Whatistheimportanceoftheimmunologicalsynapse?DanielM.DavisandMichaelL. - PDF document

Whatistheimportanceoftheimmunologicalsynapse?DanielM.DavisandMichaelL.DustinDepartmentofBiologicalSciences,SirAlexanderFlemingBuilding,ImperialCollegeLondon,SouthKensingtonCampus,London,UK,SW72AZSkirballInstituteofBiomolecularMedicineandDepartmentofPathology,NewYorkUniversitySchoolofMedicine,NewYork,NY10016,USATheimmunologicalsynapse(IS)hasprovedtobeasti-mulatingconcept,particularlyinprovokingdiscussiononthesimilarityofintercellularcommunicationcontrol-lingdisparatebiologicalprocesses.RecentstudieshaveclariÞedsomeoftheunderlyingmolecularmechanisms Correspondingauthors:DanielM.Davis(d.davis@imperial.ac.uk),MichaelL.Dustin(dustin@saturn.med.nyu.edu).TREIMM200 TRENDSinImmunologyVol.notknownNo.notknownMonth0000 ARTICLEINPRESS1471-4906/$-seefrontmatter2004ElsevierLtd.Allrightsreserved.doi:10.1016/j.it.2004.03.007 SomeofthecheckpointsforaCTLmightalsobebypassedbyactivatingsignalsassociatedwithinßam-mationorstress,suchashighexpressionofICAM-1orinductionofNKG2Dligands.NKG2DisacostimulatoryreceptorforTcellsandanactivatingreceptorforNKcells[22,23]thatrecognizesstress-inducibleMHCclassI-relatedproteinsexpressedinspeciÞcmembranemicrodomainsains.Recently,NKG2D,alongwithICAM-1,wasfoundtocontributetoformationofanantigen-independentp-SMAC-likestructure,atleastattheISformedbetweenTcellsandasupportedlipidbilayercontainingICAM-1andagonistpeptideÐMHCMHC.Thepresenceofanantigen-independentp-SMAC-likestruc-turemightacceleratelaterstagesofCTL-mediatedkilling Box1.Varietyinthestructureofimmunologicalsynapses(ISs)fordifferenttypesofimmunecellinteractionsSupramolecularorganizationofproteinshasbeenobservedinsynapsesinvolvingvarioustypesofimmunecells,includingTcells,naturalkiller(NK)cells,Bcellsanddendriticcells.ThereisevidencethattheunderlyingmolecularmechanismsbywhichproteinsarearrangedattheISinvolvesbothspontaneousprocessesandcytoskeleton-drivenmechanisms.However,betweendifferentcelltypes,ithasrecentlyemergedthattherearemanyintriguingdifferencesinthespeciÞcorganizationofproteinsattheIS.Thus,asisperhapscommonplaceafteraninitialburstofresearchinanewarea,complexitiesnowsuggestthingsarenotquiteassimpleasperhapsonceassumed.InFigureI,twocellsareshowntointeractandbelow,onecellhasbeenremovedtorevealtheorganizationofproteinsattheintercellularcontactforvarioustypesofimmunecellinteraction.InthehelperT-cellIS,LFA-1(leukocytefunction-associatedmolecule-1)andtalininitiallyclusterinthecentralsupramolecularactivationcluster(c-SMAC)withT-cellreceptor(TCR)segregatedintoaperipheralring.Aftersometime,intheorderofminutes,thisarrangementinvertssothatTCRclustersinthec-SMACwithLFA-1andtalinintheperipheralSMAC(p-SMAC)[3,5].TCRclustersinsmallareassurroundedbyLFA-1intheearlycytotoxicT-cellIS[25].Aftersometime,aperipheralringofLFA-1formstoenclosetwosegregateddomainsinthec-SMAC,onecontainingtheTCRandtheothercontainingthelyticgranules[40,41].Studiesofraregeneticdiseaseshaverecentlyrevealedsomeofthemoleculardeterminantsunderlyingthemovementoflyticgranules[42].AtthecytolyticNK-cellIS,SH2-domain-containingphosphotyrosinephosphatase-1(SHP-1)initiallyclustersinsmallareassurroundedbyLFA-1[43].Atalatertime,lyticgranulesclusterinthec-SMACwithLFA-1inthep-SMAC[9,43,44].MHCclassIclusteringinthetargetcellatthenon-cytolyticNK-cellIShasseveraldifferentpatterns[8].IthasbeenspeculatedthatthisinhibitoryNK-cellISrepresentsastagetowardsassemblyofamatureactivatingsynapsefromwhichprogressionhasbeenprevented[11].Althoughnotdepicted,TcellsalsoformanISthatisjustbeginningtobestudied[45].ItisimportanttonotethatthedatarepresentedinFigureIderivefrommanyresearchersusingavarietyofparticularsystemsandmethod-ologies.Forexample,representationoftheearlycytotoxicT-cellISderivesfromdataexaminingtheinteractionofthecytotoxicTcellwithasupportedlipidbilayercontainingintercellularadhesionmolecule-1(ICAM-1)andpeptideÐMHCandremainstobeseenincytotoxicTcellÐtargetcellinteractions.Also,itshouldbenotedthattheorganizationofproteinsatanISwillprobablyvarywithlocalconcentrationsofcytokines,chemokinesandotherenvironmentalstimuli.AdetailedtabulationofthelocationofmoleculesattheT-cellandNK-cellIShasbeenpresentedinRef.[46],althoughitisrapidlyemergingthatmanyotherproteins,suchaspotassiumchannelproteins[47]andeventranscriptionfactorsignalingintermediates[48],canalsobeorganizedattheIS.Assuch,FigureIillustratesthatthecurrentdatasuggestthattherearedifferencesintheorganizationoftypesofISbutitremainsamajorchallengetotheÞeldtoclarifytheimportanceofthesedifferencesindifferentimmunecellinteractions.FigureI.Representationofthestructureoftheimmunologicalsynapse(IS)indifferenttypesofimmunecellinteractions.MoreresearchisneededtoclarifyifandhowintercellularcommunicationisinßuencedbysuchvariationsinproteinorganizationattheIS.Abbreviations:APC,antigen-presentingcell;LFA-1,leukocytefunction-associatedmolecule-1;NK,naturalkiller;SHP-1,SH2-domain-containingphosphotyrosinephosphatase-1;TCR,T-cellreceptor. TRENDS in Immunology Time LFA-1/talinLytic granulesHelper T cellCytolytic T cellCytolytic NK cellTarget cell in contact with a non-cytolytic NK cellEffector T cell or NK cell Lysis inhibited TREIMM200 TRENDSinImmunologyVol.notknownNo.notknownMonth0000 ARTICLEINPRESS ondetectionofagonistpeptide.Thus,thedeÞnitivenatureofthepeptideÐMHCfortargetidentiÞcationmightenableCTLstopasssomecheckpointsthatareneededtopromotespeciÞcityofNKcells.Self-assemblyofproteinclustersattheISSpeciÞcpatternsofMHC/KIRcanassembleataninhibitoryNK-cellISeveninthepresenceofdrugsthatinhibitcytoskeletalorATP-dependentprocessesprocesses.Thus,supramolecularorganizationofsomeproteinscanoccurbymechanismsotherthancytoskeletalorotherATP-dependentprocessesandperhapsmicrometer-sizeddomainscouldbecreatedbyspontaneoussegregationofreceptorsandligandsspanningsimilarlysizedintercel-lulardistances.ThethermodynamicsunderlyingthisideahavebeenmathematicallyformulatedbymodellingtheISasconsistingofapposingelasticmembranescontainingtwodifferentlysizedreceptor-ligandpairspairs.ItseemsthatthelossofentropybysegregationofproteinscanbeoffsetbythegaininenergyfromincreasedreceptorÐligandinteractionsandminimisingbendingoftheoppos-ingmembranes.Consistentwiththisidea,MHCproteinwasfoundtoaccumulateataninhibitoryNK-cellISpreferentiallywherethesizeofthesynapticcleftmatchedthesizeoftheextracellularportionsofKIR/MHCKIR/MHC.Indeed,theremightbesomethingfundamentallyimport-antabouttherelativesizeoftheextracellularportionsofMHC/KIRandMHC/TCRthatallowsATP-independentself-assemblyandexclusionoflargerreceptorsandligands,suchasICAM-1orLFA-1A-1.FunctionsoftheISAmatureT-cellISformsonlyonrecognitionofagonistpeptideÐMHCMHC,anddirectcomparisonofthetimingofproteinphosphorylationwithaccumulationattheISdemonstratedthatinitialmembraneproximalTCRsignalinghaslargelyabatedbeforeamatureISformsforms.Thus,amatureISisnotrequiredtoinitiateT-cell-cellbutappearstoformasearlyTCRsignalingiswaningwaningasaconsequenceofinitialsignals.However,onligationofT-cellNKG2DaringofLFA-1A-1,atleastattheISformedbetweenTcellsandasupportedlipidbilayer,andthussupramolecularorganizationofsomeproteinsmightoccurintheabsenceofagonistpeptideÐMHC.ForsometimeithasbeenknownthatT-cellcytotoxicitycouldbetriggeredbyagonistpeptideÐMHCataconcen-trationtoolowtotriggerotherresponses,suchasIFN-secretionandinternalizationofCD3CD3.ImagesoftheCTLISinthepresenceoflowandhighconcentrationsofagonistpeptideÐMHC,revealedthatCTLcytotoxicitycanbetriggeredwithoutsigniÞcantaccumulationofCD2orphosphotyrosineattheCTLÐtargetinterfaceinterface.Polar-izationofperforinandtubulin,however,wasalmostmaximalevenatalowconcentrationofagonistpeptideÐMHC.Thus,atleastsomeaspectsofamatureISareunnecessaryforcytotoxicity.AmajornextgoalwouldbetouncovermolecularmechanismsunderlyingdifferentthresholdsforthepolarizationofperforinandCD2.ClusteringofTCRinthecentralregionoftheISmightfunctiontobalancesignaling.Recently,aMonteCarlosimulationofT-cellactivationsuggestedthatenhancedTCRsignaling,causedbyclusteringintheIS,wouldbebalancedbyincreasedTCRdownregulationdownregulation.Insupport,TcellslackingCD2APareunabletodown-regulateTCRandarehypersensitivetoantigenantigen.Also,phosphotyrosinewassigniÞcantlyincreasedinthecentralregionoftheISformedbyTcellslackingCD2AP,consistentwithsimulationofanISintheabsenceofTCRdownregulation.Thiscollaborationofmodeling,geneticsandimagingallowedpreviouslydiscussedcon-ceptsregardingadaptivesignalingprocessesprocessestobeintegratedintothespatialframeworkoftheISwiththeconclusionthatc-SMACformationmighthaveanimport-antroleinT-celladaptationtodifferentlevelsandstrengthsofantigenreceptorsignaling.Manybiologicalsystemsdisplaysomedegreeofadaptivecontrolinsignalingprocesses,however,thismightbeaparticularlycrucialproblemfortheimmunesystemgiventhediversityinamountandqualityofantigenicstructures,and Table1.Theimmunologicalsynapse(IS)canhaveseveralfunctions,withvaryingimportanceforparticularcellcellinteractionsFunctionComment(I)EstablishingcheckpointsforlymphocyteactivationStagesintheassemblyoftheIS,havingdistinctcytoskeletalrequirements,canprovideaframeworkforestablishingcheckpointsforcellularactivation.Thismightbeparticularlyimportantforintegratingpositiveandnegativesignalsinnaturalkiller(NK)cellsinwhichaninhibitoryNK-cellISmightrepresentastagetowardstheassemblyofanactivatingISfromwhichprogressionhasbeenprevented.Spatio-temporalmovementsofCD45canfacilitatediscretestagesinT-cellsignaling.ling.(II)EnhancingsignalingByitsverynature,thedenseaccumulationofproteinattheIScanincreasetherateofT-cellreceptor(TCR)triggering,atleastinitially.initially.(III)Terminatingsignalingand/oreffectorMembrane-proximalsignalingcanbeterminatedbydownregulationanddegradationofTCRandalsoperhapsbyremovalofTCRligationattheISbyintercellulartransferofpeptideÐMHC.TransferofpeptideÐMHCfromanantigen-presentingcell(APC)toacytotoxicTlymphocyte(CTL)mightalsoterminateeffectorfunctionsbyfacilitatingÔfratricideÕ.IntercellulartransferofproteinsthatcommonlyoccursatanISmightalsoberequiredfordetachmentofconjugatedcells,althoughdirectevidenceforthisislacking.lacking.(IV)BalancingsignalingBalancingtherelativeinßuenceofenhancing(II)andterminating(III)signalingtomaintainagonist-triggeredsignalsinnormalTcells.OverbalancestowardterminatingsignalinginanergicTcells.cells.(V)DirectingsecretionThiswastheÞrstfunctionproposedandthereisnowconsiderableevidenceforthisbeingamajorfunctionoftheIS.CurrentresearchaddressesexactlywhensecretionofcytokineorlyticgranulesnecessitatesamatureIS.S.TREIMM200 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perhapsthisleadtotheevolutionofaspecialstructure,thec-SMAC,toassistthisprocess.Signal-balancingproper-tiesofthesynapsemightdampenresponsestothemostabundantorhigh-qualitypeptideÐMHC,allowingÔspaceÕforotherTcellstoalsoproliferate.Thismightbeimportantinallowingtheimmunesystemtoestablishabroadresponsetopathogens.CountingofTCR-deningthresholdsintheISThemomentumofstudiesontheISisfuelledbythedevelopmentofinnovativetechnologytomeetthechal-lengeofunderstandingtheextremesensitivityofTcells,abletodetectevenasingleMHCÐpeptidecomplexcomplex.BecauseTcellsappeartohavetheabilityofsinglemoleculedetection,therewasclearlyaneedtoadapttheemergingÞeldofsinglemoleculeimagingtothestudyofT-cellactivation.ThiswasrecentlyaccomplishedbytetheringphycoerythinmoleculestosingleMHCÐpeptidecomplexesonthesurfaceofAPCsAPCs.ThenumberofMHCÐpeptidecomplexesintheinterfacewithaTcellcouldthenbedeterminedatthemomentofT-cellinteraction,andtheconsequencesofthislevelofstimu-lationcouldbequantiÞedthroughanalysisoftheCasignal.ItwasfoundthatasingleMHCÐpeptidecomplextriggersatransientCasignalandthat10MHCÐpeptideinteractionscantriggerformationofastablesynapse.Similarultrasensitiveimagingmethodswillprobablyberequiredforunderstandingtheearlystepsinsignalingcascades,whichrapidlydropbelowdetectablelevelsdespitethecontinuedaccumulationofdownstreamsignal-ingintermediatesintermediates.ItisnotyetcleariforhowthespeciÞcsupramolecularstructureoftheIShasaroleindeÞningthresholdsattheIS,beyondtheimportanceofclusteringproteinsattheintercellularcontact.Thereisstillalottobelearnt,justbywatchingItisnottheendofexcitementabouttheIS,althoughthebasisoftheexcitementhaschangedfromtheinitialhypethatthesestructuresmightexplainT-cellactivationdecisionsunilaterally.Thepictureiscurrentlymorecomplexinthatproteinclusteringandlocalizationcanbeobservedinsituationsinwhichactivationisblockedandactivationcanbeobservedintheabsenceofapparentsupramolecularorganization.However,theexcitementremainsbecausereactionsattheseinterfacesarerequiredforimmunefunctionandnewtoolstohelpunderstandthemostfundamentalaspectsofthiscommunicationcontinuetomoveusforwardrapidly.ParticularlyforunderstandingT-cellactivation,wehaveidentiÞedthekeyreceptorsandligandsandthechallengenowistounderstandhowtheyactinconcerttogethertoregulateT-cellresponses.SimilarchallengesrelatetounderstandingNK-cellactivation,althoughsomekeyreceptorÐligandinteractions,suchasligandsfornaturalcytotoxicityreceptors,alsoneedtobeAsforsomanybiologicalprocesses,alotcanbelearntbysimplyimagingwhatgoeswhereandseeingwhenknowninteractionshappen.Acaveatisthatadeepunderstandingofthestrengthsandlimitationsofeachimagingapproachisnecessarytoattachreasonableinterpretationstotheseimages.Newimagingmethods,forexampletoprobelipidphasesphases,willalsobeneededtoadvanceourunderstandingoftheIS.Anothermajortechnicalchallengeistobeabletoobservemolecularrearrangementsfacilitatingintercellularcommunicationinvivo.Aninspiringrecentobservationisthatnano-tubularstructurescanallowintercellulartransportofmembranevesiclesvesicles.Aboutapplyingcuttingedgemicroscopytounderstandwormdevelopmentinthemid-1970s,NobellaureateJohnSulstoncommented:ÔNowtomyamazement,Icouldwatchthecellsdivide.ThoseNomarskiimagesofthewormarethemostbeautifulthingsimaginableÉInoneweekendIunraveledmostofthepostembryonicdevelopmentoftheventralcord,justbyby.AcknowledgementsWethankBebhinnTreanorandDavidBaconforhelpinthepreparationofFigureI.Weapologizetoscientistswhoseimportantworkwecouldnotcitebecauseofspacelimitations.ResearchinourlaboratoriesisfundedbytheMRC,theBBSRC,theHumanFrontiersScienceProgram,andtheNIH.1Norcross,M.A.(1984)AsynapticbasisforT-lymphocyteactivation.Ann.Immunol.(Paris)135D,113Ð1342Paul,W.E.andSeder,R.A.(1994)Lymphocyteresponsesand76,241Ð2513Monks,C.R.etal.(1998)Three-dimensionalsegregationofsupramo-lecularactivationclustersinTcells.395,82Ð864Dustin,M.L.etal.(1998)AnoveladaptorproteinorchestratesreceptorpatterningandcytoskeletalpolarityinT-cellcontacts.667Ð6775Grakoui,A.etal.(1999)Theimmunologicalsynapse:amolecularmachinecontrollingTcellactivation.Science285,221Ð2276Davis,D.M.etal.(1999)ThehumannaturalkillercellimmuneProc.Natl.Acad.Sci.U.S.A.96,15062Ð150677Batista,F.D.etal.(2001)Bcellsacquireantigenfromtargetcellsaftersynapseformation.411,489Ð4948Carlin,L.M.etal.(2001)IntercellulartransferandsupramolecularorganizationofhumanleukocyteantigenCatinhibitorynaturalkillercellimmunesynapses.J.Exp.Med.194,1507Ð15179Vyas,Y.M.etal.(2001)SpatialorganizationofsignaltransductionmoleculesintheNKcellimmunesynapsesduringMHCclassI-regulatednoncytolyticandcytolyticinteractions.J.Immunol.4358Ð436710McCann,F.E.etal.(2003)ThesizeofthesynapticcleftanddistinctdistributionsofÞlamentousactin,ezrin,CD43,andCD45atactivatingandinhibitoryhumanNKcellimmunesynapses.J.Immunol.2862Ð287011Davis,D.M.(2002)AssemblyoftheimmunologicalsynapseforTcellsandNKcells.TrendsImmunol.23,356Ð36312Igakura,T.etal.(2003)SpreadofHTLV-Ibetweenlymphocytesbyvirus-inducedpolarizationofthecytoskeleton.1713Ð171613McDonald,D.etal.(2003)RecruitmentofHIVandi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