Combined Research Fields Administrative Department Quantitative Immunology ...... Yutaro Kumagai Diego Diez Immuno-Genomics ...... Alexis Vandenbon Hiromasa Morikawa
Research Planning and Management Office Core Instrumentation Facility Common Facilities Animal Resource Center for Infectious Diseases Network Administration Office Osaka University
Institute for Frontier Medical Sciences, Kyoto University President Domestic RIKEN Center for Integrative Medical Sciences Cooperative National Institute of Biomedical Innovation, Health and Nutrition Administrative Bureau Institutions Convergent Research Consortium for Immunologic Disease, Overseas Seoul, St Mary's Hospital, Catholic University of Korea Indian Institute of Science Education and Research, India
6 7 Committee and Advisory Board for IFReC International Scientifi c Advisory Board As one of the nine centers selected for the World Premier International Research Center Initiative (WPI), IFReC has been the subject The International Scientifi c Advisory Board conducts evaluations on scientifi c achievements of IFReC PIs by reviewing their reports or of evaluations including site visits and follow-ups by the WPI Program Committee. direct interviews. as of FY2016 In close cooperation with the Program Directors, the Program Offi cer and working group members, the WPI Program Committee Immunology conducts follow-up activities on progress being made by the WPI institutes including IFReC, with an eye to developing them into Jeff rey Ravetch The Rockefeller University, USA “highly visible research centers.” Christopher Goodnow Australian National University Immunology
Richard Locksley University of California, San Francisco, USA Immunology WPI Program Committee Lewis L. Lanier University of California, San Francisco, USA Immunology Program Director as of FY2016 Anne O'Garra The Francis Crick Institute, UK Immunology Toshio Kuroki Senior Advisor, Research Center for Science Systems, Japan Society for the Promotion of Science (JSPS), Japan Kiyoshi Takatsu Toyama Prefectural Institute for Pharmaceutical Research, Japan Immunology
Deputy Program Director Kazuo Sugamura Miyagi Cancer Center Research Institute, Japan Immunology Akira Ukawa Deputy Director, RIKEN Advanced Institute for Computational Science, Japan Yale Goldman University of Pennsylvania, USA Imaging
Program Committee Members Yasuyoshi Watanabe Center for Life Science Technologies, RIKEN, Japan Imaging
Toshiaki Ikoma Special Adviser, Canon Inc., Japan Masamitsu Iino Nihon University School of Medicine, Japan Imaging
Hiroto Ishida President Emeritus, Kanazawa Gakuin University, Japan Akinori Kidera Yokohama City University, Japan Informatics Shinichiro Ohgaki President, Japan Water Research Center, Japan
Maki Kawai Director General, Institute for Molecular Science, National Institutes of Natural Sciences, Japan
Kiyoshi Kurokawa Adjunct Professor, National Graduate Institute for Policy Studies, Japan All researches performed at IFReC are required to be reviewed by external scientists. Therefore an in-depth evaluation of research activities at IFReC was performed in a peer-review style by the International Scientifi c Advisory Board (ISAB), which consists of ten Honorary Professor Emeritus, High Energy Accelerator Research Organization, Japan Makoto Kobayashi Nobel Laureate in Physics (2008) highly qualifi ed scientists. Since this is the fi nal year of WPI program, the entire research activities since joining IFReC of each PI were evaluated. Norihiko Suzuki Chair of the Board/President, Akita International University The evaluation was conducted through both document and interview. For each PI, one board member was assigned for document Ryozo Nagai President, Jichi Medical University, Japan evaluation and two board members were assigned for the interview evaluation. Ten ISAB members gathered at IFReC on February 2nd rd Michiharu Nakamura Counselor to the President, Japan Science and Technology Agency (JST), Japan and 3 , 2017 for the interviews. [ ] Chairperson Director-General, Center for Research and Development Strategy, JST, Japan The document evaluation was based on a submitted research report including future prospectus, publication list, obtained patents, Ryoji Noyori Nobel Laureate in Chemistry (2001) awards, invited lectures and list of external funding. In addition, the PIs attached copies of the fi ve most important publications as an Michinari Hamaguchi President, JST, Japan appendix to their progress report. Hiroshi Matsumoto President, RIKEN, Japan The board members evaluated the following items and provided the score and the comments.
Rita Colwell Distinguished Professor, University of Maryland, USA
Richard Dasher Consulting Professor, Stanford University, USA
Victor Joseph Dzau President, National Academy of Medicine, USA Evaluation Items
Klaus von Klitzing Director, Max Planck Institute for Solid State Research, Germany Nobel laureate in Physics (1985) 1. Level of achievement
Chuan Poh Lim Chairman, Agency for Science, Technology and Research, Singapore 2. Scientifi c/technical merit
Harriet Wallberg Former president, Karolinska Institutet, Sweden 3. Research output and outcomes (research achievements, patents, invited lectures, etc.)
Jean Zinn-Justin Scientifi c adviser, IRFU/CEA, France 4. Promoting interdisciplinary research
5. Future prospects Working Group Leader and Assigned Members as of FY2016 Score 5 : Excellent, 4 : Good, 3 : Average, 2 : Fair, 1 : Poor [Program Offi cer] University Professor, Institute for Advanced Study, Kyushu University, Japan Takehiko Sasazuki
Hiroshi Kiyono Professor, Institute of Medical Science, The University of Tokyo, Japan
Nagahiro Minato Executive Vice-President for Research, Planning, and Hospital Administration, Kyoto University, Japan After summarizing the evaluation results, anonymous comments and scores made by board members were conveyed individually to PIs. The evaluations and advice received are invaluable to each PI and will surely contribute to their research in the future. Also, IFReC Professor and Chairman, Department of Allergy and Reumatology, Graduate school of Medicine, The University Kazuhiko Yamamoto will make use of the comments and suggestions to open up new paths over the next ten years. of Tokyo, Japan
Günter J. Hämmerling Professor, Tumorimmunology Program, German Cancer Research Center DKFZ, Germany
Associate (chief) scientist, Molecular Imaging Program, National Cancer Institute, National Institutes of Health, Hisataka Kobayashi USA
Philippe Kourilsky Professor, Collège de France/Honorary Director-General, The Institute of Pasteur, France
8 9 Administrative Office of IFReC
General Aff airs Section
■ Employment /acceptance of researchers and staff procedures ■ Social insurance (part-time)/ employment insurance ■ Management of work hours ■ Procedures related to patents ■ Issuing various certifi cates ■ Procedures related to international students ■ Support for international researchers
Accounting Section
■ Budget drafting / implementation / management ■ Purchasing procedures ■ Acceptance and implementation of third-party funding ■ Payment of payroll, travel expense and honorarium ■ Health insurance procedures ■ Management of buildings and assets ■ RI (Radio Isotope) procedures Laboratories Research Planning & Management Offi ce
■ Research Promotion & Support (Consultation for grants & patents, Fusion research program, etc.)
■ Establishing Research Environments (Facility & Safety management, Research agreement, etc.)
■ Fostering Young Scientists (Winter School etc.)
■ Organizing Scientifi c Events (Symposia, Colloquia, Seminars, etc.)
■ Public Relations (Publishing, Website, Outreach to citizens, etc.)
■ WPI evaluation issues (Progress report, Advisory Board meeting, etc.)
As of March, 2017
10 12 ent patternsingeneexpression. cesses anddevelopment ofvariousdiseases.Among thesedis- role inhomeostasis ofperipheralorgans,keyimmunological pro- with diversefunctions.Itisthought thatthesecellsplayacrucial of tissue-residentM2-likemacrophages. pression ofmetabolicdisordersby controllingthediff tissues. Trib1iscriticalforadipose tissuemaintenanceandsup- in variousorgans,includingbonemarrow,spleenandadipose defi M2 macrophages(whichwetermM2-likemacrophages).Trib1 of tissue-residentmacrophages—thatsharecharacteristicswith Previously wereportedthatTrib1iscriticalforthediff sponses toanti-infl host defense.Whereas,M2macrophagesareassociatedwithre- M1 macrophagesarepro-infl recognize diff ruses, andplayamajorroleininnateimmunity.IndividualTLRs ing organismsrangingfrombacteriatofungi,protozoaandvi- of microorganisms.Toll-likereceptors(TLRs)arecapablesens- recognition receptors(PRRs),whichrecognizespecifi pathogens. Innateimmuneresponsesareinitiatedbypattern lutionally conservedhostdefensemechanismagainstvarious progenitor involvedinfi Identifi Monocytesandmacrophagescomprise avarietyofsubsets Macrophages consistofatleasttwosubgroups,M1andM2. We arefocusedontheinnateimmunesystem,whichisanevo- ciency resultsin a severereductionofM2-likemacrophages Host Defense cation ofanatypicalmonocyteand committed erent microbialcomponents,andgiverisetodiff ammatory reactions,andtissueremodeling. ammatory andhaveacentralrolein brosis c structures erentiation erentiation er- their committedprogenitor. fi dritic-cell progenitors.OurresultsshowthatSatMarecriticalfor cyte/macrophage progenitors,butnotfrommacrophage/den- identifi FcεRI+ granulocyte/macrophageprogenitors,andanewly mice resultedinfi haematopoietic cells.AdoptivetransferofSatMintoCebpb-/- infl more, thedevelopmentofbleomycin-inducedfi Cebpb-defi hancer bindingproteinbeta(C/EBPβ),andarecriticalforfi share granulocytecharacteristics,areregulatedbyCCAAT/en- termed segregated-nucleus-containingatypicalmonocytes(SatM), that Ceacam1 are the specificsubtypesofmonocytesandmacrophagesthat with activationofmonocytesandmacrophages.However, fective therapies.Thedevelopmentoffibrosisisassociated Its pathogenesisispoorlyunderstood,andtherearefewef- eases, fi nied bysplenomegaly andlymphadenopathy.Regnase-1 isan mice developed spontaneousautoimmunediseases accompa- type zincfi protein Regnase-1inimmune system Regulation ofmRNAstability byCCCH-typezinc-fi brosis andthatC/EBPβlicensesdiff Regnase-1, alsoknownasZc3h12a, isamemberoftheCCCH- ammation, waspreventedin chimericmicewithCebpb-/- involved in fi ed SatMprogenitordownstreamofLy6C-FcεRI+granulo- brosis isalife-threateningdiseaseofunknownaetiology. nger domaincontaining proteins.Regnase-1knockout ciency resultsinacompletelackofSatM.Further- Shizuo Akira, Support Staff Support Visiting Scientist Research Assistant Postdoctoral Fellow Assistant Professor Associate Professor Professor +Msr1+Ly6C-F4/80-Mac1+ brosis havenotyetbeenidentifi brosis brosis. Notably,SatMare derivedfromLy6C-
MD/PhD erentiation ofSatMfrom 7 4 9 2 TanakaHiroki Kenta Maruyama Takashi Satoh Kazuhiko Maeda Shizuo Akira
monocytes, whichwe ed. We showed ed. Weshowed brosis, butnot brosis. nger phages andc-Rel,Ox40,IL-2inCD4 cleavage oftheir3'UTRssuchasIL-6andIL-12p40inmacro- endogenous RNasethatdestabilizesasetofmRNAsthrough nase-1 expressioniscriticalformodulationofTcellactivation. cell eff activity ofMalt1isinvolvedincontrolthemRNAstabilityT tion andenhancesTcellactivation.Consistently,theprotease is cleavedbyMalt1proteaseafterTcellreceptor(TCR)stimula- ■ ■ ■ Recent Publications 114, 2681-2686(2017). antiviralresponse. ribose) polymerase-mediated Kozaki T, etal. damage elicitsa Mitochondrial TCDD-inducible poly(ADP- proteins inimmunecells. (2017)inpress. Immunol. Int. by CCCH-type S.RegulationofmRNAstability K&Akira Maeda zinc-fi genitor involved in fi Satoh T, etal. Identifi ector genes.ThisindicatesthatdynamiccontrolofReg- The morphologyofSatM Figure 1. brosis. Nature 541,96-101(2017). cation of an atypical monocyte andcommitted pro- monocyte cation ofanatypical + Tcells.Regnase-1itself Proc. Natl. Acad. USA Sci. nger fective treatmentforimmune-relatedinfl derstanding oftheinnateimmunesystemandtodevelopanef- mutant miceandtissue-specifi precious roleofRegnase-1inimmunecellsbyusing ■ ■ We arestudyingtoachievethegoalofacomprehensiveun- Based onthesefi (2016). S. K & Akira Maeda CutTLR7 Structure: inZ-Loop. 45,705-707 Immunity Microbiol. Spectr. 4(6),doi:10.1128/microbiolspec. (2016). MCHD-0040 Satoh S. T &Akira Toll-Like Proteins. Receptor Inducible Signaling andIts Concept ofdisorderspecifi Figure 2. ndings, wepromoteunderstandingofthe c macrophages c Regnase-1defi ammatory diseases. cient mice.
13 Immunology 14 stone, D.L.etal, Israel (Edvardson,S.etal., genes incollaborationswithresearchgroupsfromCanadaand herited GPIdefi inherited GPIdefi teins (GPI-APs).In2016wemadeseveraldiscoveries. and medicalsignifi ing toclarifythemechanismofbiogenesis,biologicalfunctions bonded tothecarboxyl-terminus.Ourlaboratoryhasbeenwork- surface viaglycosylphosphatidylinositol(GPI)thatisamide- and complementregulatoryproteins,areanchoredtothecell as receptors,adhesionmolecules,enzymes,enzymeinhibitors GPI anchors. involved inbiosynthesis,transfer toproteinsandmaturationof genic mutationsthatcauseIGDswere identifi glucosamine tophosphatidylinositol (PI)(Figure).Todate,patho- mediates theinitialstepinGPIbiosynthesis, transferofN-acetyl- tal delay.BothPIGCandPIGParecomponentsoftheenzymethat showed gene. pound heterozygousloss-of-functionmutationsinthe intellectual disabilityandseizureshadhomozygousorcom- als fromtwofamilieshavingglobaldevelopmentaldelay,severe Characterization ofnewpathogenicmutationsresponsiblefor More than150diff Two siblingswithbiallelicloss-of-function Immunoglycobiology early-onset seizures,hypotoniaandglobaldevelopmen- ciency causedby mutations in Hum. Mol.Genet ciencies (IGDs):Wereportedthefi cance ofGPIanchors andGPI-anchoredpro- erent proteinswithvariousfunctions,such J. Med.Genet ., 2017).Threeaff ., 54:196-201,2016;John- ed in16of27genes ected individu- PIGP PIGC rst casesofin- mutations mutations and PIGC PIGP
of twoeachfourcomponents. port theideathatp24cargoreceptor isanoctamerconsisting Mol. Biol and p24δ1componentofthecargo receptor(Nagae,M.etal, we reportedcrystalstructuresofGOLDdomainsp24β1,p24γ2 transport ofGPI-anchoredproteinsfromtheER(Figure).In2016, been characterizingp24γ2complex,acargoreceptorforeffi Certain mutationin knockout HEK293cellsalsoshowedsimilarproteinaggregation. normal aggregationintheendoplasmicreticulum(ER). mutants, severalproteinsfailtobesecretedandinsteadformab- (Ihara, S.etal, show PIGNisrequiredforqualitycontrolinmammaliancells with Dr.Ihara’sgroupintheNationalInstituteofGenetics,we mannose instep8GPIbiosynthesis(Figure).Incollaboration that transfersethanolaminephosphateside-chaintothefi of individualsaff canonical functionofPIGNmaycontributetoclinicalsymptoms pendently oftheGPI-anchorbiosynthesis.Adefectinnon- tion topreventdeleteriousproteinaggregationintheERinde- show thatPIGN-1/PIGNhasaconservedandnon-canonicalfunc- in theERwithoutaff In collaborationwithDr.Yamaguchi’sgroupinRIKEN,wehave Demonstration ofnewfunctionPIGN:PIGNisanenzyme ., 428:4087-4099,2016; Taroh Kinoshita, Support Staff Support Visiting Scientist Research Assistant Postdoctoral Fellow Assistant Professor Associate Professor Professor J CellSci ected byIGDcaused ecting GPI-anchorbiosynthesis.Theseresults C. eleganspign-1 ., 130:602-613,2017).In
Proteins causedproteinaggregation 3 3 1 2 Kanzawa Noriyuki Yuko Tashima Yoshiko Murakami Yusuke Maeda Taroh Kinoshita , inpress).Theresultssup- PIGN PhD mutations. C. eleganspign-1 cient PIGN rst J. -
■ ■ ■ Recent Publications transport. J. Mol. Biol.transport. 428, 4087-4099(2016). dynamics domains: implicationfor p24complex formation andcargo and Yamaguchi Y. ofp24βandp24δGolgi andinteraction 3Dstructure Nagae M,Hirata T, K, Morita-Matsumoto Theiler R,Fujita M,Kinoshita T Genet. 98,615-626(2016). withseizures disability andhypotonia.cause intellectual Am.J. Hum. Kinoshita T, SEandMurakami Antonarakis Y. Pathogenic variantsinPIGG N,Parry DA, JohnsonCA,Matsumoto N,HamamyRoberts H, SheridanE, S,Miyatake Nak P,Makrythanasis Santoni F, K, M,SaitsuH,Nakamura M,Zaki Kato by shedding CRIPTO.in embryos J. Cell Biol. 215,705-718(2016). T. AGPIprocessing phospholipaseA2,PGAP6,modulates Nodalsignaling E, Kajikawa Takada Y, M,HamadaH,Maeda Ikawa Saito K, Y andKinoshita Lee GH,Fujita M, Takaoka Murakami K, Y, Fujihara Y, K, N,Murakami Kanzawa ashima M,IssaMY, GuipponiM,Letourneau A,Logan C, of fourcomponents. proteins, α2,β1,γ2andδ1.Wereportevidenceforanoctamericcomplexincludingtwoeach proteins arepackagedintotransportvesiclesbyacargoreceptorconsistingoffourp24 required forprotectionofproteinaggregationinadditiontoGPIbiosynthesis.GPI-anchored in PIGCandPIGP.PIGNisinvolvedstep8GPIbiosynthesis.Wereportedthat We reportedindividualswithinheritedGPIdeficienciescausedbypathogenicmutations N-acetylglucosaminine transferasethatmediatestheinitialstepinGPIbiosyntheticpathway. Biosynthesis andtransportofGPI-anchoredproteins.PIGCPIGParecomponents Figure. ■ ■ proteins. J. Biol. Chem.289,16835-16843(2014). ofglycosylphosphatidylinositol-anchoredthe recognition andtransport alpha helicalregion inp24γ2subunitofp24cargo receptor ispivotal for Theiler R,Fujita M,Nagae Yamaguchi Y, Maeda Y andKinoshita T. The 3084 (2015). dent endosome-to-TGN retrograde Mol. Biol. Cell 26,3071- transport. and Kinoshita T. Post-Golgi anterograde requires GARP-depen- transport Hirata T, D, Fujita Motooka S,Gotoh Murakami K, M,Nakamura Y, Maeda Y
15 Immunology 16 tive oxygenspecies (ROS)productionandresultant neutrophil cells, andthisbinding wasrequiredforthesuppression ofreac- brane-bound SEMA4Dinteracted withplexinB2onendothelial endothelial cells.Moreover,we foundthatneutrophilmem- MA4D. SolubleSEMA4Dexerted pro-infl AM17-mediated neutrophils-specifi MA4D wassignifi In neutrophilsfromAAVpatients,cellsurfaceexpressionofSE- in AAVpatientsandwerecorrelatedwithclinicaldiseasescores. phil cytoplasmicantibody(ANCA)-associatedvasculitis(AAV). for neutrophilsanditspathologicalinvolvementinanti-neutro- functions ofSemaphorin4D(SEMA4D)asanimmunecheckpoint metastasis, andneurodegenerativediseases.Wefocusonthe ders, suchasautoimmunity,allergy,immunedefi study intodiagnosis/therapyforhumanimmunologicaldisor- tions, wearetryingtoapplythefi peutic targetsforarangeofdiseases.Beyondsuchbasicimplica- they arenowconsideredtobepotentialdiagnosticand/orthera- immune responses,bothphysiologicalandpathological, mulative evidenceindicatesthatsemaphorinsalsoparticipatein cules thatfunctionduringneuronaldevelopment.However,cu- Semaphorins wereoriginallyidentifi guidance molecules’suchassemaphorinsandtheirreceptors. tory behaviorinvivobysolubleandmembrane-bound‘immune posed studyistheregulationofimmunecellmotilityandmigra- clinical immunology.Asbasicaspectsofourprojects,pro- We foundthatserumlevelsofsolubleSEMA4Dwereelevated Our researchteamisinvolvedintwoapproaches;basicand Immunopathology cantly down-regulated,aconsequenceofAD- c proteolyticsheddingofSE- ndings fromthisproposed ed asaxon-guidancemole- amtr eff ammatory ciency,cancer/ et on ects marker andpotentialtherapeutictargetforAAV. mediated infl of SEMA4DasobservedinAAVpatientsmayamplifyneutrophil- negative regulatorofneutrophilactivation.Proteolyticcleavage tive burst.Collectively,neutrophilsurfaceSEMA4Dfunctionsasa bition ofthepathogenorANCAIgG-inducedneutrophiloxida- through SEMA4D’sintracellulardomain,andresultedintheinhi- combinant plexinB2ledtothesuppressionofRac1activation extracellular trap(NET)formation.Treatingneutrophilswithre- Atsushi Kumanogoh, Support Staff Support Assistant Professor Professor ammatory responses.SEMA4Disapromisingbio-
5 Shohei Koyama Atsushi Kumanogoh MD/PhD ■ ■ ■ Recent Publications (2015). Pathogenic Rheumatol. 67,1481 andtherapeutic implications. Arthritis by inducinginfl to rheumatoid arthritis H, Saeki Y, Tomita T, Tanaka T, Kumanogoh 4Dcontributes A.Semaphorin M,Hosokawa Nishide K, Morimoto T, Hirano T, Shima Y, M, Narazaki Tsuboi Yoshida Y, NojimaS,Kimura ShiK, T, S,EbinaK, A,Kang Ogata D, Ito (2017). signals. andamino-acid Natureintegrates Commun. 8,14711 cytokine Kumanogoh A.Polarization ofM2macrophages requires Lamtor1 that M,Maeda Nishide Y, Takamatsu D, H,Okuzaki Yamada Mand M,Okada Hosokawa Morimoto K, T, Hayama Y, Mitsui Y, H, SakuraiN,Sarashina-Kida Kimura T, NadaS, Takegahara N,Okuno T, D, Sujin,Ito NojimaS,Kang Dis.Rheum. (2017)inpress. pathogenesis ofneutrophil-mediated autoimmune vasculitis. Ann. andisinvolved 4DinhibitsneutrophilSemaphorin activation inthe Nakanishi Y, Tsuda T, Park JH,Hirano T, Shima Y, M,Kumanogoh A. Narazaki Hosokawa K, Morimoto T, Hayama Y, Kinehara Y, Kato Y, Nakatani T, D, M,Nojima S,Ito Nishide Takamatsu S,Kimura H,Koyama S,Kang T, of neutrophils,andthisdichotomouseff cells. Inaddition,alterationoftheSEMA4D–plexinB2interactionresultsinaberrantactivation SEMA4D isshedbyADAM17.Solublehadpro-infl the generationofROSandNETformation.Incontrast,inAAVpatients,neutrophilsurface neutrophil cellssurfaceSEMA4DreceptorinhibitsRac1activationandnegativelyregulates Under healthyconditions,theinteractionbetweenendothelialplexinB2ligandand Figure. Graphicalschemeofthisstudy. production: cytokine ammatory ect isinvolvedinthepathogenesisofAAV. ■ ■ (2013). and plexinsasreceptors for semaphorins. Nat.Rev. Immunol.13,802-14 oftheneuropilins functions H.Immunological Kumanogoh A,Kikutani causes retinal degeneration.Nature Commun. 4,1406(2013). and 4Aassociates in Semaphorin withdefective endosomalsorting E,Aozasa Morii K, Takagi J, Takahashi M,Kumanogoh A.Apointmutation D, Ito S,Kimura Kang T, Yoshida Y, Maeda K, Morimoto Y, M, A,Ikawa Ogata Nojima S, Toyofuku T, J, Okuno H,Ishigami C,Kaneko Kamao T, Takamatsu H, ammatory functions onendothelial
17 Immunology 18 Because Siglec-4 isspecifi glycoprotein) associates withVZVgBandmediates VZVinfection. ture sociates withgBandisinvolvedin HSV-1infection(Ariietal. and foundthatnon-musclemyosin heavychain(NMHC-IIA)as- ther analyzedhostcellmolecules thatassociatewithHSV-1gB infection (Satohetal.Cell2008;Wang etal.J.Virol.2009).Wefur- tween PILRαandgBisinvolvedinmembranefusionduringHSV-1 protein ofherpessimplexvirus-1(HSV-1),andtheinteractionbe- found thatPILRαassociateswithglycoproteinB(gB),anenvelope munol. 2008;Kurokietal.Proc.Natl.Acad.Sci.USA.2014).We for bothsugarchainsandproteinstructures(Wangetal.J.Im- mun. 2011).PILRαisauniquereceptorthathasbindingsites new ligandforPILR(Kogureetal.Biochem.Biophys.Res.Com- et al.J.Exp.Med.2004).Inaddition,wehaveidentifi both PILRαandPILRβrecognizeCD99asahostligand(Shiratori pressed onvariousimmunecells.Wehavepreviouslyfoundthat for autoantibodies. these proteinsareinvolvedinautoimmunediseasesasatarget misfolded proteinstransportedtothecellsurfacerevealedthat port cellularmisfoldedproteinstothecellsurface.Analysesof MHC classIImoleculesfunctionasmolecularchaperonstotrans- and variouspairedreceptors.Inaddition,wehavefoundthat ruses Interaction betweenimmunereceptorsandherpesvi- PILRα isoneoftheimmuneinhibitoryreceptorsthatareex- We havebeenworkingtheinteractionsbetweenpathogens 2010).WealsofoundthatSiglec-4 (MAG,myelinassociated Immunochemistry cally expressedinneural tissues,Si- ed PANPasa Na- Microbiol. 2016). abnormalities in microbial infection(Figure1,Hirayasu etal.Nat. munity. Therefore,LILRA2isasensor todetectimmunoglobulin dation ofimmunoglobulinsisavery dangeroussituationforim- globulins arequiteimportantmolecules inhostdefense,degra- proteases butnotnormalimmunoglobulins. Becauseimmuno- recognizes abnormalimmunoglobulins cleavedbymicrobial orphan activatingreceptorexpressedonhumanmyeloidcells, lation ofinfl fi sue fi colitis (Kishidaetal.Int.Immunol.2015)aswellincreasedtis- Furthermore, PILRα-defi creased inPILRα-defi fi LPS-induced endotoxinshock.Furtheranalysesrevealedthatin- PILRα-knockout mice.PILRα-defi normalities Identifi mation PILR Biol. Chem.2015). (Suenaga etal.Proc.Natl.Acad.Sci.USA.2010;SuenagaJ. glec-4 seemedtobeinvolvedinneurotropiccharacteristicofVZV ndings indicatedthatPILRα playsanimportantroleintheregu- ltration ofneutrophilsinliverandlungwassignifi We foundthatLILRA2(alsocalledILT1,LIR-7andCD85H),an We analyzedfunctionofPILRαinimmuneresponseusing brilization (Kohyamaetal.Eur.J.Immunol.2016).These α playsanimportantroleinregulationofinfl cation ofnewsensingsystemforimmune ab- Hisashi Arase, ammation byregulatingintegrinfunction. Research Assistant Postdoctoral Fellow Assistant Professor Associate Professor Professor cient mice.(Wangetal.Nat.Immunol. 2012). cient miceshowedsevereDSS-induced cient miceweresusceptibleto MD/PhD 3 4 HirayasuKouyuki Masako Kohyama Tadahiro Suenaga Hisashi AraseHisashi cnl in- cantly am- are targetsforautoantibodiesinautoimmunediseasepatients that misfoldedproteinscomplexedwithMHCclassIImolecules ecules (Jiangetal.Int.Immunol.2013).Furthermore,wefound gens arerescuedfromproteindegradationbyMHCclassIImol- disease susceptibility.Wefoundthatcellularmisfoldedautoanti- unclear howMHCclassIImoleculesareinvolvedinautoimmune tibility tomanyautoimmunediseases.However,ithasremained defense againstpathogens(Hirayasuetal.Nat.Microbiol.2016). of degradedimmunoglobulinsbyLILRA2playsanimportantroleinhost detect immuneabnormalitiescausedbymicrobialpathogens.Therecognition receptor, LILRA2,whichspecifi to evadeimmunesystem.However,systemhasacquiredanovel Microbial pathogensproducevariousproteasesanddegradeimmunoglobulins immune receptorLILRA2 Figure 1.Microbiallycleavedimmunoglobulinsaresensedbytheinnate cules aretargetsforautoimmunediseases Misfolded proteinscomplexedwithMHCclassIImole- ■ ■ ■ Recent Publications 3787-3792 (2014). 3787-3792 (2014). susceptibility.ated withrheumatoid arthritis Proc. Natl. Acad. USA.111, Sci. Lanier LLandAraseH.Autoantibodies to IgG/HLA-DRcomplexes are associ- Takasugi OhmuraK, K, Yoshikawa H,Saito T, Atsumi T, Sasazuki T, I, Katayama ShiK, S,EbinaK, Toyama-SorimachiMatsuoka N, Yasuda S,Horita T, R, Hiwa Kohyama M,Suenaga K, Jin H,AraseN,Hirayasu T, Saito F, Tanimura K, pholipid syndrome. Blood. 125,2835-2844(2015). glycoprotein I/HLAclassIIcomplexes are novel autoantigens inantiphos- I,Sasazuki Katayama T, LanierLL,Atsumi T, Yamada HandAraseβ2- Kohyama M,Ebina Y, Yasuda S,Horita T, Takasugi Ohmura K, K, Yamamoto K, Tanimura JinH,Suenaga K, T, K, Hirayasu K, S,AraseN,Kishida Morikami crobiol. 1,1-7(2016). an innate immunesensorfor microbially cleaved immunoglobulins. Nat.Mi- Chibana H,NagaiNakamura Y, I,Colonna Katayama M,AraseH.LILRA2is Saito F, K, Hirayasu Suenaga T, K, AraseN,Oikawa ShidaK, Yamaoka T, Murota H, MHC classIIallelicpolymorphismsareassociatedwithsuscep- cally recognizesdegradedimmunoglobulinsto volved inthepathogenicityofautoimmunediseases(Figure2). targets forautoantibodiesas‘neoself’antigens,whicharein- normally wouldnotbeexposedtotheimmunesystem,can mune disease.Thissuggestedthatmisfoldedproteins,which molecules wasstronglycorrelatedwithsusceptibilitytoautoim- misfolded proteinstransportedtothecellsurfacebyMHCclassII cers (Araseeta.Br.J.Dermatol.2017).Autoantibodybindingto HLA classIIcomplexinthepatientswithrefractorycutaneousul- 2015). Inaddition,wecoulddetectautoantibodiesagainstβ2GPI/ (Jin etal.Proc.Natl.Acad.Sci.USA.2014;TanimuraBlood. 2017). 2016; Araseetal.Br.J.Dermatol.2017,HiwaArthritisRheumatol. Natl. Acad.Sci.USA.2014;Tanimuraetal.Blood2015,AraseAdv.Immunol. response toself-antigens(Jiangetal.Int.Immunol.2013;JinProc. self’ antigensbyimmunesystem,whichmightinitiateaberrant to thecellsurfacebyMHCclassIImoleculesmayberecognizedas‘neo- transported tooutsidethecells.Therefore,misfoldedproteins Cellular misfoldedproteinsaregenerallydegradedinthecellsandnot II moleculesaretargetsforautoantibodies Figure 2.MisfoldedproteinstransportedtothecellsurfacebyMHCclass ■ ■ tropic herpesviruses. Proc. Natl. Acad. USA107,866-871(2010). Sci. elin-associated glycoprotein ofneuro- mediates membranefusionandentry Suenaga T, Satoh T, P, Somboonthum Kawaguchi Y, Mori Y andAraseH.My- Immunol. 14,34-40(2012). infl Wang J, M,Arase H.Neutrophil Shiratori I,UehoriJ, infi Ikawa ammation isregulated by PILRαviamodula tion of integrin activation. Nat. Nat. tion ofintegrin activation. ltration during
19 Immunology 20 tory Tcellsintothat ofanti-infl which contributed tothealterationofcharacter ofinfl STATs (STAT1,3,5)activationunder Th17polarizingconditions, STAT3 levelinArid5adefi role ofArid5athroughstabilization ofSTAT3mRNA.Reduction shock andEAEmicemodel.Inaddition, wefoundaTcell-intrinsic Arid5a defi Arid5a hasbeenshowntostabilizeIL-6mRNA(Figure1).Infact, tive domain5a(Arid5a),whichbindstothe3'UTRofIL-6mRNA. Previously, weidentifi ing proteins,inwhichdysfunctionledtosustainIL-6production. under TLRsignaling.IL-6mRNAistightlyregulatedbyRNAbind- 6, wefocusedonIL-6geneexpressioninactivatedmacrophages syndrome (CRS). infl sion havealsobeenrevealedintheseraofpatientsfromsystemic chronic immunedisorders.However,highlevelsofIL-6expres- tibody successfullyproveditsoutstandingeffi chronic diseases,tocilizumab,ahumanizedanti-IL-6receptoran- (JIA). Regardingthesignifi such asrheumatoidarthritis(RA)andjuvenileidiopathic duction ofIL-6leadstodevelopmentvariouschronicdiseases body againstinvasionofpathogens.Incontrast,persistentpro- cells andvascularendothelialareactivatedtoprotectthe acute immuneresponses,bywhichlymphocytes,hematopoietic Introduction To pursuethemolecularmechanismsofoverproductionIL- Interleukin-6 (IL-6)isinimportantcytokineearlyphaseof ammatory responsessyndrome(SIRS)andcytokinerelease Immune RegulationImmune cient miceareresistanttodevelopment ofendotoxin- ed anRNA-binding protein,AT-richinterac- cient Tcellsledtotheimbalanceof cance ofIL-6inthepathogenesis ammatory Tcells.Importantly,we cacy against those amma- infl tory mRNAs,whichinturn,contributes tothepathogenesisof suggest thatArid5acontrolsthestabilizationofmanyinfl IL-6 andTNFα,withahighersurvivalrate.Theseresultsstrongly model, whichelicitsTh1responses,showedlowerlevelsofIFNγ, for RNAstabilizationIL-6andSTAT3mRNAs. stabilization ofSTAT3mRNA,suggestingthatArid5aisrequired revealed thatArid5aregulatesnaïveTcells-fatethroughselective cient micein covery ofmortalityinendotoxinshock.Additionally,Arid5a-defi sequently, reductionofIFNγproductioncontributedtothere- signifi ferentiation. Consistentwiththedata,serumlevelofIFNγwas which encodesT-bet,knownasmasterregulatorofTh1celldif- regulates IFNγproductionthroughTbx21mRNAstabilization, shock micemodel.Recently,ourstudieshaveshownthatArid5a defi H&E stainingoflung,liverandspleentissueindicatedthatArid5a lipopolysaccharide (LPS)mediatedshock.Histologicalanalysisby (ARDS) areclinical conditionsinvolvingacuterespiratory dys- cin-induced lunginjury(Published) Arid5a-defi stabilizing T-betmRNA.(Published) Arid5a exacerbatesIFN-γ-mediatedsepticshockby Acute lunginjury (ALI)andacuterespiratorydistress syndrome Arid5a defi ammatory andautoimmunedisease(Figure 1). cient miceareprotectedfromtissueinjuryafterLPSinduced cantly reducedinLPS-inducedArid5adefi Tadamitsu Kishimoto, Support Staff Support Postdoctoral Fellow Assistant Professor Professor cient micearehighlyresistant tobleomy- cient miceareshowntobecompletelyresistant Propionibacterium acnes
2 4 Masuda Kazuya Tadamitsu Kishimoto -primed endotoxinshock cient mice.Con- MD/PhD amma- - the anti-infl mide teratogenicity.Tounderstandthemechanismunderlying modulatory drugs(IMiDs)compoundsresponsibleforthalido- disorders. tial therapeutictargetfortreatmentofacutelunginfl sis. Therefore,thecontrolofArid5aexpressionshouldbeapoten- which indicatestheimportantroleofArid5ainlungtissuefi ciency couldprotectmicefrombleomycin-inducedlungfi munohistological dataoflungtissuesuggeststhatArid5adefi reactive oxygenspecies(ROS)expressioninlung.Moreover,im- d5a defi tant tobleomycin-inducedlunginjury-mediatedmortality.Ari- Interestingly, weobservedthatArid5adefi volved inthepathogenesisofacutelunginfl function withhighmortality.WeexaminedwhetherArid5aisin- Amplifi Figure 1. (Published) tively regulatesTLR-inducedtype1IFNproduction. Rabex-5 isalenalidomidetargetmoleculethatnega- ■ ■ ■ Recent Publications (2016). bilizing T-bet mRNA.Proc. Natl. Acad. USA.113(41),11543-11548 Sci. Zaman MM,etal. Arid5aexacerbates septicshockby IFN-γ-mediated sta- Med. Biol. 941,79-88(2016). Tanaka T, etal. RegulationofIL-6 andDiseases. Adv. inImmunity Exp. induced lunginjury. 29(2),79-85(2017). Immunol. Int. et al. Arid5a-defi Dubey PK, A recentstudyidentifi cation ofIL-6levelbyArid5a. ciencysuppressednotonlyforIL-6production,butalso ammatory propertiesofIMiDswegeneratedCere- ed Cereblonasthereceptorforimmuno- cient mice are highlyresistant to bleomycin- cient miceareresis- ammatory diseases. ammatory brosis bro- - disorders (Figure2). contributing totheeffi plex releasesRabex-5tosuppressTLRinducedSTAT1activation, We haveproposedthatdisruptionoftheCereblon-Rabex-5com- production macrophage celllineupregulatedTLRinducedtype-1interferon of CereblonwithRabex-5.KnockdownRabex-5inahuman ing protein.Signfi ing. Wesubsequentlyidentifi searched forCerebloninteractingproteinsaff might contributetotheanti-infl Cereblon-independent. ToidentifyIMiDtargetmoleculesthat anti-infl separated frompathwaysleadingtoteratogenicity.However,the of thisstudyisthatcertaintherapeuticpropertiesIMiDscanbe production occursindependentlyofCereblon.Thesignifi that theinhibitoryeff duction evenintheabsenceofCereblon.Thus,weconcluded thalidomide waseff blon defi induced type-1interferonproduction. Lenalidomide disruptsaCeleblon-Rabex-5complextosuppressTLR Figure 2. ■ ■ stabilization ofStat3mRNA.J. Exp. Med. 213(4),605-19 (2016). etal. Arid5aregulates K, Masuda naive CD4+ T cell fate through selective 113(38), 10625-30(2016). regulates Proc. 1IFNproduction. TLR-induced type Natl. Acad. USA. Sci. D,Millrine etal. isalenalidomidetarget Rabex-5 moleculethatnegatively ammatory propertiesofIMiDsareunlikelytobeentirely cient miceusingthe CRISPR/Cas9system.Importantly, via aMYD88dependentSTAT1activatingpathway. cantly, IMiDbindingpreventedtheassociation ectiveininhibitingTLR4inducedTNFαpro- ect ofthalidomideonTLRinducedcytokine cacy ofthesecompoundsinautoimmune ed Rabex-5asaCerebloninteract- ammatory propertiesofIMiDswe ected byIMiDbind- cance
21 Immunology 22 ing RINGfi that ishighlyexpressedincolonicepithelialcells.Ageneencod- genes thatarelocatedwithintheseloci,wetriedtofi ceptible lociforIBDincludingulcerativecolitis(UC).Among barrier functionofintestinalepithelialcells. environmental factorsandintestinalimmunity,analyzedthe al cellspossesssomebarrierfunctionsthatseparateintestinal tors andintestinalimmunity.Wethoughtthatepitheli- intestinal epithelialcellsbetweenenvironmentalfac- the developmentofIBD.However,thereexistsasinglelayer and intestinalimmunity(geneticpredisposition)contributeto that bothintestinalenvironmentalfactorsincludingmicrobiota tory boweldiseases(IBD).Severalrecentevidenceshaveindicated meostasis ismaintainedtorevealthepathogenesisofinfl Barrier functionofintestinalepitheliallayers uitination. Thepolyubiquitination inducedbyRNF186 wasK48- was foundtoassociate withtheseproteinsandinduce polyubiq- thelial cellsof that expressionofseveralproteins wasincreasedincolonicepi- using liquidchromatography-tandem massspectrometryshowed tination ofitssubstrates.Comparison ofproteinexpressionlevels RING fi the regulationofintestinalhomeostasis.RNF186,whichharbored lial cellsinthecolon.Therefore,weanalyzedroleof UC-susceptible loci,wasfoundtobehighlyexpressedinepithe- Genome-wide associationstudieshaveidentifi Our laboratoryisanalyzingthemechanismshowintestinalho- nger domain,acted asanE3ligasemediatingpolyubiqui- Mucosal Immunology nger protein nger Rnf186 −/− mice.Theseincludedoccludin. RNF186 186 ( RNF186 ), whichwaslocatedwithin ed severalsus- nd agene RNF186 amma- in human microbiota. SKGmicerearedinaspecifi arthritis development usingarthritis-proneSKGmice harboring Prevotella copri These RApatientsharboredintestinal microbiotadominatedby composition, calleddysbiosis,was foundinsomeRApatients. between healthyvolunteersand RA patients.Alteredmicrobiota tis (RA).Wefi microbiota isinvolvedinthepathogenesisofrheumatoidarthri- volved inarthritispathogenesis.Therefore,weanalyzedwhether models ofarthritisindicatedthattheintestinalmicrobiotaisin- orders otherthanIBD.Severalpreviousstudiesusingmouse UC-associated (SNP) inacodingregionofthe susceptible locihadfoundasinglenucleotidepolymorphism (DSS) treatment.ApreviousdeepsequencingstudyoftheUC- sensitivity tointestinalinfl doplasmic reticulum(ER)stressincolonicepitheliaandincreased ling ERstressincolonicepithelia(Figure1). responsible forthemaintenanceofguthomeostasisbycontrol- mice. Takentogether,thesefi increased sensitivitytoDSS-inducedintestinalinfl mediated, whichleadstoproteindegradation.Thesefi tein homeostasisin epithelial cellsbyinducingpolyubiquitinaton.Thedisturbedpro- dicate thatRNF186regulatesproteinexpressionthecolonic Dysbiosis triggersarthritisdevelopment We alsoanalyzedtheinvolvementofmicrobiotainseveraldis- Kiyoshi Takeda, Support Staff Support Research Assistant Assistant Professor Associate Professor Professor rst comparedthecomposition offecalmicrobiota Rnf186 . Wethenanalyzedhowdysbiosis contributesto Rnf186 SNPledtoimpairedE3ligaseactivityand
ammation afterdextransulfatesodium −/− ndings demonstratethat micecorrelatedwithenhanceden- RNF186 3 6 Mari Murkami OkumuraRyu Hisako Kayama Eiji Umemoto Kiyoshi Takeda gene.Introductionofan MD/PhD c pathogen-free c ammation in ammation ndings in- RNF186 is the microbiotaofhealthycontrolsand and colonized withhumanfecalmicrobiotafromhealthyvolunteers Then, germ-freeSKGmiceweregeneratedandthese tion ofantibioticsprotectedthesemicefromjointinfl however, eliminationofintestinalmicrobiotabyoraladministra- facility developedarthritisuponfungalcomponentinjection; infl induced celldeath,whichleadstothehighsensitivityintestinal targeted proteinwasincreased,causinghighsusceptibilitytostress- colonic epithelialcell.IntheabsenceofRNF186,expressionRNF186- RNF186 isresponsibleforthemaintenanceofproteinhomeostasisin Figure 1. RPL23A-induced Tcellresponsewasattenuatedingerm-freeSKG responses tothearthritis-relatedautoantigen,RPL23A.The lon, butnotspleen,ofRA-SKGmiceshowedenhancedIL-17 ponent injection.Lymphocytesinregionallymphnodesandco- duced. RA-SKGmicedevelopedseverearthritisafterfungalcom- microbiota fromRApatients(RA-SKG)weresuccessfullypro- ■ ■ ■ Recent Publications ammation. intestine. Arthritis Rheum. 68,2646-2661 (2016). Rheum. intestine. Arthritis ofautoreactive development viaactivation utes to arthritis T cells inthe Iida T, Saeki Y, S,& Kumanogoh A,Sakaguchi Takeda contrib- Dysbiosis K. M,FurutaMatsushita Y, S, H,Nakamura N,Kayama M,Sakaguchi Narazaki Maeda Y, Kurakawa T, D, Umemoto E,Motooka Ito Y, K, Hirota Gotoh K, RNF186. Mucosal Immunol. 10,446-459(2017). of intestinal homeostasisby theulcerative colitis-associated gene Tsuchiya K, Watanabe M,Kumanogoh A, Tsukita S,& Takeda Regulation K. H, Okumura R,FurutaKayama Y, Nar Fujimoto M, Kinoshita K, Tamura A, Tanaka D, H,Okuzaki Shimada Y, cruzi Th17 responses by suppressing M& M,Ikawa Murakami Takeda KBATF2 inhibitsimmunopathological D, H,Okuzaki S,Kayama KogaR,KobayashiKitada M,Arima Y, Kumanogoh A, Prevotella infection.J. Exp. Med. (2017)inpress. -dominated RApatients.SKGmiceharboringboth Il23a azaki M, Hatakeyama S, Ikawa M, M,HatakeyamaS,Ikawa azaki expression during Prevotella Trypanosoma -dominated ammation. cally susceptiblemice. ronmental factorthattriggersarthritisdevelopmentingeneti- tion (Figure2).Thesefi by dysbioticmicrobiotaintheintestine,causingjointinfl Thus, SKGTcellsareactivatedtovigorouslyreactautoantigens SKG micemonocolonizedwith dritic cells,respondedtoRPL23Aandrapidlyinducedarthritis. intestine. triggers arthritisdevelopmentviaactivationofautoreactiveTcellsinthe IL-17 responsestoarthritis-relatedautoantigen(RPL23A).Thus,dysbiosis regional lymphnodesandlargeintestineofthesemiceshowedenhanced the regionallymphnodesandlargeintestine.Moreover,lymphocytesin severe infl copri microbiota dominatedby A subpopulationofrheumatoidarthritis(RA)patientsshoweddysbiotic Figure 2. mice. NaïveSKGTcells,co-culturedwith ■ ■ cells. Immunity 42,279-293(2015). cells. Immunity lates ATP-dependent chronic allergic responses by basophils andmast H& Karasuyama Takeda E-NPP3(CD203c) negatively regu- Ectoenzyme K. Takeda A, Yoshikawa S,Kurashima Y, Sato S,Umemoto H, E,Kiyono Tsai M,Kusu SH,Kinoshita T, Shimada K, OkumuraR,Ikeda KayamaH, Y, 121 (2016). regation offl Yamamoto & M,Umemoto E,SanoK, Takeda Lypd8 K. promotes theseg- J, H,Nishimura Ohba H,Matsuno Osawwa Y, S,Iida Nakamura T, Kimura Gotoh K, T, N,Kusu Kamiyama T, Ueda Y, Wu H,BarmanS, H,Iijima Okumura R,Kurakawa T, Nakano T, D, M,Motoka H,Kinoshita Kayama -dominatedmicrobiotafromRApatients(RA-SKGmice)showed ammatoryarthritisaswellincreasednumberofTh17cellsin agellated microbiota andcolonic epithelia.Nature 532,117- ndings indicatethatdysbiosisisanenvi- Prevotella copri P. copri . SKGmiceharboring developedseverearthritis. P. copri -stimulated den- Prevotella amma-
23 Immunology 24 they exhibitednano-molarK ously reportedanti-DNAmAbsobtainedfromSLEmodelmice, clonal antibodies(mAbs)fromacuteSLEsubjects.Unlikeprevi- studies ofanti-DNAantibodies. pathogenicity. Thismaybeduetoalackofprecisestructural widely accepted.Itisstillunclearhowanti-DNAantibodiesexert sis anditscontributiontoclinicalsymptomsinSLEhasbeen insight intoantigenrecognitionbyanti-DNAantibodiesinSLE. teristic stackinginteraction.Ourresultprovidesanewstructural nucleobases arerigidlygrabbedbytheFabthroughcharac- matic hypermutation(SHM)totheantigenrecognition,inwhich The structuralanalysisclearlydemonstratedacontributionofso- Fab ofonemAbclone(designatedas71F12)wassolved(Figure1). tein Research,OsakaUniv),thecrystallographyofligand-bound (SPR). IncollaborationwithProf.JunichiTakagi(InstituteofPro- tibody insystemiclupuserythematosus(SLE) Structural basisforantigenrecognitionofanti-DNAan- In thecurrentstudy,weisolatedhigh-affi Anti-DNA antibodyistheestablishedcriterionforSLEdiagno- Immune RegulationImmune D insurfaceplasmonresonance nity anti-DNA mono- IgE-producing BcellsinNPs. B cells.Wesoughttounderstandthedevelopmentpathwayof high concentrationofIgEpresumablyoriginatedfrominfi sal polyposis.Nasalpolyps(NPs)ofCRSwNPpatientscontaina remains tobeelucidated. place concomitantly.ImmunecellsinvolvedinIgEproduction sal bacteria,inwhichunnecessaryclassswitchingtoIgEtakes CRSwNP isderivedfromprotectiveimmuneresponseagainstna- indicative ofsequentialclassswitchtoIgEinNPs.Takentogether, with nasalpolyposis(CRSwNP) Tracking ofallergy-relatedIgEinchronicrhinosinusitis lineages ofIgEwereoftenfoundinIgGorIgA repertoires wasperformed(Figure2).Itshowedthatmajorclonal such antigen-specifi pyogenes which normallyinhabitinsinonasalcavity,suchas The majorityofisolatedmAbsappearedtobespecifi antibody cloningandexpressionofmonoclonalIgEantibodies. We fi CRSwNP ischaracterizedbyeosinophilicinfl rst determinedantigensofIgEproducedinNPsbyusing . Next,tocomprehendthedevelopmentpathwayof Hitoshi Kikutani, Support Staff Support Postdoctoral Fellow Assistant Professor Professor c IgE,deepsequencingofNP-associatedBCR
1 1 Shuhei Sakakibara Hitoshi Kikutani MD/PhD ammation andna- 1 clonallineages, Streptococcus c tobacteria ltrating Daron M.StandleyinRIMD,OsakaUniv. lineages. Thisanalysiswasperformedby Dr.ShotaNakamura,"Dr.KazuoYamashita"andProf. across immunoglobulinisotypes.Large IgEcloneswerestronglyconnectedwithIgGandIgA Deep sequencinganalysisofexpressed BCRrepertoiresunraveledtheclonedistribution Clonal connectivitybetweenisotypes inNP-associatedimmunoglobulins. Figure 2. antigen recognitionsite. several hydrogenbonds.ItshouldbenotedthatmutatedF33oftheheavychainlocatesat The Fabbindstothyminesthroughastackinginteraction(W101oftheheavychain)and X-ray crystallographyofanti-DNAantibody71F12. Figure 1.
25 Immunology 26 tivation andextended itsbindingsiteswithinthe SEsalongTreg nome organizer Satb1, whichboundtoTreg-SEsbefore theirac- tating earlyinductionoftheassociated genes.Itrequiredthege- Foxp3 Treg-SEs developmentallybegan inTregprogenitorcellsbefore Il2ra thritis, byutilizingananimalmodelestablishedinourlaboratory. nism ofsystemicautoimmunediseases,suchasrheumatoidar- logic self-tolerance;and(iii)thecausepathogeneticmecha- organ transplants,bymanipulatingthemechanismofimmuno- autologous tumorcells,orinducingimmunologictoleranceto cells; (ii)thestrategyforelicitingeff immunologic self-tolerance,inparticulartherolesofregulatoryT sociated withtheTregsignature genes, suchas cell-specifi lineage determinationinvariouscelltypes,weidentifi of super-enhancerswithcelltype-specifi in thethymus.Thisyear,withrecentlydemonstratedassociations are initiatedandcoordinatedtodeterminetheTregcelllineage ever, howTreg-specifi 2013; Morikawaetal.,PNAS2014).Itispoorlyunderstood,how- Treg-specifi ulation, withtheirstableFoxp3expressionchiefl in thethymusasafunctionallydistinctandmatureT-cellsubpop- cal self-toleranceandhomeostasis.Themajorityofthemdevelop Foxp3, areactivelyengagedinthemaintenanceofimmunologi- Treg cells,whichspecifi This laboratorystudies:(i)thecellularandmolecularbasisof (Kitagawaetal.,Nat.Immunol.2017). Theestablishmentof transcriptionandTreg-specifi Experimental Immunology c super-enhancers(Treg-SEs),manyofwhichwereas- cDNAdemethylation(Ohkuraetal.,Immunity2012, c transcriptionalandepigeneticchanges cally expressthetranscriptionfactor ective immuneresponsesto c DNAdemethylation, facili- c generegulationand y maintainedby Foxp3 , Ctla4 ed Treg ed and mune andotherimmunologicaldiseases. cifi of Tregcellsinhumans,andhowgeneticvariationsTreg-spe- tions controlthefunction,development,andfunctionalstability studying howTreg-specifi changes controlTregcelllinagespecifi tablishment andsubsequenttranscriptionalepigenetic Our resultsthusdemonstratehowSatb1-dependentTreg-SEes- development ofsevereautoimmunityandIgEhyperproduction. cell diff ment. other immunologicaldiseasesviaaff how molecularanomalyinthisprocesscausesautoimmuneand sociated genesincluding specifi Treg-SE formationinTregprecursorcells,hinderingbothTreg- cell diff cfunctionalgenescontributetothedevelopmentofautoim- Based onthefi cDNAdemethylationandthetranscriptionofTreg-SE-as- erentiation at theprecursorstageresultedinspontaneous erentiation. Tcell-specifi Shimon Sakaguchi, Support Staff Support Research Assistant Postdoctoral Fellow Assistant Professor Professor ndings describedabovewithmice,wearenow
c transcriptionalandepigeneticaltera- Foxp3 . TheconsequentarrestofTreg c deletion ofSatb1impaired ecting Tregcelldevelop- 6 4 3 Kenji Ichiyama Noriko Sakaguchi Norihisa Mikami Wing JamesBadger Shimon Sakaguchi cation inthethymus,and MD/PhD ■ ■ ■ Recent Publications CD8 Wing JB, Adeegbe D, ofself-reactive S.Detection I,Sakaguchi Katayama Maeda Y, H,Sugiyama D, Nishikawa Ha D, HamaguchiM,Saito T, M, Nishioka trol theprognosis ofcolorectal cancers. Nature Med. 22,679-684(2016). 346, 1536-1540(2014). and Sakaguchi S. and Sakaguchi Two FOXP3 Takiguchi S, Tanoue T, Suda W, M,Doki Mori H,Hattori Morita M,HondaK, Y J,Hamaguchi M,OhkuraN,Sato E,NagaseH,Nishimura Yamamoto H, Saito T, H, Nishikawa Wada H,Nagano Y, Sugiyama D, Maeda Atarashi K, Y, (2017). establishment.Nature 18,173-183, Immunol. dent super-enhancer S.Guidance ofregulatory Sakaguchi T cell development by Satb1-depen- Yasuda D, S,KondoM, Motooka Nakamura K, Taniuchi I,Kohwi-Shigematsu T, Kitagawa Y, OhkuraN,Kidani Y, R,Vandenbon Kawakami K, A,Hirota + T cells withananergic inhealthy phenotype individuals. Science. + CD4 + as wellotherepigeneticmodifi facilitates theexpressionofassociatedTregcellsignaturegenes,including precursor cells.AsTregcelldevelopmentproceeds,enhancer-promoterlooping recruitment ofepigeneticmodifyingenzymesandbecomeTreglineage-committed (SP) stage,Treg-SEregionsundergoSatb1-dependentactivation,likelythroughthe the signaltodirectTregcelldifferentiationviaimmatureCD4single-positive from theCD4/CD8double-positive(DP)stageandboundbySatb1.Uponreceiving regulation duringthymicTregcelldevelopment.Treg-SEregionsarepoisedatleast A modelofsuper-enhancer(SE)establishmentandsubsequenttranscriptional Figure. EstablishmentofTreg-specifi structure. occupy thesiteswhereSatb1initiallybound,maintaininglocalchromatin however, mediator,cohesin,andvarioustranscriptionfactors,includingFoxp3, pre-established molecularfeaturesandSatb1transcriptionisrepressedbyFoxp3; and promoterregions.OnceTregcelldevelopmentiscomplete,Foxp3amplifies histone modificationofpromoterregionsandchromatinlooseningenhancer T-cell con- subpopulationsdistinctly cations suchasT c super-enhancersforTregcelldevelopment ■ ■ sponses via the coreceptor CTLA-4. Immunity.sponses viathecoreceptor 41,1013-1025(2014). CTLA-4. tigen-specifi Wing JB, Ise W, Kurosaki T, S.Regulatory andSakaguchi T-cells control an- ence. 346,363-368(2014). responses to aubiquitous cellular protein inautoimmune disease. Sci- S,Mimori Matsuda T, of S.Detection N,andSakaguchi T-cell Sakaguchi Furu H,Fujii M,Ito T, Nomura T, Yamazaki A, S,Morita Vignali JW, DA, Kappler Ito Y, H, H,Nishikawa OhkuraN,Morikawa Tanaka Hashimoto M,HirotaK, A, reg -specifi c expansionof Tfh cell numberandhumoralimmunere- c DNAdemethylation, Foxp3 ,
27 Immunology 28 weak stimulation induceslong-lastingmicrosynapse whereas in activationofT cells underweakstimulationconditions because quently Tcellactivation.Microsynapse isparticularlyimportant sion, whichhelptheformationof MCsandcSMAC,conse- Microsynapse supportsintegrinoutside-in signalandcelladhe- and LFA1,Paxillin,Pyk2areinthe outsideringofmicrosynapse. TCR-MCs, ZAP70,SLP76andF-actin weredetectedatthecenter, mune synapseinmicro-scale,thustermed lin, attheinitialstageofactivation.Thestructureresemblesim- focal adhesionmolecules,suchasLFA-1,paxillin,Pyk2andvincu- MC issurroundedbyanadhesion-ringcomposedofintegrinand nases, adaptors,andeff by recruitingTCRandproximalsignalingmoleculessuchaski- tion, priortoimmunesynapseformation.TCR-MCsaregenerated microclusters (MC),thesignalingclusterstoinduceTcellactiva- interface. WehavefoundthatactivationisinducedthroughTCR- ciation betweenTcellsandDCgeneratesimmunesynapseatthe with antigen-presentingcellssuchasdendritic(DC).Asso- naling regulation. tiation andhomeostasis,particularlyfromtheperspectiveofsig- determine themolecularmechanismofTcellactivation,diff results ininfectiousandautoimmunediseases.Ourgroupaimsto against infectionandtumorigenesis.AberrancyinTcellfunction generate variouseff responses. Theystartimmuneresponse,induceactivation,and T cellsareactivateduponantigenrecognitionbyassociation T cellsplaycentralrolesininitiatingandregulatingimmune Cell Signaling ector Tcells,whichfunctionforprotection ector molecules.WefoundthateachTCR- microsynapse (Figure). eren- terminus regionofRasGRP1.Massanalysisidentifi RasGRP1. ThemembranelocalizationwasmediatedthroughC- as GEFinTcells,weconcentratedonanalyzingthedynamicsof structure uponstimulation.SinceRasGRP1dominantlyfunctions MC, Ras-GRP1isaccumulatedinauniquemesh-likecytoskeletal were investigated.WhereasSosisinitiallyco-localizedwithTCR- tially regulated,dynamicsoftwoRasGEFs,Ras-GRP1andSos, tion pathways.SinceactivationoftheRas-MAPKpathwayisspa- cell activation,particularlybyanalyzingindividualcriticalactiva- sion. induce activationwiththesupportofmicrosynapseforcelladhe- duce theformationofcSMAC,however,TCR-MCsfunctionto tion signalinTcells.Underweakstimulation,cellsdonotin- apse andMCsseemtobethemajorstructuresinduceactiva- strong stimulationmerelyinducestransiently.Thus,microsyn- cells. Sincewefound thatSTINGishighlyexpressed inTcells,the cells. STINGisnow amajorintracellularDNAsensor ininnate functions andsignalingofTLRs, nucleicacidrecognitioninT and functionbyinnatesignals. We havealreadyanalyzedthe regulation ofRasGRP1throughlinking cytoskeleton. upon TCRstimulation.Thus,Rasactivationismediatedbyspatial structure ofRasGRP1andconsequentlyreducedIL-2production RasGRP1. Plectin-defi known cytoskeletallinker,asabindingmoleculetothisregionof We continuedtoanalyzespatio-andtemporal-regulationofT We havealsoinvestigatedtheregulation ofTcellactivation Takashi Saito, Professor cient Tcellsfailedtogeneratethe mesh PhD Takashi Saito ed Plectin, a and functionistoelucidatethemechanismofinducingautoim- modulators ofactivatedeff secretion. Therefore,STINGligandspossiblyserveasspecifi induces anti-viralandanti-tumorresponsesincludingtypeI-IFN STING stimulationresultedininhibitionofTcellproliferationand function ofSTINGactivationinTcellsisanalyzed.Surprisingly, ■ ■ ■ Recent Publications Ishikawa E,etal.Ishikawa Protein Dregulates ofCD4 positive kinase selection Diff Takeuchi AandSaito T. CD4 Exp. Med. 213,1609-1625(2016). essential formicroclusters structure forms synapse-like J.T cell activation. Hashimoto-Tane A,etal. The adhesion-ringsurrounding Micro each TCR (2016). ofSHP-1. through Nat. Commun. phosphorylation 7,12756mocytes The ultimateaimofourapproachestoanalyzeTcellsignaling erentiation and Function. Front. 8,194(2017). Immunol. + CTL, aCytotoxic CTL, SubsetofCD4 ector Tcells. adhesion moleculeshelpsinducingactivationsignalsthroughTCR-microclusters. (C) Adhesion-ringinmicrosynapseservesinducingoutside-insignalsthrough LFA1/focal particularly underweakstimulation. in earlyactivationandhelpformationofmicroclustercSMACT cellactivation, (B) Microsynapsecomposedofmicro-adhesionringaroundTCR-microcluster isformed microclusters (red)uponTcellstimulation. (A) FocaladhesionmoleculePyk2(green)formsmicro-adhesionring aroundTCR- essential forTcellactivation Figure. Microsynapsecomposedofmicro-adhesionringsurroundingTCR-microclusteris + T cells, Their T + thy- c analyzed inmousemodels. of autoimmunediseases.Thentheonsetautoimmunitywillbe PTPN2) whosemutationsanddefi analyzing functionandregulationbyphosphatase(PTPN22, hibit/prevent autoimmunityandallergicinfl and tobeablemodulatesignalingfunctioninorderin- mune diseasesthroughaberrantTcellsignalingandfunction ■ ■ mice. Arterioscl. Throm. Vas. 36,1141-1151(2016). Matsumoto T, prevents etal. Overexpression atherosclerosis ofCTLA-4 in lineage. J. Exp. Med. 213,123-138(2016). Takeuchi A,etal. CRTAM determines theCD4+cytotoxic T lymphocyte ciency arerelatedtoinduction ammation. Weare
29 Immunology 30 munity. tween autoreactive BandTcells,resultinginfull-blown autoim- B cellsisoneofthekeydriversfor initiatingthepositiveloopbe- titers. Thus,ourresultssuggestthat dysregulationofCD80/86on ies amelioratedaberrantactivation ofBcellsandautoantibody in lupus.Atthepromotingphase, treatmentofanti-CD4antibod- ducing co-stimulatorymolecules CD80/86attheinitiatingphase cells manifestedhyper-responsivetoBcellreceptor,therebyin- using thismousemodel,wefoundthatTet2-andTet3-defi tion ofeff Tet DNAdemethylases,Tet2andTet3,inBcellsledtoaccumula- the DNAdemethylationpathway.Weshowedthatdefi droxymethylcytosine (5hmC)andotherintermediateproductsin lyzing theconversionof5-methylcytosine(5mC)inDNAto5-hy- teins (Tet1,Tet2,andTet3).TheyactasDNAdemethylasebycata- identifi identifi creased riskofdevelopingautoimmunity.Amongthem,Tet3was hundreds ofgenepolymorphismsthatareassociatedwithanin- tolerance Involvement ofDNAdemethylases,Tet2andTet3inBcell susceptibility toseveralautoimmunediseases. protection againsttuberculosisandisassociatedwithincreased For example,thePTPN22 hanced immuneresponseandanincreasedriskofautoimmunity. The immunesystemactsinatrade-off Genome-wideassociationstudies(GWAS)haveidentifi ed Ten-eleventranslocation(Tet)dioxygenasefamilypro- ed tobeassociatedwithSLE;Tet3isoneoftherecently Lymphocyte Diff ector Tcells,autoantibodyformation,andnephritis.By R620W polymorphismprovidesincreased mannerbetweenen- ciency of ciency cient B ed Foxo1 exertsaproliferative role,ratherthanpromoting quies- development led tothelossofGCDZBcells,suggesting that cifi an experimentalsysteminwhich Foxo1isinduciblydeletedspe- B cells.ToaddressFoxo1function inGCbiology,weconducted might playacriticalroleincontrolling geneexpressionwithinGC derived lymphomasfrequentlycarry mutationsinFoxo1,Foxo1 that localizestothenucleusandcontrolsgeneexpression.AsGC- Regulation ofgerminalcenterBcellproliferationbyFoxo1 cells concomitantlywithlosingGCpotential(Fig.1). tion, therebyconferringdiff data suggestthathighCD40signalinducesstrongerT-Binterac- BCRs bysomatichyper-mutation.WefoundthattheIRF4 minal center(GC),whereantigen-specifi non, knownasaffi tigen andeffi progressive increaseintheabilityofserumantibodiestobindan- reactions Selection ofhigh-affi interaction betweenIRF4 upregulated bystrongCD40signals,therebyaff fi was increasedordecreasedbystrongerTcellhelphaplo-insuf- ma cellfateoverGCcycling.Cellsinthisplasma-pronefraction light zone(LZ)GCsubsetwithhighaffi ciency of CD40,respectively.ICAMandSAPonGCBcellswere cally inGCBcells. WefoundthatablationofFoxo1afterGC Foxo1,forkheadboxproteinO1,isatranscriptionalregulator Thehallmarkofantibody-mediatedadaptiveimmunityisthe erentiation Tomohiro Kurosaki, Support Staff Support Research Assistant Assistant Professor Associate Professor Professor ciently provideimmuneprotection.Thisphenome- nity maturation,takesplacemajorlyintheger- nity antibodiesduringgerminalcenter
+ Bcl6 erentiation potentialtowardsplasma lo cellsandT nity BCRsfavoredtheplas- 4 2 Shinya Tanaka Ryo Shinnakasu Takeshi Inoue Wataru Ise Yoshihiro Baba Tomohiro Kurosaki c GCBcellsdiversifytheir FH cells.Together,our ording stronger MD/PhD + Bcl6 lo
induced inLZBcellsaFoxo1-dependentmannerandthatde- GC Bcellproliferation.Weconfi scription factors,c-MycantBATF,bothofwhichareessentialfor adequate Tcellhelp,Foxo1 cence aspreviouslythought.Mechanistically,uponprovisionof ■ ■ ■ Recent Publications loop. Nat.Commun. 7,12596(2016). control immuneresponseTh2-type by regulating theIL-4 amplifi Yasukawa M,Sawasaki T, Kurosaki T, Yamashita M.Bach2-Batfinteractions Matsumoto A, Takemori N, Takemori Nakayama A,ShinodaK, T, OharaO, Kuwahara M,Ise W, M, S,Izumoto Maruyama J, KometaniK, OchiM,Suzuki to T cell help. J. Exp. Med. 214(4),1181-1198(2017). Foxo1tion factor controls germinalcenter Bcell proliferation inresponse Inoue T, R,Ise Shinnakasu W, C,Egawa Kawai T, Kurosaki T. The transcrip- entiation. Curr. 45,126-131(2017)Epubaheadofprint. Opin.Immunol. R,Kurosaki Shinnakasu T. Bandplasmacell diff Regulationofmemory -/- presentation toTFHcellsandexpressionofDZprogramgenes,includingBATF. upon Foxo1ablation.hasalsoimportantrolesintheLZBcells,including antigen high affinityantibodiesagainstforeignantigens.(Right)DZislostandthe GCiscollapsed (Left) WithoutFoxo1,germinalcenterstructureisdisorganizedandmicecould notproduce Figure 2.RegulationofGCBcellproliferationbyFoxo1 diff interaction withTFHistheinductionofIRF4+Bcl6loGCBcellswhichfavorplasmacell regulation ofadhesionmoleculessuchasICAM-1orSLAM.Theconsequencestrong GC-B cellswithhighaffinityBCRreceivestrongTcellhelpviaCD40,whichleadstoup- Figure 1.StrongTcellhelpconfersthediff GCLZBcellscouldnotinducetran- erentiation ratherthanGCre-cycling. rmed thatBATFwastransiently cation er- erentiation potentialtowardplasmacells (Fig.2). ed BATFup-regulationplaysanimportantroleinthisprocess gered afterreceiptofTcellhelp,andsuggestthatFoxo1-mediat- support amodelinwhichtheswitchfromLZtoDZistrig- pletion ofBATFsimilarlyledtoGCdisruption.Thus,ourresults ■ ■ plasma cells. Curr. Opin.Immunol. 33,64-9(2015). Kurosaki Kometani K, T. Diff (7), 861-9(2016). 17 Nat.Immunol. Bcellminal center compartment. cells into thememory Takahashi Y, Fukuyama H,Okada T, Kurosaki T. ofger- Regulated selection R,Inoue Shinnakasu T, S, Adachi Moriyama KometaniK, Y, M, Nakayama erentiation andmaintenance oflong-lived
31 Immunology 32 rebral malaria,an acuteanddeadlyoutcomecaused by IFreC/WPI program’saims. phy (μCT)readilyavailableatIFReC facilitiesinaccordancewith as MRI,multiphotonliveimaging andmicro-computedtomogra- anti-malarials (Figure1). new interventionsfromthisunderstandingasadjuncttherapyto by malariaindiff the understandingoftissue-specifi could bediff action betweeninfectederythrocyteandtissueenvironment rebral malariaand,respiratorydistress,suggestingthattheinter- muscle painandanemiaseverecomplicationsincludingce- tion cancausediseasemanifestationssuchasfever,vomiting, standing intosafeinterventionsuchasvaccinesanddrugs. both inanimalmodelsandhumans,translateourunder- broad andfi understand betterhowtheseparasitesinteractwiththehost.Our tion ofhost-pathogeninteractionsinthecontextmalariato sites. Therefore,researchinourlaboratoryfocusesontheelucida- potent drugsforthecompleteeliminationof millions everyyearduetoalackoffullysuccessfulvaccineor One ofthetopics wehaveinvestigatedisthepathology ofce- We havetakenadvantageofusing imagingtechnologiessuch During blood-stagelifecycleof Malaria continuestobeaglobalburdentheworldthatkills Malaria Immunology erent. Therefore,ourstudieshavemainlyfocusedon nal aimistoinvestigatehost-parasiteinteractions erent tissues/organs.Ourfi c immunopathologycaused Plasmodium nal goalistocreate Plasmodium parasites,infec- Plasmo- para- els mimickingvarious aspectsofhuman compromise bone integrity.Usingwell-established mousemod- ported inmalariaendemicregions, suggestinginfectionmay An increasedincidenceofporous bonelesionshasalsobeenre- among malaria-infectedchildren regardlessofnutritionalstatus. rica isassociatedwithinfectiousdiseases, withahighprevalence suggest thatphysicalgrowthretardation inyoungchildrenAf- enhances theriskoflymphomaformation.Growingevidences modium are asyetpoorlydefi malaria survivorsexperiencelong-term‘hidden’pathologiesthat of patientsrecoverfrommalaria.However,thereisevidencethat 2014 early diagnosismarkerforcerebralmalaria( tory functionalimpairment(lossofsmell)couldbeavaluable storm. Withthesefi cro-bleeding, eventsassociatedwithhighfeverandcytokine show parasiteaccumulationandcellocclusionfollowedbymi- plex trabecularsmallcapillariescomprisingtheolfactorybulb photon imagingoftheolfactorybulbconfi and functionallydamagedby ultra-high fi experimental cerebralmalaria.Deepinvestigationofthebrainby cerebral malariapathogenesisusingultra-highfi sites inmice.Wehaverecentlyreportedanewunderstandingof dium falciparum On theotherhand,despiteseverecomplications,majority ). parasitespromotesprolongedimmuneactivationand Cevayir Coban, eld MRIshowedthattheolfactorybulbisphysically Support Staff Support Research Assistant Postdoctoral Fellow Professor infectionsinhumansand ndings, wehaveprovidedevidencethatolfac- ned. Forinstance,chronicexposureto
Plasmodium 2 2 2 Cevayir Coban MD Plasmodium parasites.Livemulti- P. berghei rmed thatthecom- Cell HostMicrobe, eld MRIduring ANKA para- infection, Plas- fi of bonelossinmalaria-infectedpatientsandthepotentialbene- analog, couldpreventboneloss.Theseresultshighlighttherisk loss. Importantly,supplementationofalfacalcidol,avitaminD3 mozoin, amajorPlasmodiumby-product,didnotcausebone site withimpairedhemoglobindigestionthatproduceslittlehe- genesis favoringboneresorption.Infectionwithamutantpara- creased RANKLexpressionandover-stimulationofosteoclasto- responses inosteoclastandosteoblastprecursors,leadingtoin- bone marrownicheinducesMyD88-dependentinfl stasis, butsustainedaccumulationof mice. Acutemalariainfectionseverelysuppressesbonehomeo- term bonelossinadultmice,andgrowthretardationyoung we recentlyshowedthatinfectioncausessignifi ■ ■ ■ ts ofcouplingbonetherapywithanti-malarialtreatment( Recent Publications Cerebral Cell Malaria. 15(5),551-63(2014). HostMicrobe Plays aKeyRoleinSpatiotemporal PathogenesisZhao Hetal. Olfactory of 61 (2016). safetymozoin evaluation. adjuvant:Apreliminary Vaccine 34(18),2055- Lee MSJ, Igari Y, Tsukui T, Coban C.Current Ishii KJ, statusofsynthetiche- in press. in thebonemarrow andpromote chronic boneloss. Immunol. (2017) Sci. Coban C. S,IshiiKJ, SM,KurodaKhan E,Akira Lee Fujita MSJ, K, Maruyama Y, S,Horii KonishiA,Lelliott PM, Itagaki T, LinJW, caused by approach tothisproblemisuseimagingtechnologiesunderstandimmunopathology investigated topreventandtreatacutechroniccomplicationscaused bymalaria.Our in theeffortstoeliminatemalaria.Thereforeadditionaltreatmentmodalities shouldbe retardation. Highprevalenceofdrugresistanceandlooksuccessfulvaccines aredrawbacks the incompleterecoveryfrominfectioncauseschronicillnessessuchasbone lossorgrowth develop partialimmunityandsufferfrommildsymptoms.Recentevidences suggestthat Blood stage Figure. Plasmodium Plasmodium Plasmodium Plasmodium parasites. productsinthe cant andlong infectioncausesdeadlycomplications.However,mostpeople products persist products ammatory Lee et al., ScienceImmunology,2017,inpress safe adjuvantthatdisplaysverylowautoimmuneproperties. procedures inratsandconcludedthatsynthetichemozoinisa Vaccine, 2016 cology studiesinseveralanimalsandinfectionmodels(Leeetal., and completeditspreliminaryGLPnon-clinicalsafetytoxi- mozoin, asyntheticanalogof effi vants, becauseifrationallydesigned,adjuvantsimprovevaccine opment ofpotentvaccinesagainstmalaria,wefocusontheadju- treat disease(s).Giventhatthereisstillmoretodoforthedevel- standing ofhost-pathogeninteractionsintovaccinesordrugsto ■ ■ cacy. Wehavedevelopedanewadjuvant calledsynthetiche- The fi and Microbe 7(1),50-61(2010). and Microbe modium falciparumInvolves Hemozoin Malarial andHost TLR9. Cell Host Coban of C etal. Immunogenicity Whole Parasite Vaccines Against Plas- 12(5),705-16(2012).Host Microbe to blood-stagemalariainfection by reinforcing hostiron metabolism.Cell Zhao Hetal. Lipocalin2bolsters innate andadaptive immuneresponses nal goalofourresearchisthetranslationunder- ). ThesestudieswereperformedaccordingtoGLP Plasmodium ). -produced hemozoin,
33 Immunology 34 in thelungs(Kuroda Eetal study, weexaminedthedetailed mechanisms ofiBALTformation for localIgEresponsesinducedby particulateinthelungs.Inthis lungs. WesuggestthatiBALTformation playsanimportantrole bronchus-associated lymphoidtissue (iBALT)formationinthe tern). ReleasedIL-1αcontributes to IgEproductionandinducible and releaseIL-1αasaDAMP(damage-associatedmolecularpat- have shownthatparticulatesinducealveolarmacrophagedeath lates triggerallergicresponsesareunclear.InFY2016-2017,we mune responses.However,themechanismsbywhichparticu- ticulates haveastrongadjuvantactivityandinduceType-2im- in allergicinfl Old, butnewlyevolvingadjuvantresearch ox/fl ammation, andmightbeagood MD/PhD ox mice,asamodelofTfh-de- 1 6 4 4 Natsuko Kishishita Takuya Yamamoto Norifumi Iijima Etsushi Kuroda Ken J. Ishii ammation ed cines (manuscriptinpreparation). markers topredictsafetyandimmunogenicityofadjuvantedvac- liminary resultssuggestserummiRNAmayprovideusefulbio- of BiomedicalInnovation,HealthandNutrition(NIBIOHN).Pre- groups includingourlabinIFReCandthoseNationalInstitute relevant immunologicaldisordersarebeingconductedbyfour tained fromvolunteersofvaccineclinicaltrialsandpatients the future. and immunology,contributenewadjuvantdevelopmentin this approachwillbeanimportantplatformforadjuvantbiology standardized corefornewadjuvantcharacterization.Webelieve juvant demonstratedthatcurrentdataworkasareliableand actions (Manuscriptsubmitted).AddinganotherdataofAS04ad- macrophage andIL-1. Lung-specifi Figure 1. ■ ■ ■ Recent Publications target. 7(31),48860-48869(2016). mor microenvironment to release immunogenicdeadtumor cells. Onco- Kitahata Y, et al. Circulating nano-particulate TLR9 agonistscouts outtu- Lymphoid Tissue Formation. Immunity. 45(6),1299-1310 (2016). Releaseto Promoteand Interleukin-1α Bronchus-Associated Inducible Kuroda Fine E,etal. Inhaled Alveolar Particles Induce Macrophage Death (2017) inpress. cles viaDNAvaccination results inrobust CD8+ T cell responses J. Immunol. Kanuma T, vesi- into etal. extracellular CD63-mediated antigendelivery Cohort aswellretrospectiveanalysisofhumansamplesob- c allergicinfl ammation byPM2.5andotherallergenviaalveolar exchanges ofresearchers. novation, HealthandNutrition(NIBIOHN)accompanyingactive these diseasestogetherwithNationalInstituteofBiomedicalIn- continue innovativeresearchanddevelopmentvaccinesagainst er's disease,andmanyotherlifestyle-relateddiseases.Wewill clude abroadrangeofdiseasessuchascancer,allergy,Alzheim- not onlyrestrictedtoaframeworkofinfectiousdiseasesbutin- delivery systemandadjuvants.Vaccinetargetdiseasesarenow contributions. Theseincludesearchingofmodalityantigen ies inmylabatIFReCarealsoexpectedtomakeconsiderable molecular designing.Towardsuchanend,outcomesfromstud- generation vaccinesisvergingonpracticalpossibilitybased comprehensive transcriptomeanalysis. Adjuvant databaseprojecttoevaluateadjuvanteffi Figure 2. Future prospect ■ ■ The ForefrontVaccineDevelopment:Developmentofnext- tics. 28(7),329-38(2016). Immunol. Int. Temizoz B, etal. Vaccine adjuvantsaspotential cancer immunotherapeu- J. Immunol.46(5),1142-51(2016). response induced by humanizedCTL CpGcomplexed Eur. withβ-glucan. role IIFNsand IL-12 etal. oftype Species-dependent Kobiyama K, inthe cacy aswellsafetyby 35 Immunology 36 quired immunityinvolvingaparasite-specifi which ruption ofthemembranesparasitophorousvacuole(PV), the cell-autonomousinnateimmuneresponse,causingdis- cellular protozoanpathogen kine, interferonγ(IFN-γ),againstinfectionoftheobligatoryintra- IRGs andGBPstothe Atg5, Atg7,andAtg16L1,positivelycontroltherecruitmentof m3), andsomeessentialautophagycomponents,suchasAtg3, latory IRGs(calledGMS-IRGs),suchasIrgm1andIrgm3(Irgm1/ branes andarethoughttocooperateinPVdisruption.Theregu- such asIrga6andIrgb6,arerecruitedtothe binding proteins(GBPs).GBPsandeff Pases, suchasimmunity-relatedGTPases(IRGs)andguanylate- been showntoeffi tein suchasovalbumin(OVA)thatissecretedintothePVs,has nous parasite-derivedproteins,includingGRA6,oramodelpro- to sponse, whichisimportantforbothacuteandchronicresistance tween thePVand thehostendoplasmicreticulum (ER)providesa TAP1-dependent manner.Furthermore,thedirect interactionbe- major histocompatibilitycomplex classI(MHCI)moleculesina cytes, processthevacuolarproteins andpresenttheantigenson fessional nonhematopoieticcells, suchasfi such asdendriticcells(DCs)andmacrophages, aswellnonpro- cells. Professionalhematopoietic antigen-presenting cells(APCs), When ahealthyhostisinfectedwith The hostimmunesystemproducesaproinfl T. gondii T. gondii Immunoparasitology andprotectiveimmunityagainstinfection.Endoge- formsinsideinfectedcells,byIFN-γ-inducibleGT- ciently elicitstrongparasite-specifi T. gondii Toxoplasmagondii PVs. ector IRGs(calledGKS-IRGs), T. gondii broblasts andastro- c CD8 c T. gondii . IFN-γstimulates ammatory cyto- ammatory , itdevelopsac- + Tcellre- PVmem- c CD8 c + T cifi tor alpha(TNF-α)furtherenhancestheactivationofantigen-spe- The primingofmacrophagesbytheIFN-γandtumornecrosisfac- route ofentryfortheMHCI-restrictedantigenpresentedonDCs. plays aspecifi ubiquitin tothe elimination ofintracellularpathogens,isrecruitedtogetherwith binding domainandthatisinvolvedintheselectiveautophagic CD8 autonomous innateimmunityandvacuolar-antigen-specifi tion. However,themolecularlinkbetweenIFN-γ-inducedcell- potential roleforIFN-γ-inducibleGTPasesinantigenpresenta- vation afterPVdisruptionbyIFN-γ-inducibleGTPases. c CD8 c Here, weshowthatp62/Sqstm1,aproteinwithubiquitin- + Tcell-dependentacquiredimmunityremainsunclear. + TcellsinanIrgm3-dependentmanner,suggestinga Masahiro Yamamoto, Support Staff Support Assistant Professor Professor croleinvacuolar-antigen-specifi T. gondii PVs,followingIFN-γstimulation,and 1 Youngae Lee Jisu Ma Sasai Miwa Masahiro Yamamoto cCD8 PhD + Tcellacti- c ■ Recent Publications AIM2 and Caspase-11-NLRP3 Infl AIM2 andCaspase-11-NLRP3 Kanneganti TD. Ligandsfor IRGB10Liberates by Bacterial Sensing the Kuriakose Malireddi RK, T, Peters JL,NealeG,Brown SA, Yamamoto M, R,SasaiM,Place DE,Kesavardhana S, SM,Karki Man Temirov J, Frase S,ZhuQ, ammasomes. Cell. 167,382-396 (2016). Toxoplasma VacuolarAntigen p62 PlaysaSpecifi Figure. c RoleinInterferon-γ-InducedPresentationofa ■ Reports 13,223-233(2015). Reports presentationinterferon-γ-induced ofa Toxoplasma vacuolarantigen. Cell Yamamoto M(Corresponding author).p62plays aspecifi Lee Y, N,OhshimaJ, J, SasaiM,Ma BandoH,Saitoh Sakaguchi T, S, Akira c role in 37 Immunology 38 non-specifi their fl membrane. Amongthesethreefl translocate PtdSerfromoutertoinnerleafl and ATP11C)theirsubunitCDC50Aasfl sperm, agederythrocytes,exosomes,andenvelopedvirus. sure isalsoobservedinactivatedlymphocytes,capacitated the activatedplateletsforbloodclottingfactor.ThePtdSer-expo- works asan“eatme”signalofapoptoticcells,andscaff to PtdSer-exposure.ThePtdSer,thusexposedthecellsurface, tween theinnerandouterleafl scramblase(s) thatnon-specifi asymmetrical distributionofphospholipidsisdisruptedbya When cellsundergoapoptosis,orplateletsareactivated,the and phosphatidylethanolaminefromoutertoinnerleafl ATP-dependent phospholipidfl asymmetrical distributionofphospholipidsismaintainedbyan ma membranes,isexclusivelylocalizedintheinnerleafl (PtdSer), oneofmostabundantphospholipidsineukaryoticplas- tributed betweeninnerandouterleafl destroyed duringapoptosis.Ahigh concentrationofCa tion sitesinthemiddleofmolecules, andtheirfl cytes. ATP11AandATP11Ccontain twoorthreecaspase-recogni- expressed invariouscellsincluding lymphocytesandhepato- expressed inbrains,whileATP11AandATP11Careubiquitously There aretwofamilies ofmembraneproteinsthat support We recentlyidentifi Phospholipids inplasmamembranesareasymmetricallydis- ippase activity,too. Biochemistry andImmunology Biochemistry c scramblingofphospholipids atplasmamembranes. ed threeP4-typeATPases(ATP8A2,ATP11A cally scramblesphospholipidsbe- ets ofplasmamembranes,leading ippase, thattranslocatesPtdSer ippases, ATP8A2isspecifi es Phosphatidylserine ets. ippases thatactively ets oftheplasma ippase activityis 2+ olds on inhibits et. This et. cally ets. Ca family, thatcontain10transmembranesegments,functionas Five members(TMEM16C,16D,16F,16Gand16J)oftheTMEM16 scramblase toexposePtdSerthecellsurface(Figure1). ibly inactivatesthefl a Ca intestines. HumanandmouseplateletsexpressonlyTMEM16Fas are tissue-specifi TMEM16F isubiquitouslyexpressedinvariouscells,whileothers Among thesefi re-establishes theasymmetricaldistributionofphospholipids. are activated,thehighintracellularconcentrationofCa specifi macrophages for engulfment.Xkr4andXkr9are rathertissue- Xkr9 scramblases,toquicklyandirreversibly exposePtdSer(Figure2). and ATP11Cfl apoptosis, caspasecleavesand irreversibly inactivatesATP11A cleaved bycaspaseduringapoptosis. Thus,whencellsundergo caspase-recognition siteintheC-terminaltailregionandare bling phospholipidsduringapoptosis.Thesememberscontaina supposed tocarry10transmembranesegments,supportscram- loss offunctionmutationinTMEM16F. drome, ararecongenitalbleedingdisorder,werefoundtocarry thrombin forbloodclotting.Infact,humanpatientsofScottsyn- cannot exposePtdSer,leadingtothereducedabilityproduce When theintracellularCa The PtdSer,thusonthedeadcell’s surface,isrecognizedby Three members(Xkr4,Xkr8andXkr9)oftheXkrfamilythatare 2+ -dependent scramblases by forming a dimer. Thus, when cells -dependent scramblasesbyformingadimer.Thus,whencells 2+ -dependent scramblase.TheTMEM16F cally expressedinthebrainsand intestines, respectively. Shigekazu Nagata, Support Staff Support Visiting Scientist Research Assistant Postdoctoral Fellow Associate Professor Professor ippases, andcleavesactivatesXkr4,Xkr6, c; theyarespecifi ve membersthatfunctionasscramblases, ippases, andreversiblyactivatesTMEM16F 2+ levelreturnstonormal,thefl cally expressedinthebrainsor 1 1 3 4 SegawaKatsumori Shigekazu Nagata Shigekazu -/- activatedplatelets PhD 2+ revers- ippase are releasedfromdeadcells.Thismayactivatetheimmunesys- lieved toundergosecondarynecrosis,andintracellularmaterials engulfed bymacrophage.Theunengulfeddeadcellsarebe- undergo apoptosis,theydonotexposePtdSerandcanbe In activatedplatelets,theintracellularCa and aCa The flippasecomprisedofP4-ATPase(ATP11AorATP11C)andCDC50A, Ca Figure 1.ThemolecularmechanismforPtdSerexposureincellswithhigh on theintracellularconcentrationofATPandCa transiently exposedtothecellsurfaceinthisprocess,andlikelydepends P4-ATPases resumeflippingPtdSerandPtdEtn.Thus,isonly returns tonormallevel,TMEM16Fstopsscramblingphospholipids,while macrophages. of PtdSerpreventsthePtdSer-exposingcellsfrombeingengulfedby ATPases andreducestheirflippingactivity.WhentheCa activates TMEM16Ftoscramblephospholipids,whileitinactivatesP4- Xkr8 Whereas, Xkr8isubiquitouslyexpressedinvarioustissues.When ■ ■ ■ Recent Publications 2+ motility. Mol. Cell Biol. 36,645-659(2016). Gyobu S,etal. Arole of ascramblingdomaininsperm TMEM16E carrying 9509-9514 (2016). in apoptotic phosphatidylserineexposure. Proc. Natl. Acad. USA113, Sci. J, E,NagataS. Imanishi Suzuki phospholipid scramblingcomplexThe Xkr8 Rev. Immunol. 17,333-340(2017). Nagata S, Tanaka M.Programmed cell deathandtheimmunesystem. Nat. -concentration. -/- mouseembryonalfi 2+ -dependent scramblase(TMEM16F)areschematicallyshown. broblasts (MEF)andfetalthymocytes 2+ concentrationincreasesand 2+ . Theconstantflipping 2+ concentration the fl membranes, andalsothephysiologicalpathologicalrolesof and scramblasesfl disease. tem, leadingtosystemiclupuserythematosus-typeautoimmune recognized bymacrophagesforengulfment. is anirreversibleprocess,andthePtdSerexposedoncellsurface while thesamecaspasescleaveandinactivateATP11AATP11C.This of thecaspase-cascadecleavesXkr8toactivateitsscramblaseactivity, When cellsundergoapoptosis,caspase3or7inthedownstream (ATP11A/ATP11C associatedwithCDC50A)areschematicallyshown. A caspase-dependentphospholipidscramblaseofXkr8,andflippase Figure 2.ThePtdSer-exposureinapoptoticcells. ■ ■ We arestudyingthemolecularmechanismofhowfl apoptotic phosphatidylserineexposure. Science344,1164-1168(2014). cleavage etal. ofphospholipidfl Caspase-mediated Segawa K, mouse platelets. Proc. Natl. Acad. USA112, 12800-12805(2015). Sci. releasephosphatidylserine exposure inactivated andmicroparticle Fujii T, NagataS. S,Eto K, A,Nishimura Sakata TMEM16F isrequired for ippases andscramblases. ip andscramblephospholipidsatplasma ippase for ippases 39 Immunology 40 stimulation arelimited andthereiscompetition amongTcells in additiontoinhibition ofactivation-signalingwhen ligandsfor tion withAPCiscrucialformaintaining stableunresponsiveness and experimentalstudies.Modeling showedthatTcellinterac- teractions whichstabilizethetolerant statewithboththeoretical water exposure. tion ofMnandevaluatedtheaccumulationwithdrinking cedure wedevelopedsuccessfullydelineatedthespatialdistribu- green signal)appearedmostprominentlyinamygdala.Thepro- histogram indicatesthatMnaccumulation(pixelswithhigh ter (5:3:2),andimagedusingtwo-photonmicroscopy.The nol solution,embeddedinamixtureofglycerol,ethanolandwa- sue sectionswerestainedwith20mMMorinhydratein60%etha- harvested andcryo-sectionedusingthestandardprocedures.Tis- neurons throughvoltagedependentCa ic contrastagentinmagneticresonanceimaging.Mn Immune responsesmodelling in tissue,micewereexposedtoMn fl Use ofMorinhydrateformanganesemapping ing watercontaining50mMMnCl visualization ofthedistributionMn metals shiftsfromaredtogreenemissionpeak,andthusitallows ical isolatedfromMaclurapomifera,whichwhenitbindsdivalent as anindirectmarkerofcalciumchannelactivity.Morinisachem- uorescence microscopy.TodemonstratethedetectabilityofMn We showedthatregulatoryTcells (Treg)cellsenhancecell-in- Recently, manganese(Mn)becameanimportantparamagnet- Single MoleculeImaging 2 . Thebrainsofthesemicewere 2+ for5weeksbywayofdrink- 2+ 2+ atacellularlevelunder channels,andthusacts 2+ canenter specifi (Fig. 1).TorobustlyinhibitTcellproliferationinthepresenceof manipulation ofimmuneresponsesinvariousclinicalsettings. dissociation, whichgivesinsightintoimmunetoleranceand experiments revealedanovelabilityofTregcellstoinhibitcell- combined studiesoftheoreticalmodelingandimmunological ation ofantigenspecifi vivo. TransientreductionofTcellnumberenhancedtheprolifer- model, wealsofoundawaytomanipulateimmuneresponsein amined invitroexperiments.Basedonthepredictionsof the proliferationpatternsofTcellsundervariousconditionsex- nate reactivityunderselfandnon-selfconditionswellreproduced latory ligandsonAPC.Notably,themodeloptimizedtodiscrimi- sion ofadhesionmoleculesandinhibittheexpressioncostimu- showed antigen-specifi tion andactivation.Supportingthistheory,invitroexperiments cally, Tregcellswererequiredtoinhibittwoprocesses;dissocia- cent Ca was monitored.Tcellactivationcan bemonitoredusingfl we manipulatedsingleTcelltointeractwithAPCandactivation Observation ofsingleTcellactivation the immunesystem distinguishesbetweenselfand non-selfde- that activationof Tcellsisnotdeterministic.Tounderstand how showed activationevenwhenantigen wasabsent,indicating antigen waspresentedbyAPC.On theotherhand,11%ofTcells attached toAPC,wefoundthatonly 22%becameactivatedwhen To answerhowmanyTcellsareactivatedbycontactwithAPC, c TregcellsevenwhereindividualTbehavestochasti- 2+ -reporter. Bycountingthenumber ofactivatedTcells Toshio Yanagida, Support Staff Support Visiting Scientist Postdoctoral Fellow Associate Professor Professor c Tcellsindraininglymphnodes.Thus,the c Tregcellsactuallyenhancetheexpres- 1 2 1 Yutaka Komai Tomoyuki Yamaguchi FujitaHideaki Toshio Yanagida PhD uores- ence ofTregcells,activationTcellswassuppressedinthe gated theeff spite thenon-deterministicmannerofTcells,wefurtherinvesti- ■ ■ ■ Recent Publications troscopy. Rep. Sci. 6,37562(2016). and Fujita spec- H.Non-labelimmunecell state using Raman prediction Ichimura T, ChiuLD, Fujita H, Machiyama K, Yamaguchi T, Watanabe TM One. 11(9),e0163134(2016). AntigenPresentationsNon-Self by Regulatory T Cell Suppression. PLoS Furusawa C, Yamaguchi T. RobustandAccurate DiscriminationofSelf/ fl scattering andanti-Stokes ofRaman by thesimultaneousobservation and Fujita Protein K. expression guidedchemicalprofi Chiu LD, Ichimura T, Sekiya T, H, Machiyama Watanabe T, Fujita H,Ozawa T, uorescence Rep. emission.Sci. 7,43569(2017). ect ofTregcells(Fig.2),andfoundthatinthepres- cells asafunctionofthefractiontargetantigen(right). T cellpopulationwithrespecttoantigenpresentation(middle),andaveragenumberofTreg Multicellular modelofTcellresponse.Schematicillustrationthe(left),response Figure 1. present onAPC. fl Rhod2 stainedTcells(red)wereheldwithaglassmicro-needleandplaced onAPCand Counting TcellactivationuponengagementwithAPCinthepresenceofregulatory Tcell. Figure 2. uorescence wasmonitored.RegulatoryTcell(blue)andconventional(green) arealso ling oflivingcells show thatTregcellsactasafaucetforleakyimmunesystem. not changeinthepresenceofantigenpeptide.Thisobservation absence ofantigenpeptide,whereasactivationprobabilitydid ■ ■ 13788 (2016). caused by interplay between RNApolymerasesonDNA.Nature Comm. 7, M, Fujita Iwaki K, Yanagida T. Transcriptional burstingisintrinsically sin VI heads. Nature Comm. 7,13715(2016). origami nanospringthatreveals force-induced adjacent bindingofmyo- M, K, Iwaki S,Ikezaki Wickham Yanagida T, Shih W. Aprogrammable DNA 41 Imaging 42 Clin. Invest.2012). Moreover,wedemonstrated that vitaminD ing knockoutmice defi S1P-mediated migratorycontrolof bonecellsbyS1Pgenerat- Exp Med2010).Wealsoshowed thesubstantialcontributionof precursors inconcertwithvarious chemokines(Nature2009;J a keyfactorincontrollingthemigratory behaviorofosteoclast this methodology,weshowedsphingosine-1-phosphate (S1P)as physical couplingwithbone-reformingosteoblasts.Byutilizing mode ofbone-resorbingfunctioninvivoandthefunctional gration andpositioningoftheirprecursormacrophages, behaviors ofbone-resorbingmacrophage,osteoclast,i.e.,themi- visualizing insidethebones.Wehavefi tion invariouskindsoftissuesandorgans(seetheFigure). commitment andother‘niches’,celldynamicsduringinfl well asbiologicalsciencesuchhematopoiesisandcelllineage dating thedynamicsystemsinbonebiologyandimmunologyas dition tothespatialandstructuralinformation.Wearenoweluci- nature ofdiff vanced imagingtechniques,wehaveinvestigatedthedynamic various kindsoftissuesandorgansinvivo.Bymeansourad- stand thefundamentalprinciplecontrollingcellulardynamicsin nicating atdefi types aremigratingthroughoutthebodyandmutuallycommu- teoblast dynamicsinvivo Intravital boneimagingrevealingosteoclastandos- Our labhasoriginallyelaboratedthenovelimagingsystemfor The immunesystemishighlydynamicandavarietyofcell Immunology andCellImmunology Biology erent celltypesinatime-dependentmanner,ad- ned sites.Theaimofourlaboratory istounder- cient forendogenousS1Ptransporter (J. rst elucidatedtheinvivo amma- with bone-formingosteoblastsand otherbonecelltypes. bone resorptionwhichwasfi this probe,wecouldgrasptherealtimecourseofosteoclastic bone resorptionbyosteoclasts(Nat.Chem.Biol.2016).Byusing veloped anewchemicalprobefordetectingprotonsecretionin collaboration withDr.KikuchiintheframeofIFReC,wehavede- nism underlyingboneerosioninarthriticjoints.Furthermore, teoclasts directlytoconvertthemtheRstate,acriticalmecha- stimulate osteoclasticbonedestructionbycontactingNstateos- novel imagingsystemshowedthatRANKL-bearingTh17could on theirsurface,althoughitsfunctionalroleremainselusive.This Th17 cells,abonedestruction-proneTcellsubset,expressRANKL tively looselyattachedandwrigglingalongthebonesurface. trix bysecretingacids,andnon-resorbing(N)osteoclastsrela- teoclasts fi mature osteoclastfunctionalstates;i.e.,bone-resorbing(R)os- Clin. Invest.2013).Thisnovelvisualizationidentifi ceeded invisualizingthefunctionofmatureosteoclastsvivo(J. Natl. Acad.Sci.USA2013). ed migrationcontrolofosteoclastprecursormonocytes(Proc. signifi understood. Using metabolomicsapproach,wefound thatmeta- ‘metabolic reprogramming’, butitsroleinosteoclast isnotfully cell diff Immunometabolism ofbone homeostasis Through theimprovementofboneimagingsystem,wesuc- Osteoclasts dramaticallyaltertheir metabolicactivityduring cantly blocksbonedestructionbymodulatingS1P-mediat- erentiation. Thischange inthemetabolicstatusistermed rmly adheringtobonesanddevouringthebonema- Masaru Ishii, Support Staff Support Visiting Scientist Research Assistant Assistant Professor Associate Professor Professor nely regulatedbycell-cellcontact MD/PhD 3 1 2 Yutaka Uchida(SA) Szandor Simmons(SA) Takahiro Matsui (SA) Takao Sudo Mizuno Hiroki Junichi Kikuta Keizo Nishikawa Masaru Ishii ed twodistinct in theregulationofcellularmetabolismanddiff osteoporosis. Thus,werevealedtheroleofepigeneticprocesses tion bytheafl number ofosteoclasts.Furthermore,inhibitionDNAmethyla- defi fi mt3a-defi via epigeneticrepressionofanti-osteoclastogenicgenes.Dn- ated DNAmethylationbyDnmt3aregulatesosteoclastogenesis changes toosteoclastdiff mt3a) asatranscriptionfactorthatcouplesthesemetabolic more, weidentifi tion reactions,includinghistoneandDNAmethylation.Further- methionine cycle.SAMistheuniversalmethyldonorformethyla- intracellular S-adenosylemethionine(SAM),ametaboliteofthe bolic reprogrammingduringosteoclastdiff ■ ■ ■ ciently intoosteoclastsandthatmicewithanosteoclast-specifi Recent Publications S100A8 inobesefat. Proc. Natl. Acad. USA112,2058-66 (2015). Sci. R,etal. Sekimoto Visualized macrophage dynamics andsignifi probe. Nature Chem.Biol. 12,579-85(2016). usingdesigned smallmolecular andmotility with cell osteoclast activity H, etal.Maeda Real-timeintravital imaging ofpHvariation associated 19(5), 530-541.doi:10.1038/ncb3509(2017). oftransformed cellsextrusion through metabolicchanges. Nat.Cell Biol. Kon S,etal. Cell competition withnormalepithelialcells promotes apical ciencyinDnmt3ahaveelevatedbonemassduetoasmaller cient osteoclastprecursorcellsdonotdiff avin-3,3΄-digallate abrogatedbonelossinmodelsof ed the ed de novo erentiation. WefoundthatSAM-medi- DNAmethyltransferase3a(Dn- vivo behaviorswithadvancedimagingtechniques. principle controllingthedynamicnatureofimmunecellsbyvisualizingin organs, byforminga‘soft-wired’network.Weareelucidatingthebasic Immune cellsarehighdynamicandinterconnectingvarioustissues Figure. Intravitalimagingforvariousimmunesystems. erentiation increased erentiate ef- erentiation, cance of cance c strategy foravarietyofbonedisorders(NatMed2015). which mayprovidethemolecularbasisforanewtherapeutic future. sues, andwewillbeabletodissecthumanimmunologyinthe ceeded invisualizingnon-labelledhumannormalandcancertis- tion tohumantissuesandorgansinvivo.Todate,wehavesuc- AMED), wearedevelopinganewmicroscopysystemforapplica- human samples.Bycollaboratingwithcompanies(supportedby limited inanimalmodelsandmaynotbeapplicableforanalyzing system dynamicsinvivo.However,theapplicationiscurrently intact tissuesandorgansthususefulforstudyingimmune doubtedly powerfultoolfordissectinglivingcellulardynamicsin secting humanimmunology Application ofintravitalimagingtechniquesfordis- ■ ■ ■ Intravital imagingwithmulti-photonmicroscopyisanun- D. Proc. Natl. Acad. USA 110,7009-13(2013). Sci. vitamin ofactive a criticalpointofcontrol action inantibone-resorptive J,Kikuta etal. S1P-mediated osteoclast migration precursor is monocyte J. Clin. Invest.clast function. 123,866-873(2013). J, visualizationofRANKLand Kikuta etal. Dynamic Th17-mediated osteo- Med. 21,281-7(2015). metabolic pathway.pling to anS-adenosylmethionine-producing Nat. etal. Dnmt3aregulates K, osteoclast diff Nishikawa erentiation by cou- 43 Imaging 44 relative toSUVinbilateralthalami (SUVt)wassignifi also assessed. and thelesionnumberinT1-T2-weightedMRimageswere C-11 acetateuptake.Correlationbetweenuptake Mapping 8(SPM8)toevaluatepathologicalchangeofregional metric analysiswereperformedbyusingStatisticalParametric tion techniqueforco-registeredMRI.Voxel-basedstatisticalpara- gray matter(GM)and/orwhite(WM)basedonsegmenta- were producedforanalysis.Volume-of-interest(VOI)analysis and theratioofregionalSUVtomeaninthalamus(SUVt) method. Parametricimagesofstandardizeduptakevalue(SUV) images werereconstructedusingafi started 20minafterintra-venousinjectionofC-11acetate.The PET scanningwasperformed.Astatic20-minacquisition were enrolledinthepresentstudy.Inallsubjects,MRIand old (yo))andhealthyvolunteers(HV)(6females,58+/-5.8yo) means ofPETwithC-11acetate. astrocyte intheCNSandtoclarifyactivityMSvivoby in theCNS.Weaimedtovisualizeandinvestigateactivationof revealed tobeselectivelyuptakenandmetabolizedbyastrocytes vous systemofmultiplesclerosis(MS)patients.Acetatehasbeen As aresult,standardizeduptake value(SUV)of11Cacetate Patients withrelapsing-remittingMS(6females,46+/-5.9year Astrocyte activationisrepresentativepathologyincentralner- Nuclear Medicine lee-ak projection ltered-back cnl in- cantly tool. static C-11acetatePET,whichmay beaneff changes inpatientswithMScanbedetectednon-invasive MS pathophysiology. Thus, thealteredastrocyteactivationispresumablyinvolvedin edge ofacuteMSlesions,followedlaterbyastrocyticscarring. an animalmodel;astrocytehypertrophyoccursattheleading ing WMandGM.Astrocytepathologyprecedesdemyelinationin ordinary MRI,pathologicalchangesexisteveninnormalappear- pathology. Inadditiontothedemyelinatedlesionsdetectedby widespread involvementofcorticalastrocytesinMSpathology. axonal damageandcorticalastrogliosis.Theseresultsimplythe tate uptakerefl tate uptakeinwhiteandgraymatter.ItseemsthattheC-11ace- of tissuedestructionweresignifi numbers ofT2lesionsandT1blackholesrecognizedasamarker matter especiallyinsubcorticalwhiteMSpatients.The elucidated signifi nifi The ratioofSUVinwhitemattertograywassig- creased inbothwhiteandgraymatterMScomparedwithHV. Our presentstudysuggeststhatpathologicwhitematter Infl cantly higherinMSthanHV.Voxel-basedstatistical analysis ammatory demyelinationinWMisacardinal feature inMS Jun Hatazawa, Support Staff Support Research Assistant Assistant Professor Associate Professor Professor ecting reactiveastrocytescorrelateswithboth cant increaseinuptakebroadareaofwhite cantly correlatedwithC-11ace- 1 2 Tadashi Watabe Kato Hiroki Jun Hatazawa MD/PhD ective MSdiagnostic ■ ■ Recent Publications pone.0159505 (2016). Based StatisticalAnalyses. PloSone11,e0159505,doi:10.1371/journal. Can Improve of theLesion Detectability inBrainSPECT/CT Voxel-rection H,Shimosegawa E,FujinoKato K&Hatazawa J. CT-Based Attenuation Cor- halation PET. Res7,16.doi:10.1186/s13550-017-0263-6 (2017). EJNMMI tratumoral hypoxia: 18F-fl Watabe T, etal. Quantitative evaluation ofoxygen metabolisminthe in- uoromisonidazole gasesin- and15O-labelled uptake (SUVt)inpatientswithMScomparedtoHVpatients. volunteers. ColoredvoxelsindicateT-scoresrepresentingsignifi Voxel-based statisticalcomparisonC-11acetateuptakebetweenMSpatients andhealthy Figure 3. ratio ofregionalSUVtomeanthalamicSUV.) Standardize uptakevalueofC-11acetateintheMSpatientsandhealthyvolunteers.(SUVt: Figure 2. A representativecaseofMS.(SUV:standardizeduptakevalue,) Figure 1. ■ jnumed.115.164004 (2016). andDizygotic Monozygotic Twins. J. Nucl. Med. 57,392-397.doi:10.2967/ mental Infl Watanabe H,Shimosegawa S,Kato E&Hatazawa J. GeneticandEnviron- uences on Regional Brain Uptake of 18F-FDG: A PET Studyonuences onRegional BrainUptakeof18F-FDG:APET cantly increasedC-11acetate 45 Imaging 46 duce thebenefi or havelimitationintheavailability offl ated usingfl the currentprobesavailablefor no-washimagingwerecre- even whenfreeprobesarenotwashed outfromcells.However, imaging ofcell-surfaceproteins withlowbackgroundsignals, tact quenchingmechanism.Thisfl creases fl be introducedintotheprobe.Inaddition,NBquencherde- fl (NB) quencher.Thismodularscaff cinnamic acidligand,afl labeling reactionwithPYP-tag.FCANBconsistsofa4-hydroxy- the fl bacteria. Thenoticeablefeatureofthislabelingmethodisthat protein tag,whichisasmall(14kDa)derivedfrompurple tein (PYP)taganditsspecifi tein labeling/imagingsystembasedonphotoactiveyellowpro- multicolor pulse-chaseimaging.Previously,wedevelopedapro- addition ofprobeswithdiff cause targetproteinsareconditionallylabeledbythetemporal protein taganditsspecifi tained. Inthistypeofimaginganalyses,proteinlabelingusinga cise spatio-temporalinformationonproteinsinlivingcellsisob- ing thesetechniqueswithpulse-chaseimagingofproteins,pre- localization andmovementofproteinsinlivingcells.Bycombin- uorophores emittingfl In thisresearch, to achievemulticolorproteinimaging with Multicolor imagingtechniquesareusefulforinvestigatingthe uorescence ofFCANBisquenchedandincreaseduponthe Chemical Imaging Chemical Imaging Techniques uorescence intensityofthefl uorophores emittingshort-wavelengthfl t inmulticolorimaginganalyses. uorescence atdiff uorophore, alinker,andnitrobenzene cprobesisanattractiveapproach,be- c probe,FCANB.PYP-tagisanovel eet fl erent uorogenic propertyallowsthe old isattractive,sincevarious uorophores thatallowthe uorophore basedonacon- uorophore, andthusre- erent wavelengths can uorescence ferent long-wavelengthfl high contrast,wesynthesizednewPYP-tagprobeswithtwodif- 570 nmand670nm,respectively.Thesefl Cy5, whichemitfl length fl cence intensityuponthelabelingreactions.Aslong-wave- or insituimagingofvariousproteins. fl proach fordesignoffl strategy basedonDNBquenchingwouldoff wash imagingofthecell-surfaceproteinswithhighcontrast.The the goodfl aged EGFRinlivingcells(Fig3).Particularly,Cy5DNB2showed well asfl FCANB. ItturnedoutthatDNBincreasedthelabelingreactionsas labeled PYP-tagmorerapidlythanthepreviouslyreportedprobe, enhanced uponlabelingreactionsofPYP-tag(Fig2).Theprobes covalently. Asweexpected,fl SA-PAGE analysesrevealedthatboththeprobesbindtoPYP-tag fold oftheprobetodevelopCy3DNB2andCy5DNB2(Fig1).SD- based oncontactquenching.WeintroducedDNBintothescaf- employed asaquencherthatreducesfl fl and havebeenappliedtoanumberofbiologicalstudies.As uorophores. Theseprobesthus willbeattractivetoolsforinvivo uorescence quencher,3,5-(dinitrobenzyl)acetamide(DNB)was uorophores, wechosesulfonatedcyaninedyes,Cy3and uorescence intensity.Boththeprobesspecifi Kazuya Kikuchi, uorogenic andkineticpropertiesallowedno- Support Staff Support Visiting Scientist Research Assistant Postdoctoral Fellow Assistant Professor Associate Professor Professor uorescence withthemaximumwavelengths of uorogenic probeswithlong-wavelength uorophores, whichincreasefl uorescence intensityoftheprobes 1 2 2 1 Masafumi Minoshima Yuichiro Hori Kazuya Kikuchi Kazuya PhD uorophores arebright ursec intensity uorescence er apromisingap- cally im- cally uores- ■ ■ Recent Publications Osteoclast Activity. Nat. Chem.Biol. 12,579-585(2016). ofpH Real-time Intravital Imaging K. Kikuchi Variation Associated with H,KowadaMaeda T, S,IshiiM, J, Kikuta Furuya M,Mizukami M,Shirazaki Chem. Biol. 12,853-859(2016). veal aRoleofGLUT4 N-glycosylation inIntracellular Traffi S,Hori Hirayama Y, BenedekZ,Suzuki T, Fluorogenic K. Kikuchi Probes Re- absence ofPYP-tag,respectively. fluorescence spectraofCy5DNB2inthepresenceand reactions withPYP-tag.Redandbluelinesrepresent Fluorescence enhancementofCy5DNB2uponlabeling Figure 2. Principle offl Figure 1. uorogenic PYP-taglabelingsystemusingCy5DNB2. kn, Nat. cking, ■ Live-Cell Imaging of Intracellular Proteins, ofIntracellular Imaging Live-Cell 7,308-314(2016). Chem. Sci., Design ofaProtein tagandFluorogenic Probe for Structure withModular Kamikawa Y, Hori Y, Yamashita S,StandleyD.M, JinL,Hirayama K, K. Kikuchi Live-cell imagingofPYP-EGFRusingCy5DNB2. Figure 3. 47 Imaging 48 the natureof measurements,mostcrucially the laserfocus a measurementof theendogenousmolecules,but dependingon cell typeinamixedgroupofcells. TheRamananalysisisbasedon cell typewasdistinctenoughthat itcouldbeusedtopredictthe al., 2016).Ofnoteisthefi to discriminatebetweendiff markers. Werecentlycompleted a studyusingRamanscattering munological functionarethepresenceorabsenceofsurface levels, usuallyinlymphocytesandmacrophages. as analysisofcellphenotype,andtodiscriminateactivation that wehavedevelopedtostudyprocessesinimmunecells,such derway, wearenowmovingmoretowardsapplyingthetools measurements. Withseveralyearsofdevelopmentalreadyun- can beusedinacomplementarymannertotheRaman-based sue sample.Wealsoinvestigatehowotherlight-basedmethods ical distributionsofendogenousmoleculesinalivingcellortis- age wheretheimagecontrastisdefi expressed atdiff in thewavelengthofscatteredlight.Withdiff ments, usingahighintensitylaserbeam,andmeasuringtheshift mation. WerelyheavilyonRamanscatteringbasedmeasure- can beexploitedtomeasuresamplesandextractcellularinfor- tral data,orboth.Severaldiff surements, withoutfl cal processes.Mostofourresearchinvolvesusingopticalmea- techniques andapplythemtostudybiologicalimmunologi- For TandBcells,theusualfeaturesfordiscriminationofim- The Biophotonicslabresearchthemeistodevelopoptical Biophotonics erent locationsinthecell,wecanproduceanim- uorescent labels,toprovideimages,spec- nding thattheRamansignatureofeach erent lymphocytecelllines(Hobroet erentphysicalinteractionsoflight ned bythespatialandchem- erent molecules of ametalsurface inthecell,andarenotstrictlylabel-free butthe lion times.These particularexperimentsrequirethe introduction face. Theenhancementfactorcan thenbeintheorderof1mil- chemical bondsthatformbetween targetmoleculesandthesur- the scatteringsignalisamplifi suffi el) inthecell,however,Ramanscattering alonecannotgenerate of drugsinsolution. samples (e.g.Hobroetal.,2013),evaluationoftissue,oranalysis required. Thisincludesdetectionofpathogensinbloodorserum cuvette-based samples,orothersampleswhereimagingisnot table Ramandetectionsystemwhichcanbeusedforanalysisof man withsomeadvantagesoverothermethods.Webuiltapor- molecules suchasmRNA,andproteinscanbemeasuredbyRa- small moleculeinteractions).Theinteractionsbetween we canmeasurepurifi ration modelofindividualcelllines. and specifi individual celltypesfromoneanother,resultedinhighsensitivity method, usingitasamethodofdiscriminatingTvsBcellsand/or plications ofthisapproachgobeyondasimplediscrimination more usualwet-labmethodsofcellphenotyping.Whiletheim- nents ofthecell.Thisgivesacomplementaryapproachto be morerelatedtothegenomeortranscriptome-basedcompo- surface markers.ThereforetheRamansignatureshouldinstead and opticalsectioning,itshouldnotdetectcontributionsfrom For theultimatesensitivity(approachingsingle-moleculelev- We alsousedRamananalysisinanon-imagingmode,where cient signal,butbyusing plasmonicRamanenhancement, city (Hobroetal.,2016).Figure1showsaPLS-DAsepa- Nicholas IsaacSmith, Support Staff Support Assistant Professor Associate Professor ed samplesofinterest(e.g.mRNAand ed atametallicsurfaceand/or by 1 Nicolas Pavillon Alison Hobro Nicholas IsaacSmith Nicholas PhD needed toformanimage(Pavillonetal.,2016).Oneofthemain reconstruction modelstoreducetheamountofdatapoints ect oncompressedsensing,usingsub-samplingandoptimized best tooptimizethemeasurements,andcompletedourfi thereby creatingtheRamanenhancementdirectlyinsidecell. nanoparticles in-situbyphotoreduction(Smithetal.,2014), lution, whichcanpassthroughtheentirecell,andtoform regions. Anovelmethodtosolvethisisintroducegoldsaltso- the nuclearmembrane,restrictingmeasurementstocytosolic cles, andevenforthosethatdo,theparticlesdonottypicallypass can bediffi surface insidethecell,usuallyaninertmetalnanoparticle.This enhanced Ramanexperimentsistheneedtointroducemetal ecules withoutanyconjugatedtargeting.Onediffi uses onlytheinterfaceenhancementtodetectendogenousmol- ally similartothemorewell-knownSPRsensors,ourapproach tradeoff 2016. a dividinglinebetweentheB-andT-cellspectra.FigurefromHobroetal. set asideandusedtovalidatethemodel.Thereddashedlinerepresents separation modelisbuiltfrompartofthedata,withaportiondata cell lineisdenotedbydiff sample representsoneRamanspectrafromanindividualcell,whereeach B- andT-celllinediscriminationbylabel-freeRamanmeasurement.Each Figure 1. ■ ■ ■ Recent Publications mozoin. Analyst 140,2350-2359 (2015). imaging ofthemacrophageRaman response to themalariapigment he- Hobro Pavillon AJ, N,FujitaM,Coban CandSmithNI.Label-free Ozkan K, (2016). for cell label-free linediscrimination. Analyst lymphocyte 141,3756-3764 Hobro Kumagai AJ, Y, spectroscopy asatool SandSmithNI.Raman Akira Opt. Express 24(26),30038-300052 (2016). Pavillon NandSmithNI.Compressed sensinglaserscanningmicroscopy. Due tothechallengesofRamanimaging,weconsideredhow isoftenworthwhile,dependingonthetarget.Conceptu- cult sincesomecellsdonotreadilyuptakenanoparti- erent symbolsdescribedinthefi culty for these gure key.The rst proj- groups thatusefl tobleaching whichisveryusefulforcollaborationswithother nifi multiphoton fl plied tolaserscanningmicroscopyingeneral,suchasconfocalor we showedthatthecompressedsensingapproachcanbeap- loss. Althoughourlabisprimarilyfocusedonlabel-freeimaging, the imagecanstillbereconstructedwithminimalinformation process. Asshowninfi ing properties,wecansignifi set ofmeasurements,incombinationwithamodeltheimag- measurements intypicalimagingconditions.Byusingareduced aging system,thereissomeamountofredundancybetween quired. However,duetotheinherentblurinmicroscopeim- sample damage.Therefore,relativelylongimagingtimesarere- er laserpowerisusuallynotpossibleduetothedesireavoid problems inRamanmicroscopyisthesignal-to-noiseratio.High- from Pavillonetal2016. C-H stretchingband(2925cm−1,green)andlipids(2853red).Figure the compressedsensingmethod.Thecolorcontrastisgeneratedfroma reduction insamplingstillproducesusableimageswhencombinedwith only 1/3(c1)or1/5(c2)oftheoriginaldata,showingthatuptofi are thenmeasuredatfullresolution(b),andreconstructedusing microscope, measuredwith100nmfl measurements. Image(a)showsthepointspreadfunctionofRaman can signifi Raman measurementsonMEFcells,showinghowcompressedsensing Figure 2. ■ ■ Biophys. J. 105(5),1123-1132 (2013). Composition Revealed by SimultaneousMultimodalLabel-Free Imaging. Pavillon andSmithNI.Cell andMolecular N,Hobro OpticalDensity AJ ticles insidecells. Nat.Commun. 5(5144), 1-9(2014). Fujita KandKumagai Y. Laser-targeted photofabrication of goldnanopar- S,Pavillon H,Ichikawa N,Hobro Niioka AndoJ, AJ, K, Smith NI,Mochizuki cant improvementinimagingspeedand/orreductionpho- cantlyimproveRamanimagingspeedbyreducingtherequired uorescence imaging.Thiscanthenleadtoasig- uorescent probes. gure 2,usingupto5timeslesssampling, cantly speeduptheRamanimaging uorescent beads.TheRamanimages vetimes 49 Imaging 50 cyte traffi Physiological signifi adaptive immuneresponseswas unclear. (Fig. 1).However,thephysiological roleofthismechanismin tion oflymphocytes,andconsequently inhibitstheirLNegress CCR7 andCXCR4,chemokinereceptorsthatpromoteLNreten- vation oflymphocyteβ Control oflymphocytetraffi phocyte egressfromLNsthroughβ recently reportedthatinputsfromadrenergicnervescontrollym- to otherlymphoidorganscontinueantigensurveillance.We cytes exitfromtheLNintolymph,returntobloodfl and adaptiveimmuneresponses. role ofadrenergicnervesinthecontrollymphocytetraffi among lymphoidorgans.Therefore,wehavebeenstudyingthe the inputsfromadrenergicnervesaff a richsupplyofadrenergicnerves.However,itwasunclearhow receptors (ARs).Likeothervitalorgans,lymphoidorgansreceive induces cellularresponsesthroughα the sympatheticnervoussystemandproducenoradrenalinethat largely unclear.Adrenergicnervesconstitutetheeff cellular andmolecularbasisforneuralregulationofimmunityis sponses areinfl ceptors The activityofadrenergic nervesdisplaysacircadian rhythm After spendingseveralhoursinalymphnode(LN),lympho- It haslongbeenproposedthatvariousaspectsofimmunere- Immune ResponseDynamics Immune cking uenced bynervoussystemactivity.However,the cance ofadrenergic controloflympho- 2 ARs enhancestheresponsivenessof cking through β 2 ARs (Nakaietal.,2014).Acti- 1 , α ect lymphocytemigration 2 , β 1 , β 2 andβ 2 -adrenergic re- ow, andtravel erentarcof 3 adrenergic cking These fi ished whenlymphocyterecirculationthroughLNswasstopped. was dependentonβ the daytime.Thediurnalvariationofhumoralimmuneresponses more robusthumoralimmuneresponsesthanimmunizationin night time,whenlymphocytenumbersinLNswerehigh,induced mune responses.Wefoundinmicethatimmunizationthe the chanceofantigenencounterandpotentiateadaptiveim- during theperiodofhighadrenergicnerveactivitymayincrease encounters withpathogensaremore likelytooccur(Fig.2). have evolvedtomaximizetheeffi diff tive immuneresponses(Suzukiet al.,2016).Thetime-dependent phocyte traffi neural inputstolymphocyteβ in mice.ThediurnalvariationofLNegresswasdependenton lymphocyte egressfromLNswasrestrictedduringthenighttime and blood.Consistentwiththeseobservations,wefoundthat cyte numbersinLNsandtheirconcomitantdecreaselymph nerve activityinLNswasaccompaniedbyanincreaseoflympho- ward thenighttimeinmice.Thesurgeofadrenergic dents. Indeed,thenoradrenalinecontentinLNswaselevatedto- the daytimeinhumans,whereasitreachesapeakatnightro- noradrenaline releasefromadrenergicnervesincreasesduring that issynchronizedwiththerest-activitycycleofspecies.The We hypothesizedthattheaccumulationoflymphocytesinLNs erences inthemagnitude ofadaptiveimmuneresponsesmay ndings suggestthattheβ Kazuhiro Suzuki, Support Staff Support Research Assistant Assistant Professor Professor cking contributestothedailyfl 2 AR-mediated neuralsignalsandwasdimin- 2 ARs (Suzukietal.,2016). ciency ofhostdefensewhen 2 AR-mediated controloflym- 2 2 Nakai Akiko Kazuhiro Suzuki MD/PhD uctuation ofadap- ing toaddressthesequestionsrevealthewholepictureofad- and infl be visualizedhowadrenergicnervesinteractwithlymphocytes β solved. First,themolecularmechanismofcrosstalkbetween by thenervoussystem.However,afewquestionsremainun- Future perspectives ■ ■ 2 Recent Publications ARs andchemokinereceptorsisunclear.Second,itremainsto through lymphnodes. J. Exp. Med. 213,2567-2574(2016). the adaptive immune response recirculation by diurnallymphocyte Hayano K, Suzuki Y, A,Furuta Nakai FandNodaM.Adrenergic control of by adrenergicmunity Clin.Exp. nerves. Neuroimmunol. 8,15-22(2017). A.Control traffi KandNakai oflymphocyte Suzuki Our fi uence theirbehaviorsinLNs.Intheshortterm,wearego- ndings haveaddedanovellayerofimmuneregulation pathogens duringtheactivephase. of lymphocytetraffi lymphocyte accumulationinLNspromotesadaptiveimmuneresponses.This diurnal variation which leadstoanincreaseoflymphocytenumbersinLNs.Immunizationduring theperiodof During theperiodofhighadrenergicnerveactivity,lymphocyteegressfrom LNs isrestricted, Figure 2.Diurnalcontrolofadaptiveimmunitybyadrenergicnerves. Figure 1.ControloflymphocyteegressfromLNsthroughβ promote lymphocyteretentioninLNs,andinhibitsegressfromLNs. lymphocyte β Inputs fromadrenergicnervesstimulateβ 2 ARs enhancessignalsthroughCCR7andCXCR4,chemokinereceptorsthat cking mayhaveevolvedtomaximizetheeffi cking andadaptive im- cking 2 ARs expressedonlymphocytes.Activationof for ourselvesinthisrapidlygrowingfi lar attentioninbiomedicalscience.Wearegoingtocarveaniche nerves. Currently,theneuro-immuneinteractionsattractparticu- functions arerefl plan toclarifycellularandmolecularmechanismsbywhichbrain infl sympathetic nervoussystemandtherebyadrenergicnervesis renergic controloflymphocytetraffi ■ ■ egress from lymphnodesthrough β A,Hayano Nakai Y, Furuta F, Control K. oflymphocyte NodaMandSuzuki roimmunol. 7,10-17(2016). A.Autonomic control KandNakai ofinfl Suzuki 211, 2583-2598(2014). uenced bypsychologicalconditions.Inthelongerterm,we 2 ARs. ciency ofhostdefenseagainst ected inimmuneresponsesthroughadrenergic 2 -adrenergic receptors. J. Exp. Med. eld. cking. Theactivityofthe ammation. Clin.Exp. Neu- 51 Imaging 52 ways. tory responsethroughdescendingbrain-to-spinalcordpath- about injurytothebrain,andmodulatelocaltissueinfl peripheral andspinalcordcircuitsthattransmitinformation by topicalapplicationthecompoundcapsaicin.Thissensitizes to studyneurogenicinfl to probetheroleofnervoussystemintissueinfl cally), andregulatingautoimmunity(e.g.asinpsoriasis).Oneway ception), modulatingtheimmuneresponse(locallyandsystemi- fl tions formation science.Theresearchfocusesontwomainques- the brain,andcombinesimmunology,neurosciencein- My labstudiesinteractionsbetweentheimmunesystemand fl the brainresponse thatusesinformationabout peripheralin- odologies, wehavebeenableto chartthedynamicsignatureof Imaging (fMRI))inhumans.Bycombining thetwoimagingmeth- toencephalography (MEG)andfunctional MagneticResonance Brain responsesduringneurogenicinfl ammatory response,bysensingbacterialantigens(immuno- ammation tocontrolbehaviour. Wehaveidentifi We havebeenstudyingcapsaicin infl The nervoussystemplaysacriticalroleinthelocaltissuein- 2. Canweisolatebrain-basedbiomarkersforfatigueandpain 1. in infl fl How doesthebrainrespondtoperipheralneurogenicin- ammation? Brain-Immune Interaction ammatory arthritis(humansandrodentmodels)? ammation, whichcanbedoneinhumans ammation using(Magne- ammation. ed abrainre- ammation is amma- during jointinfl tifi model) inrodents(incollaboration withShionogi).Thishasiden- been studyingbrainactivityfollowing jointinfl in termsofsymtpomsfatigue andpain.Inparallel,wehave tients, weaimtoprospectivelypredict theirtreatmentresponse start anti-TNFtherapy,andlongitudinally followingupthesepa- toid arthritis.Furthermore,ourcohortinvolvespatientsaboutto thritis, andwearenowapplyingthesetopatientswithrheuma- by ‘decoding’patternsofbrainactivityforpatientswithosteoar- can predictbothpainanddepressionwithreasonableaccuracy but alsofortranslationwithanimalstudies. which isamajorproblemnotjustforhumanandclinicalstudies, lem isthatwehavenoobjectivebiomarkersforthesesymptoms, ease, andhowitrelatestoimmuneactivation.Partoftheprob- we understandaboutthenatureoffatigueininfl fl immunologic therapiessuchasantiTNF,despitethefactin- their disease.Critically,thesesymptomsoftenrespondpoorlyto pain arethemostseveresymptomstheyexperienceaspartof Biomarkers forfatigueandpainininfl can specifi mise furtherinjury.Thisisthefi information toguideprotectiveresponsesorientatedmini- gion thatseemscriticalforsensinginfl ammatory markersoftenrespondwell.Thishighlights howlittle ed abrain region(theanteriorcingulate cortex) thatisactive Patients withinfl Using bioinformaticapproaches,wehavedevelopedtoolsthat cally planbehaviourbasedoninfl Ben Seymour, Postdoctoral Fellow Assistant Professor Associate Professor Professor ammation,and is associatedwiththeleveloffa- ammatory arthritisreportthatfatigueand rst demontsrationthatthebrain MD/PhD 1 Maruyama Masaki Aya Nakae Ben Seymour ammation andusingthis ammatory arthritis. ammation orinjury. ammation (CFA ammation ammatory dis- ammatory neurogenic infl Overview ofourstudyprogramidentifyingthebrainresponseto Figure 1. infl tigue (indexedbytheanimalsburrowingbehavior)induced model ofpost-operativepain,andhasbeguntocharacterizethe ■ ■ ■ ■ Recent Publications 0006 (2016). thatbypasses conscious exposure.neural activity Nat.Hum.Behav. 1, M,LauH.FearKawato withoutfear reduction through reinforcement of A, ShibataK, Cortese Koizumi A,AmanoK, Yoshida W, B, Seymour 13209. doi:10.1038/ncomms13209 (2016). controlsbrain plasticity paininphantom limbpatients. Nat.Commun.7, Yokoi M, H,Hirata Yoshimine T, Kamitani Y, Saitoh Y. sensorimotor Induced Yanagisawa T, Fukuma H,Shimizu Kishima B, R,Seymour HosomiK, T, 208. doi:10.1038/mp.2016.81 (2016). inmajordepression.rupted Mol. habenulafunction Psychiatry 22(2),202- Lawson R,Nord B, C,Seymour Thomas D, Dayan P, Pilling S,RoiserJ. Dis- srep32509 (2016). come valueinParkinson's disease. Rep. Sci. 6,32509.doi:10.1038/ toulation ofthesubthalamicnucleusmodulates decisionout- sensitivity B,Seymour BarbeM,Dayan P, Shiner T, DolanR,Fink GR.Deepbrainstim- In otherwork,AyaNakaehasbeendevelopinganovelrodent ammatory signals. ammation. infl of theanteriorcingulatecortexassociatedwithfatigueinCFA Sequential imagesthroughtheratbrainshowingfunctionalconnectivity Figure 2. procedures. chronic painstatethatresultsfromnumerouscommonsurgical is tounderstandhowimmunologicalfactorsmaycontributethe of thepersistentpainstatethatresults.Avisionforthisresearch immunological, endocrinologicalandneurophysiologicalbasis ■ ■ ■ ■ ammatory arthritismodel. spinal cord. Mol. Pain 12,1-16(2016). mice viasuppression of injury-model striction TNF-α expression inthe (GJG) ameliorates allodyniainchronic con- Hagihara K.Go-sha-jinki-Gan A,Kishida M, Nakae Nakanishi Y, N,ShibataM, Sakashita BabaK, Yoshikawa H, adult population:amulticenter study. Pain. 157(5),1156-63(2016). Valls-Solé, J. Normative datafor heatevoked Aδcontact potentials in Granovsky Y, AnandP, A,Nascimento O, Nakae SmithB, Sprecher E, 10.1186/s12991-016-0095-1 (2016). study. disorder: acase-control Ann.Gen.Psychiatryspectrum 15,8.doi: Hagihira S, Takeda cognitive abnormalitiesofpaininautism M.Sensory Yasuda Y, H,OhiK, A,Kang HashimotoYamamori R, Nakae H,Fujimoto M, ing 2017. with Combined EEGandPhysiological Data.IEEEEMBSNeuralEngineer- Lancaster J, H,Callan Mano D, B. M,Seymour Kawato Decoding Acute Pain 53 Imaging 54 The MonteCarlomethodisappliedtothetravelingpulseand cells coupledwithreaction-diff is developedbasedonakineticchemotaxisequationformotile A MonteCarlosimulationmethodofchemotacticmotilecells due tothenoiseinfl ry. Alimitationoftheproposedmethodisthatitmightsuspend rophages intherealmousebrainandvisualizes3-Dtrajecto- the macrophageswell.Themethodsuccessfullytracksmac- artifi MR imagesandartifi and theHungarianalgorithm.Itwasvalidatedusingmousebrain numerically. Themethodisbasedonbackgroundsubtraction the methodto3-Dprocessingandevaluatedperformance method in11.7time-lapse2-DMRimages.Inthisyear,weextend year, weproposedanautomatedmethodbasedonHungarian algorithm Three dimensionalanalysisofmacrophagelearning-tracking A MonteCarlosimulationmethodforchemotacticmotilecells It iseff cially synthesizedimagesshowedthatthemethodtracked Information Systems ective toanalyze3-Ddynamics ofmacrophages.Last uence. cially synthesizedimages.Theresultswith usion equationsforchemicalcues. and extractionofdominantregionfromMDCTImages. sualizes them.Theresultsshowaclinicalabilityofvisualization and aregiondominatedbytheextractedarterialsubtreevi- method extractsanarterialsubtreeassociatedwithaseedpoint MDCT imagesasthebasisofaregionextractionalgorithm.The to extractthelungregiondominatedbyanarterialsubtreefrom ment eff arterial subtreewhichhasthrombi,physiciansfi nary arteries.Bymeasuringthesizeofaregiondominatedbyan one ofthelungdiseasescausedbythrombioccurringinpulmo- Region ExtractionAlgorithm cal foundationtotheMonteCarlomethod. tion problemofchemotacticbacteriaprovesasolidmathemati- Carlo simulationsandtheoreticalanalysisonthepatternforma- eling pulseofchemotacticbacteria.ComparisontheMonte Carlo methodcanreproducetheexperimentalresultsoftrav- pattern formationproblems.ItisdemonstratedthattheMonte Chronic thromboembolicpulmonaryhypertension(CTEPH)is ect point. Thisresearchproposesanautomatedmethod Yutaka Hata, Support Staff Support Assistant Professor Associate Professor Professor PhD 1 Manabu Nii Shinichiro Shima Shugo Yasuda Syoji Kobashi Yutaka Hata nd ahighertreat- diff of themouse.Linesaretrajectoriesmacrophages.Differentcolors 3-D trackingresultinthetime-lapseMRimages.(Dotsarebrainsurface Figure 1. ■ ■ Recent Publications erent macrophages). (2017). cation to thetraveling populationwave. J. Comput. Phys. 330,1022-1042 Yasuda Carlo S.Monte chemotaxismodel:anappli- simulationfor kinetic (preprint). model. arXiv:1703.08386 chemotaxis analysis ofakinetic Instability bacteria: tumble chemotactic Perthame B. and Yasuda pattern formation S.Self-organized ofrun-and- extracted regioncorrespondingtothepulmonaryartery). Extraction resultofdominantregion(reddenotespointedpulmonary arteryandpurpleregionisthe Figure 3. concentration ofchemicalcuealongthechannel. The insetshowsthesnapshotofpopulationdensitybacteriaand chemotactic bacteriumandtheircolorsrepresentthemovingdirection. chemotactic bacteriainmicrochannel.Theparticlesrepresenteach The snapshotofMonteCarlosimulationforthetravelingpulse Figure 2. ■ ■ Learning andCybernetics (2016). MRimages, Conf. Int. algorithmintime-lapse ing-tracking onMachine Tashita M,Mori A,Kobash,S,Nii Y, Yoshioka Y, &Hata Y. Macrophage learn- netics andMechanics, 52,479-486(2016). image, J. Int. Appl. from Electromag- MDCT domainregion extraction tery Yukawa A, KonoNishii T, N,KobashS,&Hata Kamiura Y. ar- Pulmonary 55 Informatics 56 RBPs, BCRs,TCRs,pathogens,etc.). as curateddatabasesforsequenceandstructuresofinterest(e.g., ment methods,bothaspubliclyavailablesoftware(MAFFT)and mune receptorsequencedata;and(3)multiplealign- of immuneresponses;(2)Functionalanalysislarge-scaleim- protein-RNA complexesinvolvedinpost-translationalregulation software targetingthreemainareas:(1)structuralmodelingof groups inotherareas,ourpriorityistodeliverstate-of-theart methods, andcollaboratewithawiderangeofexperimental munology. Althoughwestilldevelopgeneralbioinformatics eral bioinformaticstoseveralspecifi these issueswas to integrateinformationfrommultiple sequence noisiness ofsome RNAbindingsites.Ourinitialgoal toaddress due tothefl starting fromsequenceinformation. Thisapproachischallenging are workingonthepredictionof protein-nucleotidecomplexes RBPs tootherwithsimilardomains. Forthesereasons,we makes itdiffi chemical pathways.However,thelackofstructuralinformation important RBPsandRNAtargetsaff mediated responseindiff structural motif;however,eachoftheseproteinsshapestheIL6- and Roquineachtargetinterleukin-6(IL6)throughaconserved (e.g., infl responses, astheyeff RNA-binding proteins In 2016 Systems Immunology ammatory responses). Asanexample,Arid5a,Regnase-1 , thesystemsimmunologylabbegantoshiftfromgen- exibility ofsingle-strandednucleotides aswellthe cult togeneralizethemodes ofactionknown ect thestrengthanddurationofactivation erent ways.Therearelikelytobeother (RBPs)areessentialforproperimmune c topicsincomputationalim- ecting awide rangeofbio- tional interpretation oftheresultingBigSequence Dataisamajor and TCRsequences fromindividualdonorsonalarge scale.Func- proteomics-based technologies, it isnowpossibletocollectBCR receptor (BCR)andTcell (TCR)sequencingalongwith antigens. Withtheemergenceof novelhigh-throughputBcell past andpresentantigensaswell immuneresponsestothese tigen exposure.BandTcellrepertoires thuscontainanimprintof intrinsic andextrinsictothehost,includinggenetics,agean- in regulationofimmunereponses. to identifynewRBP-RNAinteractionsthatarelikelyplayarole els (manuscriptinpreparation).Withthesetoolsplaceweaim ate all-atomrepresentationsofthecoarse-graineddockedmod- limitation, wearenowdevelopinganextensionthatcangener- quired topredictbasespecifi sentation ofthenucleotidechainandthuslackresolutionre- Our fl laborative projects(Nyati,etal.,FukudaSakakibara,al.) tides. Thisstrategyhasalsopaidoff drive dockingsimulationsoffullyfl The nextgoalwastoutilizepredictedbindingpropensities experimental groups(Masudaetal.,Minoal,Yokogawaal). benchmarks aswellinsmall-scalecollaborativestudieswith and itscompanion,aaDNA,havebeenvalidatedinthird-party been largelysuccessfulandourresultingtool,aaRNA(Lietal), nucleotide bindingpropensitiesfornewRBPs.Thisstrategyhas alignments ofknownprotein-RNAcomplexesinordertopredict B andTcellrepertoires exible dockingsimulationsutilize acoarse-grainedrepre- Daron MStandley, Support Staff Support Research Assistant Postdoctoral Fellow Assistant Professor Associate Professor Professor areshapedbyvariousfactors,both cityofagivenRBP.Toaddressthis exible single-strandednucleo- , asevidencedbyrecentcol- 1 2 1 Li Songling Katoh Kazutaka Daron MStandley PhD based scoringfunction.Preliminarytestsindicatethatpairsof several newfeaturesinMAFFTandderivingaknowledge- modeling steptowithinfi mashita). In2016,wesucceededinspeedingupthestructural robust enoughtoidentifyhitsacrossmultipledonors(Shirai,Ya- effi sues, wehavedevelopedastructuralmodelingapproachthatis ty, specifi known abouthowsmallsequencevariationsaff over, evenifasignifi tated sequences,isunlikelytoyieldmanyfunctionalhits.More- tween donorsorbetweenadonorandreferencesetofanno- TCRs, comparingreceptorsconventiallyatthesequencelevelbe- of billionsuniqueBCRsandhundredsmillions the humanimmunesystemcantheoreticallyproducehundreds sequences withwhichtocomparethedata.Givenfactthat challenge, however,astherearefewwellannotatedBCRorTCR ■ ■ ■ cient enoughtohandlelarge-scalesequencedatasetsandyet Recent Publications (2016). clear antibodiesin acute systemic Submitted lupuserythematosus. S,etal. Clonalevolution andantigen-recognitionSakakibara ofanti-nu- veals criticalresidues for itsRNAassociation.Submitted (2016). Fukuda insightofAPOBEC3GN-terminal H,etal. domainre- Structural mRNA stabilizingprotein Arid5a. NucleicAcids Res. 45,2687-2703(2017). etal. K, Nyati TLR4-induced NF-κBandMAPK signaling regulate theIL-6 city oraffi nity toknownantigens.Toaddresstheseis- cantly similarsequenceisdetected,little B, FlexibledockingofssDNAtoSLEantibody(Sakakibaraetal). A. PutativebindingmodeofmouseAUF-1proteinwiththehomologymodel AUF-1(Nyatietal). Figure. ve secondsperstructurebyutilizing ect thesensitivi- large-scale analyses. larger numberofsequences,inrespondingtothedemands on anextensiontomakemoreaccurateoptionsapplicablea sequences forphylogenetictreeinference,etc.Weareworking mation ofnon-codingRNAsandproteins,interactiveselection Recent featuresinclude:Inclusionofsecondarystructuralinfor- ent optionsfornewlyemergingtypesofdataandanalyses. curacy, speedandutilityinpracticalsituations,providingdiff lease ofMAFFTin2002,wehavecontinuouslyimproveditsac- the mostpopularprogramsforbuildingMSAs.Sincefi comparative analysesofbiologicalsequences.MAFFTisone in preparation). likely thanrandompairstotargetthesameantigen(manuscript BCR orTCRmodelswithinthesamestructuralclusterarefarmore ■ ■ Multiple sequencealignment 1933-1942 (2016). the MAFFT multiplesequence alignment program. Bioinformatics 32, KandStandleyD.M.Katoh Asimplemethodto control over-alignment in chained guidetrees. Bioinformatics 32, 3246-3251(2016). alignment program to large data—reexamination oftheusefulness Yamada K.D, Tomii ApplicationoftheMAFFT sequence K. KandKatoh (MSA)isanimportantstepin rst re- er- 57 Informatics 58 data. derstand interferonregulationfrom suchtimelapseimaging computational algorithmstoextract importantquantitiestoun- upon antiviralresponses.Weare alsodevelopingautomated fl feron expressionundermicroscope iscombinedwithmultiple ways: timeandperturbation.TimelapseimagingoftypeIinter- are tryingtoincreasethe“dimension”ofobservationintwo quantitative monitoringofIFN-a6(Kumagaietal.2007).Nowwe have developedafl understanding thedynamicsofcomplexsystems.Previously,we Quantitative approaches specifi tem. Someoftheseprojectsaredescribedbelowtohighlight to getinsightintospecifi ets. Theseapproachesarecombinedinseveralprojectsthataim system’s dynamicsthroughtheanalysisofthesemassivedatas- opment ofmathematicalframeworkstounderstandtheimmune data” frommultiplesourcesintonetworkmodels,and(3)devel- measurement ofmoleculardynamics,(2)integration“big diff understand howtheimmunesystemworksbycombiningthree munology, bioinformaticsandtheoreticalphysics.Ouraimisto searchers withexpertiseindiff uorescent proteinknock-incells tomonitorgenesinduced Receptor molecule dynamicssuchasdimerization andcluster- Accurate quantifi The QuantitativeImmunologyResearchUnitisateamofre- erent butcloselyinterconnectedapproaches; (1)quantitative c topics. Quantitative Immunology cation ofbiologicalresponsesiscriticalfor uorescent proteinreporter systemforthe cquestionsrelatedtotheimmunesys- erent scientifi c fi elds includingim- et al.2015).Acommon featureofthesediseasesis thatinfl chronic obstructive pulmonarydisease(COPD)and silicosis(Diez to studythemechanismsbehind respiratorydiseases,suchas and generegulatoryimmunenetworks. Weapplythesemethods tion datafrompublicdatabases, to obtaininsightintosignaling ent experimentalconditions,and withprotein-proteininterac- transcription factorbindingwith transcriptome datafromdiff that revealstherelationbetweenbiologicalcomponents. es thatintegrate,summarizeandextractrelevantinformation brought biologyintothebigdataeraandneedforapproach- Data integration ing pathways. be usedtodescribethedynamicsofimmunesystem’ssignal- lecular densityconditions.Thishighlyquantitativetechniquecan dynamics ofsinglemoleculeswithoutbias,evenunderhighmo- level. Wedevelopedanovelalgorithmtoquantifythediff fully monitoredTLRsandtheiradaptorsatthesinglemolecular We areusingthistechniquetostudyTLRsignaling,andsuccess- (TIRFM) tomonitorthedynamicsofsingleimmunemolecules. ratories inIFReC,TotalInternalRefl we areapplying,incollaborationwithRIKENQBiCandotherlabo- for mostimmunesignalingpathways.Toaddressthisproblem, resolution, thedetailsofthisprocessarestillpoorlyunderstood signaling. However,becauseoflimitationsinbiochemicalassay ing withdownstreammoleculesisimportantforimmunesystem We aredevelopingmethodsthatintegratemeasurementsof The accumulationofhigh-throughput(“omics”)datasetshas Support Staff Support Research Assistant Postdoctoral Fellow Assistant Professor Associate Professor ection FluorescentMicroscopy 2 1 1 Yutaro Kumagai Diego Diez amma- usion er- the integrationofmassivedataopendoortoapproach fl study theregulatorypathwaysassociatedwithirreversiblein- ments oftranscriptionalactivityinamousemodelsilicosis,we ca forsilicosis).CombiningATAC-seqandRNA-seqmeasure- exposure totheharmfulcomponents(tobaccoforCOPDandsili- tion anddiseaseprogressionareirreversibleevenafterremoving Mathematical modeling validated. trolling RNAdegradationduringtheimmuneresponseisbeing 2016). Wealsoidentifi the RNA-bindingproteinsRoquinandRegnase-1(Kumagaietal. including thestem-loopstructurecontainingmotifsboundby quences ofeachcluster.Weidentifi and structuremotifidentifi terns withsimilardegradationkinetics.Then,denovosequence degradation kineticsbyclusteringthetimecoursedataintopat- after LPSstimulation.Transcriptomicdataisusedtodetermine rithm withRNAdegradationtimecourseinmousedendriticcells ■ ■ ■ ammation. Recent Publications Crit. Care Med. 190,981-988,doi:10.1164/rccm.201403-0421PP (2014). of chronic diseases. respiratory From basicsto application.Am.J. Respir. Diez D, Agusti Aand Wheelock CE.Network analysis intheinvestigation entifi synaptic pruningby inmicroglia. complement up-regulating factors Sci- JH,DiezDandHanayamaBahrini I,Song R.Neuronal exosomes facilitate 016-3325-7 (2016). motifsstructure inmRNAs. BMCGenomics17,1032,doi:10.1186/s12864- stimulation facilitates identifi patterns indendriticcells LPSmap ofRNAdegradation after kinetics Kumagai Y, Vandenbon A, Teraguchi SandSuzuki S,Akira Y. Genome-wide The accuratequantifi We areintegratinganoveldenovoRNAmotifsearchalgo- c reports 5,7989,doi:10.1038/srep07989 (2015). c reports ed novelmotifsforwhichtheirroleincon- cation ofimmunologicalresponsesand cation of primary sequence and secondary sequence andsecondary cation ofprimary cation isappliedtothe3’UTRse- ed knownstructuralmotifs, quantitative, integrativeandtheoreticalapproaches. The QuantitativeImmunologyResearchUnitcombines Figure. tective immunityandimmunetoleranceisachieved?”. main questionsinimmunology,“howthebalancebetweenpro- presentation. Thisframeworkallowsustoaddressoneofthe erogeneous behavioroftheindividualcellsinvolvedinantigen ic dynamicsoftheimmunesystemfromstochasticandhet- the immunesystem.Inthisframework,wederivemacroscop- munological research,wearedevelopingahierarchicalmodelof network dynamicsofimmunesignaling. dimensional data.Nowweareapplyingthissystemtoinferthe parameters ofthenetworkregulatorypathwaysfrommulti- developed asystemtoautomaticallyidentifythestructureand neous natureofcellpopulations(Teraguchietal.2011).Wehave which alsoallowsustorepresentthestochasticandheteroge- matical frameworkcalledStochasticBinaryModeling(SBM), eters. Tocircumventthisproblem,wehavedevelopedamathe- lular signalingisthebiochemicaldeterminationofmanyparam- description oftheimmunesystem. developing novelmathematicalframeworksforthequantitative immunology researchfromthetheoreticalperspective.Weare ■ ■ In anotherattempttoutilizethepowerofmathematicsinim- A commonobstacleforconstructingdynamicalmodelsofcel- 84, 062903(2011). equations.chemical reaction Phys. Rev. Matter Phys. E,Stat.Nonlin.Soft modelingofcells incontinuous timeasanalternativetic binary to bio- Teraguchi S,Kumagai Y, Vandenbon SandStandley DM.Stochas- A,Akira e1003323, doi:10.1371/journal.pcbi.1003323 (2013). duringtheinnate immuneresponse.activated PLoS Comp. Biol. 9, changes over timeinstimulated dendriticcells to identifygenenetworks Patil A,Kumagai Y, transcriptional LiangKC, Linking Suzuki K. Y andNakai 59 Units for Combined Research Fields 60 TSS-seq timeseries data. bioinformatics analysisofnewlyobtained ChIP-seq,RNA-seqand modifi charide (LPS)stimulation.Genome-wide patternsinhistone these eventsindendriticcells(DCs) inresponsetolipopolysac- nology ResearchUnit,wehave been studyingtheorderingof stood. course ofashort-termresponsetopathogensarenotwellunder- histone modifi ed. However,thecausalrelationshipsbetweendynamicsin modifi cations indendriticcells The controlofstimulus-induceddynamicsinhistonemodifi ects weareinvolvedin. approaches. Here,webriefl from limitedexperimentaldata,usingintegrativebioinformatics gies forextractingthemaximumamountofinformationpossible ics ResearchUnit,ouraimistoestablishandapplymethodolo- and betweengeneproductsisrequired.IntheImmuno-Genom- work, theinteractionsandregulatoryprinciplesbetweencells achieve thisgoal,acompleteunderstandingoftheimmunenet- causes ofdiseases,andtheirtreatmentinhumanpatients.To In collaborationwithDr.KumagaioftheQuantitativeImmu- The importanceoftranscriptionfactors(TFs)andepigenetic The ultimategoalofimmunologyishealthcare,elucidation cation changes andgeneinductionwerestudiedthrough cations inthecontrolofgeneexpressioniswidelyaccept- Immuno-Genomics cations, TFbindingandgeneinductioninthe y introducesomeoftheresearchproj- - man RNA-seqsamples, coveringmorethan60cell types. paring alargerdataset including>44,900mouseand >38,700hu- from 24celltypes261studies intotal.Wearecurrentlypre- from 191studies,and3,434mouse microarraysamples,obtained 4,639 humanmicroarraysamples, obtainedfrom19celltypes (Vandenbon etal.,PNAS,2016). The currentdatabaseincludes sion databaseforcellsofthehuman andmouseimmunesystem inference framework Establishment ofacomprehensivecelltype-specifi histone modifi the linkbetweenSTAT1/2bindingandestablishmentofthese Analysis ofTRIF,IRF3,andIFNRknock-outdatafurtherconfi tion ofH3K9K14ac,andwasfollowedbyincreasesinH3K4me3. fi of TFactivationandthetimingaccumulationhistonemodi- LPS-treated DCs,werevealedpotentiallinksbetweenthetiming of H3K4,andfi acetylation ofH3K9K14occursrapidly,followedbymethylation tional inductionofthepromoters.Forexample,LPS-induced within well-defi mulation ofseveralhistonemodifi data onlypartlysupportedthisview.Rather,LPS-inducedaccu- modifi cations. Especially,bindingbySTAT1/2 coincidedwithinduc- Last year,wepublishedImmuno-Navigator,ageneco-expres- Through integrativeanalysiswithbindingdatafor>20TFsin Although researchersoftenassumethatactivatinghistone cations areaprerequisiteforinducingtranscription,our Visiting Scientist Research Assistant Assistant Professor nally methylationofH3K36. cations. ned timewindows,independentofthetranscrip- cations atpromotersoccurred 1 1 Hiromasa Morikawa Alexis Vandenbon c network rmed eral aspectsofregulationtheimmunesystem. we areinvolvedinseveralinterdisciplinaryprojectscoveringsev- Interdisciplinary collaborativeresearchprojects Figure 1. between diff ferentiated celltypesaswellsimilaritiesinsignalingpathways leverages knownrelationshipsbetweenprogenitorcellsanddif- well asoverstimulus-inducedsignalingpathways.Themethod neously overextensivepartsofthediff methodology inwhichbiologicalnetworksareinferredsimulta- ■ ■ ■ Recent Publications Proc. Natl. Acad. USA.113,E2393–E2402(2016). Sci. database, reveals cell type-specifi Vandenbon, A.,etal. Immuno-Navigator, coexpression abatch-corrected motifsstructure inmRNAs. BMC Genomics 17,1032(2016). stimulation facilitates identifi patterns indendriticcells LPSwide mapofRNAdegradation after kinetics Kumagai, Y., Vandenbon, A., Teraguchi, S.&Suzuki, S.,Akira, Y. Genome- (2017). 18,173–183 establishment.Nat.Immunol. dependent super-enhancer Kitagawa, Y,. etal. Guidance ofregulatory T cell development by Satb1- In additiontoprojectsthataremostlybioinformatics-driven, At thesametime,wearedevelopinganewnetworkanalysis erent celltypes.Aconceptualschemeisshownin networks canfi knowledge ofcelltypediff expression data(1)isusedtoinferbiologicalnetworksusingdefault,naïve methodologies (2).Usingtheprior Conceptual fl Figure. cation of primary sequence and secondary sequence andsecondary cation ofprimary c genenetworks intheimmunesystem. ow ofourcomprehensivecelltype-specifi nally beusedfordownstreaminterdisciplinarycollaborations(4). erentiation lineagetreeas erentiation, ourmethodology improvestheinferrednetworks(3).Thesehighquality c networkinferenceframework.Alargeamountofgene cells (Ikebuchietal.,Scientifi ysis ofsingle-cellgeneexpressiondatasubsetsmigratoryT cells (Kitagawaetal.,NatureImmunology,2017),andtotheanal- lishing super-enhancersduringthedevelopmentofregulatoryT Chemistry, 2016),totheelucidationofroleSatb1inestab- irov etal.,AdvancesandApplicationsinBioinformatics (Mino etal., further analysisofpost-transcriptionalregulationbyRegnase-1 (Kumagai etal.,BMCGenomics,2016).Wealsocontributedtothe these forthepredictionofRNAsecondarystructure dation inresponsetoLPSstimulationofDCs,andemployed Unit wehaveanalyzedthegenome-widepatternsofRNAdegra- ■ ■ In collaborationwiththeQuantitativeImmunologyResearch 1058–1073 (2015). fl Mino, T., andRoquinRegulate aCommon etal. Element inIn- Regnase-1 (2016). /Gzmbinhibitsthecutaneousimmuneresponse.Il10 Rep. Sci. 6,35002 R.,etal. ArareIkebuchi, regulatory subsetofskin-tropic T cells expressing mRNAs Mechanisms. by Cell Spatiotemporally Distinct ammatory 161, Cell , 2015)andcirculatingserummicro-RNAs(Nos- c Reports,2016). 61 Units for Combined Research Fields IFReC's New PIs New Principal Investigators from FY2017 IFReC welcomed four new principal investigators with a new framework for governing structure in 2017. We expect them to add new impetus to IFReC. Laboratory of Stem Cell Biology and Developmental Immunology Laboratory of Molecular Immunology Additionally, we identifi ed a population of reticular cells ex- Mincle, MCL, Dectin-2 and DCAR are C-type lectin receptors that pressing CXCL12 at high levels, termed CXCL12-abundant reticular recognize M. tuberculosis. Our objective is to identify and study (CAR) cells within bone marrow (Immunity 2006) and indicated novel immunoreceptors and their ligands, in order to elucidate the that CAR cells are adipo-osteogenic progenitors and the major pro- mechanisms underlying ligand recognition as well as their poten- ducer of CXCL12 and SCF, creating the special microenvironment tial roles in immune disorders. Based on these results, we also aim (niche) for HSCs and B cells (Immunity 2010). Recently, we found to design new methods to orient the immune responses. that the transcription factor Foxc1 was preferentially expressed in To this end, our research centers on the following axes: CAR cells in the marrow, enhancing CXCL12 and SCF expression 1) Signifi cance and mechanisms of recognition of “aberrant self” or and was essential for inhibiting adipogenic processes in CAR cell “non-self pathogens” by C-type lectin family receptors. progenitors, and development and maintenance of niches for 2) Identifi cation of “self” through the TCR as well as its role in T cell HSCs and immune cells (Nature 2014). We are studying the roles of development. CXCL12-CXCR4 signaling and CAR cells in the spatiotemporal regu- 3) Study of newly discovered T cell subsets involved in autoim- lation of lymphohematopoiesis during homeostasis and diseases. mune diseases. Takashi Nagasawa, MD/PhD Sho Yamasaki, PhD We isolated a chemokine, CXCL12 (SDF-1/PBSF) as a molecule The immune system uses a wide range of receptors for the pro- that stimulates the growth of B cell precursors (PNAS 1994) and tection against pathogens and their clearance. Innate immune re- found that CXCL12 and its receptor CXCR4 are essential for coloni- ceptors elicit rapid responses, while a more complex one is trig- zation of bone marrow by hematopoietic stem cells (HSCs)(Nature gered by adaptive immune receptors. 1996; Immunity 2003), maintenance of a pool of HSCs in bone mar- The analysis of the C-type lectin family of innate immune recep- row (Immunity 2006), development of immune cells, including B tors has made signifi cant progress in the past decade. cells, pDCs and NK cells (Nat. Rev. Immunol. 2006), vascular forma- We have found that these receptors can sense both “damaged tion and cardiogenesis (Nature 1998). Conceptual Figure self” and “non-self pathogens”. Notably, our team showed that Laboratory of Statistical Immunology Laboratory of Molecular Neuroscience However, little is known regarding how to develop methodolo- cess from the perspective of the functional network of biological gy to integrate large-scale human genome data with diverse bio- systems. Further, we analyzed the mechanism by which the spatio- logical and immunological resources. The theme of our laboratory temporal dynamics in those biological systems control a series of is to develop such methods and apply to the latest large-scale dis- processes. ease genome and omics data. We have demonstrated that the dis- Particularly, the ultimate goal of this study was to elucidate the ease risk genes were signifi cantly enriched in overlap with the tar- control mechanism exerted by the associations among the ner- get genes of the drugs currently used for treatment of the diseases, vous system, immune system, and vascular system. Additionally, and could be promising resources of drug repositioning (e.g. we aimed to elucidate the principles involved in the operation of CDK4/6 inhibitors for rheumatoid arthritis, Okada Y et al. Nature living organisms with neural network disorders within the central 2015). Construction of the novel analytical methodologies of statis- nervous system by observing such disorders and their functional tical genetics, such as the HLA recovery process with respect to the dynamics of the entire bio- imputation method and the logical system and by conducting a comprehensive analysis of the miRNA network-based GWAS association between each system. analysis (Okada Y et al. Nat Genet 2015, Sci Rep 2016), is Yukinori Okada, MD/PhD also the focus of our labora- Toshihide Yamashita, MD/PhD Statistical genetics is a research fi eld that evaluates causality of tory. Through such transla- Disorders of the central nervous system, such as cerebrovascular human genetic variations on diseases, using statistical and bioin- tional approaches, we would diseases, cerebrospinal trauma, and encephalomyelitis, often formatics approaches. Recent developments of sequencing tech- like to empirically show the cause spatiotemporal changes in the nervous system and in vari- nologies have provided human genome data of hundreds of thou- value of statistical genetics to ous biological systems, such as the immune system and vascular sands of the subjects, and successfully identifi ed comprehensive dissect disease immunology system. In this study, we analyzed disorders of the neural networks catalogues of genetic susceptible loci of immune-related diseases. and novel drug discovery. in the central nervous system and the subsequent restoration pro- 62 63 Chugai Lab Joint Research Chair of Innovative Drug Discovery in Immunology Kunihiro Hattori (Visiting Professor) Ryusuke Omiya (Visiting Associate Professor) Junichi Hata (Visiting Researcher) As well as other researchers and technicians. Symposia & Seminars Members of Chugai Lab Drs. Kunihiro Hattori, Junichi Hata, and Ryusuke Omiya (left to right) The laboratory was set up on the basis of the comprehensive sive collaboration including performing joint feasibility studies collaboration agreement for cutting-edge research formed be- prior to advancing joint research. The synergy of IFReC’s recent tween Osaka University and Chugai Pharmaceutical Co., Ltd. research results with Chugai’s proprietary antibody engineering The Chugai researchers in this laboratory and IFReC research- technologies and molecular library etc. has high potential for the ers will promote close exchanges of information and comprehen- discovery of seeds for new drugs. Interview with Dr. Hattori I don’t know if it is the tradition of Osaka University largely in- nologies developed over many years at Chugai to collaborate fl uenced by Professor Kishimoto, but the researchers at IFReC are with and accelerate research as well as create a win-win relation- all thoroughly devoted to pure basic research. It may be that they ship in which we strategically acquire intellectual property and have not had the opportunity to view their research from the per- share knowhow on effi cient application methods and research spective of drug discovery. processes. I hope that this new laboratory, set up under the comprehen- Following the global success of the monoclonal antibody sive collaboration agreement between Chugai and Osaka Univer- Actemra® commercialized under the direction of Professor Kishi- sity, will act as a pathway to drug discovery based on IFReC’s moto, we seek second and third drugs from seeds originating cutting-edge research. from Osaka University and in the near future expect to expand Our job is to not simply receive experiment data from the uni- the laboratory located here to 20 researchers. versity and make drugs but is to utilize the drug discovery tech- 64 International Symposium on Advanced Immunology Nov. 1 Opening remarks Shojiro Nishio (President of Osaka University) Hiroo Imura (Former President of WPI Program) Shizuo Akira (Director of IFReC) Speaker Title Symposia & Seminars Shimon Sakaguchi Control of immune responses by regulatory T cells (Osaka University) Vijay K. Kuchroo Single cell genomics identifi es novel regulators of autoimmunity and (Harvard University) anti-tumor immunity Richard Flavell Non-coding RNAs in the regulation of infl ammation ( Yale University) Tasuku Honjo Cancer immunotherapy by PD-1 blockade (Kyoto University) Tak W. Mak Beyond Checkpoint Blockade: Emerging Strategies in Immunotherapy (Princess Margaret Cancer Centre, Canada) Atsushi Kumanogoh Involvement of semaphorins in pathogenesis of autoimmune, allergy and (Osaka University) infl ammatory diseases Josef S. Smolen Insights into pathways to infl ammatory rheumatic diseases - novel therapies and (Medical University of Vienna) response prediction Tadamitsu Kishimoto IL-6, Tocilizumab and Arid5a; Immunopathology and Therapy of Autoimmune Diseases (Osaka University) Nov. 2 Speaker Title Frederic Alt Recurrent DNA Break Cluster Genes in Neural Development, Diversifi cation and (Harvard University) Disease: Potential analogies to Lymphocyte Rearrangement Processes Tomohiro Kurosaki Fate decisions of germinal center B cells into the memory B cell or plasma cell (Osaka University) compartment Max Cooper Evolution of Lymphocyte Lineages (Emory University) Kiyoshi Takeda Regulation of intestinal homeostasis by epithelial barriers ■ Date : November 1-2, 2016 (Osaka University) ■ Venue : Conference Room 1003, 10th floor of Osaka International Gabriel Nuñez Role of the Microbiota and Immunity in the Control of Pathogens at the Intestinal Barrier Convention Center (University of Michigan) Tadatsugu Taniguchi th Recent advances in the regulation of infl ammation and its associated diseases This symposium was held for commemorating the 10 anniversary of IFReC and (University of Tokyo) the 77th anniversary of Prof. Tadamitsu Kishimoto’s birth. In the symposium, the Ruslan Medzhitov world’s leading scientists discussed current progress in elucidating immune reac- Infl ammation, Homeostasis and Disease (Yale University) tions and mechanisms that mediate and regulate immune responses. The partici- pants had an excellent opportunity to exchange information and ideas to accelerate Shizuo Akira Identifi cation of a novel monocyte subset involved in lung fi brosis and further the progress of immunology to the future discovery of new therapies for (Osaka University) immune diseases. Closing talk & remarks by Fritz Melchers 66 67 IFReC Seminars Date Speaker Affi liation Title 2016 MHC class I dependent education of NK cells: impli- Apr. 28 Klas Kärre Karolinska Institutet, Sweden cations for immunotherapy to tumors Self-assembling nanoparticle-based immunogen Jun. 17 Masaru Kanekiyo NIAID, NIH, USA design: mimicry of virus and beyond DNA damage and immune recognition of cancer Jul. 8 Stephan Gasser National University of Singapore cells Symposia & Seminars Center for Translational Medicine, Interleukin 10 in regulation of cardiac hypertrophy Sep. 27 Suresh Kumar Verma Temple University, USA and heart failure Transcriptional regulation of innate lymphoid cell Oct. 20 James Di Santo Institut Pasteur, France development and plasticity Role of the microbiota in the regulation of immunity Oct. 31 Gabriel Nunez University of Michigan School of Medicine, USA against systemic infections Impact of HIV and SIV on lymph node structure and Nov. 4 Richard A. Koup NIAID, NIH, USA function Towards the ultimate diagnostics: learning how to HudsonAlpha Institute for Biotechnology iCubate Nov. 15 Jian Han diagnose diseases from our immune system by NGS Inc., and iRepertoire Inc., USA of immune repertoire 2017 John Curtin School of Medical Research, A voyage into the discovery of the genetic Gaetan Burgio Australian National University mechanisms for host resistance to malaria Jan. 13 A voyage into the discovery of the genetic Richard Culleton NEKKEN, Nagasaki University, Japan mechanisms for malaria parasite resistance to hosts Intra-tumoral delivery of TLR agonists for cancer Jan. 26 Dennis Klinman The National Cancer Institute (NCI), NIH, USA therapy Modeling Zika virus in mice: what have we learned Jan. 27 Daniela Verthelyi The Food and Drug Administration (FDA), USA so far? European Molecular Biology Laboratory / Promoting tissue regeneration by modulating the Feb. 1 Mikael Martino Australian Regenerative Medicine Institute, immune system an example with bone regeneration Monash University, Australia Systems approaches to studying the immune Feb. 2 Anne O'Garra The Francis Crick Institute, UK response in tuberculosis: strategies to improve mouse models of human disease Feb. 3 Lewis L. Lanier University of California, San Francisco, USA Tracking the fate of memory Natural killer cells Exploring human intrahepatic Treg and IL-17 University of Birmingham Liver and Hepatobiliary Feb. 22 Ye Htun Oo secreting cells to develop translational Treg cell Unit, UK therapy Global Head, Roche Pharm Research & John C. Reed Bolstering immune responses Early Development Mar. 23 Global head, Immunology, David M. Lee Infl ammation & Infectious Diseases, Restoring immune homeostasis IFReC holds seminars throughout the year with speakers from a variety of disciplines Roche Pharma Research & Early Development including immunology, imaging and informatics with the aim of promoting collabora- School of Medicine Hospital, Marmara University, Haner Direskeneli Immunopathogenesis of Behcet's Disease tive research, as well as to inspire and educate the next generation of scientists. Istanbul, Turkey Since its establishment, IFReC has held more than a hundred seminars, which have Ehime University Proteo-Science Center and Mar. 24 Junya Masumoto Infl ammatory signaling in familial Behcet's Disease served as a forum for eff ective interaction between researchers beyond national borders Graduate School of Medicine, Japan and academic disciplines. This program has certainly contributed to IFReC’s mission of Aziz Sancar Institute of Experimental Medicine, Gunnur Deniz Natural Killer cells in Behcet's Disease promoting internationalization and interdisciplinary research. Istanbul University, Turkey 68 69 IFReC Colloquia Speaker Title Regulation of gut homeostasis by the ulcerative colitis-associated Kosuke Fujimoto (Mucosal Immunology) gene RNF186 Immuno-Navigator: a co-expression database for cell type-specifi c 25st Alexis Vandenbon (Immuno-Genomics Unit) network inference in the imune system Paroxysmal nocturnal hemoglobinuria caused by PIGT Yoshiko Murakami (Immunoglycobiology) mutations: Atypical PNH Some ATG proteins play a critical role in non-autophagic cell-autonomous Miwa Sasai (Immunoparasitology) immunity to vacuolar pathogens st Symposia & Seminars 26 Alison J. Hobro (Biophotonics) Multimodal label-free imaging of cellular responses to stress Takashi Satoh (Host Defense) NF-IL6 licenses pro-fi brotic monocyte diff erentiation from its progenitor Akiko Nakai (Immune Response Dynamics) A novel molecular mechanism of chemoattractant receptor signaling In vivo 3-D macrophage labeling and tracking method using time-lapse 27st Syoji Kobashi (Information Systems) MR images Michelle Sue Jann Lee (Malaria Immunology) Plasmodium infection modulates bone homeostasis David Millrine (Immune Regulation) The anti-infl ammatory properties of thalidomide Mohammad Mahabub-Uz-Zaman (Immune Regulation) Pathogenic role of Arid5a in septic shock 28rd Hironori Kawahara (Immune Network) Regulation of glial infl ammatory responses by neuronal exosomes A distinct subset of CD25 negative T-follicular regulatory cells localizes in James Wing (Experimental Immunology) germinal centers Tracking allergen-reactive IgE+ B cells in patients with eosinophilic Kazuya Takeda (Immune Regulation) chronic rhinosinusitis Newly developed animal post-operative persistent pain model and its Aya Nakae (Brain-Immune Interaction) behavior and pituitary changes after experimental pain in humans th 29 Dynamics of capsaicin-induced skin infl ammation and its infl uence on Masaki Maruyama (Brain-Immune Interaction) decision behaviors Tristan Nakagawa (Brain-Immune Interaction) The representation of capsaicin-induced tissue infl ammation in the brain ■ DateDate : 2255 th CollColloquium:oquium: AprilApril 20, 2016 2266 th Colloquium: June 8, 2016 Hiroki Kato (Nuclear Medicine) 11C-acetate PET in patients with multiple sclerosis: a pilot study 2277 th Colloquium: August 10, 20120166 28 th Colloquium: December 14, 2016 29 th Colloquium: February 8, 2017 ■ VenueVenue : Taniguchi Memorial Hall, Osaka University IFReC colloquia are a series of discussion meetings for IFReC members held once every other month since FY2011. At each colloquium, three speakers from IFReC laboratories give talks about their latest research progress followed by intensive discussion. After the colloquium, a small social gathering is held to further the discussions and encourage the exchange among IFReC members in an informal setting. These events serve as a platform to promote fusion researches and deepen understanding of researches conducted in IFReC. 70 71 Events The 6th NIF Winter School on Advanced Immunology Lecturer Title Shizuo Akira Functional diversity of macrophage/monocyte subsets revealed IFReC, Japan by gene targeting Veronique Angeli Events Homeostatic function of arterial macrophages National University of Singapore Burkhard Becher How T cells talk to myeloid cells in infl ammation University of Zurich, Switzerland Antonio Bertoletti Immunotherapy with T-cell receptor redirected T cell targeting viral Duke-NUS Medical School, Singapore antigen in viral related tumors Subhra Biswas Immuno-metabolic re-programming of monocytes/macrophages Singapore Immunology Network Ana Cumano Lymphocyte commitment during development Institut Pasteur, France Wendy Havran Immune crosstalk in the skin The Scripps Research Institute, USA Tomohiro Kurosaki Selection of germinal-center B cells into memory B cell or plasma IFReC, Japan cell compartment Paul Macary Analyzing the impact of antibody-mediated BloT5 neutralization on National University of Singapore airway hypersensitivity Laura Mackay Development and Function of Tissue-Resident Lymphocytes Doherty Institute, Australia Evan Newell High-dimensional cellular immune profi ling in health and disease Singapore Immunology Network Christiane Ruedl Intestinal dendritic cells Nanyang Technological University, Singapore Shimon Sakaguchi Control of immune responses by regulatory T cells IFReC, Japan Federica Sallusto Challenges and opportunities of human immunology Institute for Research in Biomedicine, Switzerland Kazuhiro Suzuki Control of lymphocyte traffi cking and adaptive immune responses IFReC, Japan by adrenergic nerves Filip Swirski Harvard Medical School and Unleashing the infl ammatory cascade in cardiovascular disease Massachusetts General Hospital, USA ■ Date : January 22-26, 2017 ■ Venue : Grand Copthorne Waterfront Hotel, Singapore The sixth Winter School on Advanced Immunology was jointly organized with Singapore Immunology Network (SIgN). Forty-four young researchers, who were competitively selected from 85 applicants and 16 world leading immunologists, got together in Singapore on 22-26 January 2017. Five young IFReC researchers participated in the school. The partici- pants shared intriguing insights and fi ndings in immunology, discussed new ideas and forged friendships that will fuel networking and future collaborations. (Visit the following website for details; http://ifrec-sign-winterschool.org) 74 75 Japanese Lessons Japanese language classes are held for overseas researchers / students to alleviate any stress and inconvenience in research or daily life that may be caused by the language barrier. Outreach Activities We off er two lecture-style classes, "Class A: Elementary to Pre-intermediate" and "Class B: Intermediate to Advanced". Class members are expected to learn verb and adjective conjugations to have basic knowledge of Japanese grammar in Class A , to learn grammar, vocabulary of intermediate/ advanced level, Kanji etc. to improve each member of skills in Class B. Comment by Ms. Kaori Tajima (Japanese instructor) Many researchers attend the classes because at IFReC Japanese is necessary when carrying out daily research activities in the laboratory. In order for the participants to start using Japanese sooner in communication with their colleagues, we learn words such as experiment, analysis, business trip, thesis/paper etc. at an early stage. Many of the participants eke out time for the classes from their full schedule so words such as busy, overtime, deadline, are often used in class. The participants fi nd the classes a welcome break from the laboratory and emphasis is placed on listening and speaking using pair and group work to encourage networking with researchers from other laboratories. At the end of a two year course, most communication in the class is in Japanese. This is the time when the participants can really start to enjoy their life in Japan by taking what they have learnt in the classes and using it to broaden their weekend activities. 76 Science Cafe Super Science High School Students Fair The series of science cafes is a long-lasting IFReC outreach IFReC science cafe was held in Osaka University Suita campus activity to promote communication among researchers and in FY2016. About 70 participants enjoyed a novel topic in im- the general public. It also enhances people’s understanding of munology in a relaxing atmosphere. immunology researches and the researchers involved in them. Outreach Activities ■ Date : August 10-11, 2016 ■ Venue : Kobe International Exhibition Hall ■ Host : MEXT, JST ■ Support : Boards of Education (Kobe Prefecture and Kobe City) Super Science High Schools (SSH) are selected high schools in Japan, which promote advanced math/science education as well as collaborative researches with universities, and activities to develop international perspectives. The SSH Student Fair FY2016 was held in Kobe and more than 200 schools, including several schools from overseas, held booths with posters to present their researches. WPI institutes held a collaborative booth and introduced the research activities of each insti- tute using posters, booklets and demonstrations. “Science Café on the Edge” at 2016 Icho Festival Students Visit ■ Date : May 1, 2016 Nara Prefecture super science high school, ■ Venue : Techno Alliance Building (Osaka University, Suita Campus) science tour ■ Date : August 18, 2016 ■ Guest : Etsushi Kuroda (Vaccine Science, IFReC) IFReC welcomed thirty students from Nara Prefectural Nara Senior High School and Unebi Senior High School. They heard a lecture by Prof. Kurosaki (Lympho- cyte Diff erentiation), and visited Akira Lab., Arase Lab. and Kishimoto Lab. to try some experiments and to talk with researchers. Osaka Prefectural Tennoji High School ■ Date : October 7, 2016 IFReC welcomed seventeen students from Osaka Prefectural Tennoji High School. They heard a short lecture from an IFReC Assistant Professor, Yutaro Kum- agai (Quantitative Immunology), introducing immunology and innate immunity and tried some experiments using nucleotide database. They also visited the Center for Information and Neural Networks (CiNet), and listened to IFReC Pro- fessor Yoshichika Yoshioka (Biofunctional Imaging) and Associate Professor Aya Nakae (Brain-Immune Interaction) explain their research. 78 79 The Forum Commemorating the 10th Anniversary of WPI AAAS 2017 Annual Meeting “Toward the future of Science in Japan” In celebration of the 10th anniversary of WPI program, a lecture ■ Date : December 17, 2016 meeting for general citizens was organized by MEXT, JSPS, and ■ Venue : Lecture Hall, 3F MEXT East Building WPI institutes. The nine leading researchers from all the WPI institutes were invited as the speakers, and the meeting was highly acclaimed by the audience. Greetings Outreach Activities Yasunao Seki (MEXT) 13:30－13:45 Yuichiro Anzai (JSPS) Hiroo Imura (Former WPI President) Keynote Lecture “Toward the free cross-border science” 13:45－14:00 Toshio Kuroki (WPI Program Director) Lectures Part 1 “Mathematics, Physics, and Earth Science” 14:00－14:20 Hitoshi Murayama (Director, Kavli-IPMU, University of Tokyo) 14:20－14:40 Shigeru Ida (Vice Director, WPI-ELSI, Tokyo Tech) 14:40－15:00 Motoko Kotani (Director, WPI-AIMR, Tohoku University) ■ Date : February 16-20, 2017 Lectures Part 2 “Life Science” ■ Venue : Hynes Convention Center (Boston, U.S.A) 15:20－15:40 Ryuichiro Kageyama (Vice Director, WPI-iCeMS, Kyoto University) The American Association for the Advancement of Science (AAAS) is the biggest international scientifi c society in the world 15:40－16:00 Masashi Yanagisawa (Director, WPI-IIIS, University of Tsukuba) and its mission is to “advance science and serve society”. The AAAS 2017 Annual Meeting was held in Boston and it off ered a 16:00－16:20 Shimon Sakaguchi (Vice Director, WPI-IFReC, Osaka University) broad range of activities including lectures, symposia, seminars and exhibits, with the theme of "Serving society through sci- ence policy ". Lectures Part 3 “Chemistry and Material Science” WPI institutes held a collaborative booth to introduce the WPI program and the institutes’ activities using posters, booklets 16:40－17:00 Masakazu Aono (Director, WPI-MANA, National Institute for Materials Science) and a demonstration experiment. More than 300 participants visited the booth and gained interest in the WPI program and world leading researches in Japan. 17:00－17:20 Kenichiro Itami (Director, WPI-ITbM, Nagoya University) 17:20－17:40 Seiji Ogo (Group Leader, WPI-I2CNER, Kyushu University) Closing “The future of Japan led by Science” 17:40－17:50 Ryoji Noyori (President of WPI Program, Nobel Laureate 2001) 17:50－18:30 Freewheeling discussion by audience and speakers 80 81 Research Projects Support Programs for Fusion Researches Papers in Fusion Research Fields Fusion Fields Other Fields % of Fusion Fields One of the missions of the WPI program is to generate novel a young postdoc, who obtained a Ph. D degree less than two research fi elds through fusion of existing research fi elds. Under years previous, is supervised by two mentors, whose specializa- this mission, IFReC has been committed to the creation of in- tion lies in different research fields, and is provided financial 250 30% novative immunology fi elds through integration with imaging support of ¥3 million a year for three years by IFReC. 28% and bioinformatics technologies for comprehensive under- These programs have eff ectively encouraged IFReC research- 26% 26% standing of immune dynamics. To facilitate fusion research, er’s interest in fusion research. By FY2016, a total of 27 projects 200 23% IFReC launched the Research Support Program for Combined had been selected for the program. Many interdisciplinary ap- 18% Research Fields in October 2009. This program provides fi- proaches were made and 36 papers successfully produced, con- 16% Research Projects nancial support for research proposals combining different tributing to the advancement of the fusion research at IFReC. 150 14% research areas. The project teams must consist of researchers The papers of fusion research published by IFReC members, in- from diff erent fi elds or backgrounds. The IFReC board of repre- cluding those produced from this program, accounted for more 143 131 100 106 sentatives screened the projects based on proposals and the than 30% of the total papers produced at IFReC in FY2015. This 111 126 projects selected are given ¥3 million a year for three years. demonstrates that our continued eff orts for the development 99 IFReC also started the Dual Mentor Program in October FY2012 of fusion research have come to fruition over the years of WPI 50 with the aim of helping young researchers engage in fusion program. 67 research at IFReC. In a selected project, a graduate student or 42 49 52 46 22 39 38 0 8 11 2008 2009 2010 2011 2012 2013 2014 2015 2013 － 2016 Project Leader Title Collaborators Visualizing information processing of immune cells via combination of Jun Kozuka, RIKEN QBiC Yutaro Kumagai fl uorescent reporter and single molecule imaging Shunsuke Teraguchi, IFReC Visualizing activation of germinal center B cells using genetically encoded Kazuhiro Suzuki Yoshihiro Baba, IFReC calcium indicators Development of an epigenome-based computational classifi cation system Alexis Vandenbon, IFReC Naganari Okura for the treatment of autoimmune diseases Shota Nakamura, RIMD Shimon Sakaguchi, IFReC Identifi cation of key factors for inducing functionally stable regulatory Alexis Vandenbon Hiromasa Morikawa, IFReC T cells Naganari Ohkura, IFReC Rikinari Hanayama, IFReC Evaluation Workshop The dynamics of novel signaling networks of macrophages exposed to Diego Diez Yutaro Kumagai, IFReC pathogens All the Combined Research Program and Dual Mentor Program projects are re- Shunsuke Teraguchi, IFReC quired to present their results once a year to IFReC members. At an annual Evalu- ation Workshop, IFReC PIs evaluate each project on the basis of specific criteria, such as “Achievement of the interdisciplinary research project mission “, Innovation of methodology” and “Prospects”. The evaluators have sometimes added severe comments to encourage improvement. Each project leader is given the evaluation result after the workshop. In the workshop, a comment sheet was provided to the audience and the comments from the audience were also given to the project lead- ers. In the 2016 workshop, fi ve project leaders of the Research Support Program pre- sented their research outcomes in front of IFReC members. The projects that gained a high score from IFReC PIs were awarded “Best Project Award.” Also, two of the past project leaders, selected by the director according to their outcome, reported on their outstanding interdisciplinary research achievements in the workshop. 84 85 Young Scientist Support Program for Research Abroad To strengthen our international research network and our ba- to develop the practical skills and abilities of young research- sis for international collaborative research, IFReC has provided ers in international collaborative research and to develop their fi nancial support to young researchers who wish to participate network with researchers overseas. Seven researchers used this in research activities at overseas institutions. The program aims support program in FY2016. Young Scientist Support Program for Research Abroad Name Country Conferences attended Kazuya Masuda USA Gene expression and signaling in the immune system Michelle Lee Sue Jann Australia International Congress of Immunology 2016 1. Institutes Seminar Series of the DRFZ Szandor Simmmons Germany 2. 46th Annual Meeting of the DGFI 3. Collaboration meetings with scientist of the Max Planck Institute Alexis Vandenbon Singapore 15th International Conference on Bioinformatics (InCoB2016) Data Kouyuki Hirayasu Italy 16th Annual Meeting of the Society for Natural Immunity Alison Jane Hobro USA SCIX conference Yutaro Kumagai Singapore 15th International Conference on Bioinformatics (InCoB2016) Since the start of this program, IFReC has provided support for 49 visits overseas by young researchers. Support provided for 43 overseas visits by young researchers in the past fi ve years Europe 15 America 20 Asia 4 Africa 1 Australia 3 IFReC has supported the active participation of young scientists in research activities overseas 86 Common Facilities Kishimoto Foundation Fellowship IFReC and its parent institution, the Research Institute for Mi- Animal Resource Center consists of three buildings for specifi c IFReC launched the Kishimoto Foundation Fellowship program The Kishimoto Foundation was established in 2008 in honor crobial Diseases (RIMD) are located on the same site, constitut- pathogen-free (SPF) animals and the live immuno-imaging for researchers in various fields of immunology in 2010. The of Tadamitsu Kishimoto, who, during the 1980s and 90s, elu- ing a large research complex. It contains the Core Instrumen- facility. With a large capacity animal-breeding facility in IFReC, program is supported by the Kishimoto Foundation and de- cidated the function of interleukin-6 (IL-6), a key molecule for tation Facility, the Animal Resource Center and the Network researchers are able to choose animal rooms suitable for their signed to support overseas researchers in order to promote and stimulating immune responses. He later developed the anti-IL6 Data Administration Offi ce, all of which are jointly operated by IFReC experiment purpose. develop immunological research and international exchanges receptor-based therapy, tocilizumab, to treat immune disorders and RIMD. The Core Instrumentation Facility is equipped with Using these common facilities, IFReC researchers are able to at IFReC. The fellowships are open to international postdoctoral such as Castleman's disease or rheumatoid arthritis. various highly advanced instruments and skilled technicians effectively and smoothly carry out their experiments to pro- researchers who seek to collaborate with IFReC researchers. The provide in-house services to IFReC and RIMD researchers. The mote their world-leading research at IFReC. recipients are provided with a salary and an airfare to Japan. IFReC–RIMD Research Complex at Suita Campus of Osaka University FY2016 Kishimoto Fellowship Recipients Position of recipient Name (initials) Nationality Host researcher Period Specially Appointed Researcher T. N. Germany Seymour Apr. 1, 2015 - Mar. 31, 2017 1 Specially Appointed Researcher P. L. Australia Coban Oct. 1, 2015 - Sep. 30, 2016 6 2 Specially Appointed Researcher D. H. Australia Sakaguchi Oct. 16, 2015 - Oct 15, 2016 3 5 Specially Appointed Researcher J. P. Vietnam Kishimoto Apr. 1, 2016 - Mar. 31, 2018 Quantitative Specially Appointed Researcher N.T. Slovenia Immunology Oct. 1, 2016 - Mar. 31, 2018 Research Unit 4 Photo : S. Higashiyama 1. IFReC Research Building 2. Integrated Life Science Building 3. Main Building, Research Institute for Microbial Diseases 4. South Building, Research Institute for Microbial Diseases 5. Cutting-edge Research Building for Infectious Diseases 6. Animal Resource Center for Infectious Diseases Animal Resource Center for Infectious Diseases Network Administration Offi ce ■ Specifi c pathogen-free (SPF) animal facility ■ Provision and maintenance of network infrastructure: ■ Sperm/ embryo freezing and preservation LAN system and servers (web, mail, mailing lists, etc.) ■ In vitro fertilization and embryo transplantation ■ Intracytoplasmic sperm injection Core Instrumentation Facility ■ Transgenic and knock-out animals ■ Basic and advanced instruments ■ Genome editing in experimental animals ■ In-house service ■ DNA sequencing, cell sorting, electron microscopy, mass Live Immuno-Imaging Facility spectrometry and next-generation DNA sequencing analysis ■ SPF animal experiment facility with 11.7T MRI, in-vivo imager ■ Radio isotope facility & two-photon microscope. ■ DNA chip center 88 89 Major Awards Shimon Sakaguchi The Crafoord Prize Five IFReC researchers Highly Cited Researchers 2016 Data Shimon Sakaguchi was awarded the Crafoord Prize Highly Cited Researchers are researchers with papers that Biology & Biochemistry in Polyarthritis 2017 for his discoveries relating to have a large number of citations from all over the world as • Shizuo Akira, Director of IFReC selected by Thomson Reuters. In 2016, some 3,000 researchers regulatory T cells, which counteract harmful immune Immunology from 21 fi elds were selected as being some of the most infl u- reactions in arthritis and other autoimmune diseases. • Shizuo Akira, Director of IFReC ential researchers in the world. The award ceremony was held at the Royal Swedish • Shimon Sakaguchi, Deputy Director of IFReC Six selections were from IFReC for a total of fi ve researchers Academy of Sciences. • Ken Ishii, Vaccine Science, IFReC (Director Akira were selected for two categories). Sakaguchi is the fourth Japanese prize winner, of • Kiyoshi Takeda, Mucosal Immunology, IFReC which three are currently or used to be IFReC re- • Masahiro Yamamoto, Immunoparasitology, IFReC searchers. Kiyoshi Takeda Erwin von Bälz Prize/Osaka Science Prize Shimon Sakaguchi, other laureates, and crown princess Victoria at the ceremony on May 18, 2017. Tadamitsu Kishimoto The King Faisal International Prize Kiyoshi Takeda (second left) at the award Tadamitsu Kishimoto was awarded the King Faisal ceremony of the Bälz Prize International Prize for Medicine 2017. The awarded (Photo : Katsumi Yanagiya) topic is “Biologic Therapeutics in Autoimmune Dis- eases”. The award ceremony was held at Riyadh, the Kingdom of Saudi Arabia. The King Faisal International Prize is launched by the King Faisal Foundation, and is an award includ- ing Science and Medicine. Many prize winners have gone on to receive other prestigious prizes, such as Kazuhiro Suzuki the Nobel Prize. Medical Research Encouragement Prize of the Japan Medical Association Astellas Awards for the Best Biomedical Research King Salman bin Abdulaziz Al-Saud of Saudi Arabia and Tadamitsu Kishimoto at the award ceremony on April 4, 2017. Kazuhiro Suzuki with the certifi cates of commendation for the prizes 90 91 Composition Finance Number of IFReC Staff Breakdown of total expenditure at IFReC in FY2016 Data 300 28 Administrative Staff 28 24 250 Research Support Staff 29 Individual Research Researchers 25 74 89 Project 82 28 63 28 25.7% Personnel 200 45 21 32.3% 77 Total Expenditure 150 31 74 15 3,423,108,667 173 183 186 100 23 171 (JPY) 135 179 13 127 115 50 3 89 Equipment Project 52 21.9% Activities 0 19.9% Mar.2008 Mar.2009 Mar.2010 Mar.2011 Mar.2012 Mar.2013 Mar.2014 Mar.2015 Mar.2016 Mar.2017 Travel 0.3% Number of Researchers Travel 200 26% 0.6% 30% 23% From Overseas 32% 31% Equipment Grant-in-Aid for Japanese From the 48 5.6% Scientifi c Research 55 42 University 150 20.3% 31% 54 54 20.8% 31% 29% Project WPI Grant Other Sources 42 Activities 40 1,241,101,000 2,182,007,667 100 27% 33 36.9% Contract (JPY) Research (JPY) Personnel Management 24 138 137 128 57.0% 19.9% Expenses Grant 117 119 15% 24.5% 50 8 93 87 82 Donation 65 14.4% 44 0 Mar.2008 Mar.2009 Mar.2010 Mar.2011 Mar.2012 Mar.2013 Mar.2014 Mar.2015 Mar.2016 Mar.2017 92 93 Selected Articles Two FOXP3+CD4+ T-cell subpopulations distinctly control the prognosis of colorectal cancers. Regulated selection of germinal center cells into the memory B cell compartment. Data Nat Med. 22:679-84 (2016). doi: 10.1038/nm.4086. Nat Immunol. 17:861-869 (2016). doi: 10.1038/ni.3460. Takuro Saito T, Nishikawa H, Wada H, Nagano Y, et al. Shinnakasu R, Inoue T, Kometani K, Moriyama S, et al. CD4+ T cells that express the forkhead box P3 (FOXP3) tran- whereas strategies that locally increase the population of FOX- How memory cells are selected and generated during germi- scription factor function as regulatory T (Treg) cells and hinder P3lo non-Treg cells could be used to suppress or prevent tumor nal-center (GC) reactions remains unclear. effective immune responses against cancer cells. Abundant formation. Tomohiro Kurosaki and his group found that light-zone (LZ) Treg cell infiltration into tumors is associated with poor clini- GC B cells with B cell antigen receptors (BCRs) of lower affi nity cal outcomes in various types of cancers. However, the role of were prone to enter the memory B cell pool. Mechanistically, Treg cells is controversial in colorectal cancers (CRCs), in which cells in this memory-prone fraction had higher expression of the FOXP3+ T cell infiltration indicated better prognosis in some transcriptional repressor Bach2 than that of their counterparts studies. with BCRs of higher affi nity.Haploinsuffi ciency of Bach2 resulted Shimon Sakaguchi and his group showed that CRCs, which in reduced generation of memory B cells, independently of sup- are commonly infiltrated by suppression-competent FOXP3hi pression of the gene encoding the transcription factor Blimp-1. Treg cells, can be classified into two types by the degree of Bach2 expression in GC cells was inversely correlated with the additional infi ltration of FOXP3lo nonsuppressive T cells. Func- strength of help provided by T cells. Thus, they propose an in- tionally distinct subpopulations of tumor-infi ltrating FOXP3+ T structive model in which weak help from T cells maintains rela- cells contribute in opposing ways to determining CRC progno- tively high expression of Bach2, which predisposes GC cells to sis. Depletion of FOXP3hi Treg cells from tumor tissues, which enter the memory pool. would augment antitumor immunity, could thus be used as an effective treatment strategy for CRCs and other cancers, Microbially cleaved immunoglobulins are sensed by the innate immune receptor LILRA2. Real-time intravital imaging of pH variation associated with osteoclast activity. Nat Microbiol. 1:16054 (2016). doi: 10.1038/nmicrobiol.2016.54. Nat Chem Biol. 12:579-585 (2016). doi: 10.1038/nchembio.2096. Hirayasu K, Saito F, Suenaga T, et al. Maeda H, Kowada T, Kikuta J, Furuya M, Shirazaki M, Mizukami S, et al Hisashi Arase group found that immunoglobulins disrupted Intravital imaging by two-photon excitation microscopy by microbial pathogens are specifi cally detected by leukocyte (TPEM) has been widely used to visualize cell functions. How- immunoglobulin-like receptor A2 (LILRA2), an orphan activat- ever, small molecular probes (SMPs), commonly used for cell ing receptor expressed on human myeloid cells. The microbially imaging, cannot be simply applied to intravital imaging be- cleaved immunoglobulins but not normal immunoglobulins cause of the challenge of delivering them into target tissues, as stimulated human neutrophils via LILRA2. In addition, stimula- well as their undesirable physicochemical properties for TPEM tion of primary monocytes via LILRA2 inhibited the growth of L. imaging. pneumophila. When mice were infected with L. pneumophila, Kazuya Kikuchi and Masaru Ishii groups designed and devel- immunoglobulins were cleaved and recognized by LILRA2. oped a functional SMP with an active-targeting moiety, higher More importantly, cleaved immunoglobulins were detected photostability, and a fluorescence switch and then imaged in patients with bacterial infections and stimulated LILRA2- target cell activity by injecting the SMP into living mice. The expressing cells. combination of the rationally designed SMP with a fl uorescent Their fi ndings demonstrate that LILRA2 is a type of innate im- protein as a reporter of cell localization enabled quantitation of mune receptor in the host immune system that detects immu- osteoclast activity and time-lapse imaging of its in vivo function noglobulin abnormalities caused by microbial pathogens. associated with changes in cell deformation and membrane fl uctuations. Real-time imaging revealed heterogenic behaviors of osteoclasts in vivo and provided insights into the mechanism of bone resorption. 94 95 Arid5a exacerbates IFN-γ-mediated septic shock by stabilizing T-bet mRNA. Identifi cation of an atypical monocyte and committed progenitor involved in fi brosis. Data Proc Natl Acad Sci USA. 113:11543-11548 (2016). Nature 541: 96-101(2017). doi : 10.1038/nature20611. Zaman MM, Masuda K, Nyati KK, Dubey PK, et al. Satoh T, Nakagawa K, Sugihara F, et al. Adenine-thymine (AT)-rich interactive domain containing pro- They conclude that Arid5a regulates the augmentation of IL-6 “Pulmonary fi brosis is one of a family of related diseases called tein 5a (Arid5a) is an RNA-binding protein that has been shown and IFN-γ in response to LPS, which possibly works synergisti- interstitial lung diseases. As the lung tissue becomes scarred, it to play an important immune regulatory function via the sta- cally for amplifi cation of various other cytokines that ultimately interferes with a person's ability to breathe. Most cases of pul- bilization of IL-6 and STAT3 mRNA. However, the role of Arid5a cause the development of septic shock in mice. monary fibrosis have no known cause. These cases are called in the overwhelming and uncontrolled immune response that idiopathic pulmonary fi brosis.” (American Lung Association) leads to septic shock is unknown. Monocytes and macrophages comprise a variety of subsets Tadamitsu Kishimoto group reported that Arid5a-deficient with diverse functions. It is thought that these cells play a crucial mice are highly resistant to lipopolysaccharide (LPS)-induced role in homeostasis of peripheral organs, key immunological endotoxic shock and secrete lower levels of major proinfl amma- processes and development of various diseases. Among these tory cytokines, including IFN-γ, IL-6, and TNF-α, than WT mice diseases, fi brosis is a life-threatening disease of unknown aetiol- in response to LPS. Their previous study suggests that Arid5a ogy. control the IL-6 level in vivo in response to LPS by stabilization Shizuo Akira group identifi ed a new type of macrophage, Seg- of IL-6 mRNA. They also observed that neutralization of IFN-γ regated nucleus Atypical Monocytes (SatM). They found SatM and IL-6 signifi cantly recovered the mice from endotoxic shock. have a bi-lobed segmented nuclear shape, and are ‘disorder- specific monocyte/macrophage subtypes’ corresponding to fi brosis. It may also now be possible to develop novel, more spe- cifi c therapeutic targets for this intractable disease in the future. Adrenergic control of the adaptive immune response by diurnal lymphocyte recirculation Inhaled fi ne particles induce alveolar macrophage death and interleukin-1α release to promote through lymph nodes. inducible bronchus-associated lymphoid tissue formation. J Exp Med. 213:2567-2574 (2016). Immunity 45:1299-1310 (2016). doi: 10.1016/j.immuni.2016.11.010. Suzuki K, Hayano Y, Nakai A, Furuta F, and Noda M, et al. Kuroda E, Ozasa K, Temizoz B, Ohata K, Koo CX, et al. It has long been proposed that various aspects of immune Particulate pollution is thought to function as an adjuvant responses are infl uenced by nervous system activity. However, that can induce allergic responses. However, the exact cell the cellular and molecular basis for neural regulation of immu- types and immunological factors that initiate the lung-specifi c nity are largely unclear. immune responses are unclear. Kazuhiro Suzuki group revealed that neural inputs to β2- Ken Ishii group found that upon intratracheal instillation, par- adrenergic receptors (β2ARs) expressed on lymphocytes gener- ticulates such as aluminum salts and silica killed alveolar mac- ate the diurnal variation in the frequency of lymphocyte egress rophages (AMs), which then released interleukin-1α (IL-1α) and from lymph nodes, which is refl ected in the magnitude of the caused inducible bronchus-associated lymphoid tissue (iBALT) adaptive immune response. During the period of high adren- formation in the lung. IL-1α release continued for up to 2 weeks ergic nerve activity, lymphocyte egress from LNs is restricted, after particulate exposure, and type-2 allergic immune re- which leads to an increase of lymphocyte numbers in LNs. Im- sponses were induced by the inhalation of antigen during IL-1α munization during the period of lymphocyte accumulation in release and iBALT formation, even long after particulate instilla- LNs promote adaptive immune responses. This diurnal variation tion. Recombinant IL-1α was suffi cient to induce iBALTs, which of lymphocyte trafficking may have evolved to maximize the coincided with subsequent immunoglobulin E responses, and effi ciency of host defense against pathogens. IL-1-receptor-deficient mice failed to induce iBALT formation. Therefore, the AM-IL-1α-iBALT axis might be a therapeutic tar- get for particulate-induced allergic infl ammation. 96 97 Publications Barman, S; Kayama, H; Okuzaki, D; Ogino, T; Osawa, H; Matsuno, Hamaoka, S; Naito, Y; Katoh, H; Shimizu, M; Kinoshita, M; Akiyama, Hubber, A; Kubori, T; Coban, C; Matsuzawa, T; Ogawa, M; Kawabata, Kakizuka, T; Ikezaki, K; Kaneshiro, J; Fujita, H; Watanabe, TM; 1 H; Mizushima, T; Mori, M; Nishimura, J; Takeda, K. Identifi cation K; Kainuma, A; Moriyama, K; Ishii, KJ; Sawa, T. Effi cacy compari- 27 T; Yoshimori, T; Nagai, H. Bacterial secretion system skews the 42 Ichimura, T. Simultaneous nano-tracking of multiple motor pro- of a human intestinal myeloid cell subset that regulates gut 14 son of adjuvants in PcrV vaccine against Pseudomonas aeru- fate of Legionella-containing vacuoles towards LC3-associated teins via spectral discrimination of quantum dots. Biomedical homeostasis. International Immunology 28, 533-545 (2016). ginosa pneumonia. Microbiology and Immunology 61, 57-74 phagocytosis. Scientifi c Reports 7, 44795 (2017). Optics Express 7, 2475-2493 (2016). (2017). 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Emoto, T; Yodoi, K; Matsumoto, T; Nakajima, K; Kita, T; Takeda, M; Mizoguchi, T; Hayashi, T; Sasaki, Y; Hatakeyama, M; Taguchi, K; Sugawara, R; Lee, EJ; Jang, MS; Jeun, EJ; Hong, CP; Kim, JH; Park, A; The articles were published between April, 2016 and March, 2017. The data were acquired using Web of Science Core CollectionTM on May 12, 121 Washio, K; Sakaguchi, S; Malissen, B; Nishigori, C; Hirata, K. UVB Yun, CH; Hong, SW; Kim, YM; Seoh, JY; Jung, Y; Surh, CD; Miyasaka, Exposure Prevents Atherosclerosis by Regulating Immunoin- 132 M; Yang, BG; Jang, MH. Small intestinal eosinophils regulate 2017, and sorted by alphabetical order of the fi rst authors. fl ammatory Responses. Arteriosclerosis Thrombosis and Vascu- Th17 cells by producing IL-1 receptor antagonist. Journal of Ex- lar Biology 37, 66-74 (2017). perimental Medicine 213, 555-567 (2016). 102 103 Access Map SUITA CAMPUS Osaka University ADDRESS 2Fl. 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