whichnormallyretainNF- Bwithinthecyto-plasmofunstimulatedcells.Thepast5yearshavewitnessedtremendousadvancesinourun-derstandingofthisbranchoftheTNFsignalingnetwork.Especiallynoteworthywastheidenti-ficationofthemultiproteinI Bkinase(IKK)complexthatmediatesphosphorylationofI BinaTNF-dependentmanner(4).ThecoreoftheIKKcomplexconsistsoftwocatalyticsubunits,IKK andIKK ,andaregulatorysubunit,NF- Bessentialmodulator(NEMO,orIKK ).Inaddition,theIKKcomplexcontainsakinase-specificchaperoneconsistingofCdc37andHsp90thatplaysaroleinshuttlingthecomplexfromthecytoplasmtothemembrane.TheIKKcomplexisalsorecruitedtoTNF-R1,whereitbecomesactivatedwithinminutesofTNFtreat-ment.ThisactivationdependsonRIP,indicatingthattheIKKactivationwithinthereceptorcom-plexlikelyoccursthroughaRIP-dependentin-termediatefactor,perhapsakinase.Geneknock-outstudiesinmicehaveestablishedessentialrolesforIKK inTNF-inducedactivationofNF- B,andforNEMOinregulationofIKKcomplexactivationinresponsetonumerousup-streamsignals.Incontrast,IKK playsonlya

minorroleinTNF-inducedactivationofNF- B,butithasotherimportantfunctions,suchasservingasaNF- B2/p100kinaseinBcells.AninterestingfeatureoftheTNFsignalingnetworkistheexistenceofextensivecrosstalkbetweentheapoptosis,NF- B,andJNKsignal-ingpathwaysthatemanatefromTNF-R1.IntheabsenceofNF- Bactivity,cellularsusceptibil-itytoTNF-inducedapoptosisincreases,where-asenforcedactivationofNF- Bprotectsagainstapoptosis.Similarly,TNF-inducedJNKactivationisstrongerandmoreprolongedincellslackingNF- B,andtheproductsofseveralNF- B–activatedgenesinhibitactivationofJNKbyTNF.Moreover,NF- BactivationpromptstheresynthesisofI Bandotherinhib-itorymolecules,suchasthecIAPs,therebyadd-inganotherlayerofregulationofthedurationandamplitudeofTNFsignaling.

Todate,mostoftheplayersintheTNFpathwayhavebeenvalidatedbybothbiochem-icalandgeneticmeans,thusprovidingarichsourceofpotentialdrugtargetsforthedevelop-mentofanewgenerationofanti-inflammatoryagents.However,manyquestionsremainunan-swered.Forexample,whatMAPKKKinitiates

VIEWPOINT

thekinasecascadethatactivatesJNK,andhowisthiskinaserecruitedtoTNF-R1andactivatedwithinthereceptorcomplexinresponsetoTNF?InthecaseofIKKcomplexactivation,thepossibilityremainsthatanintermediatefactororkinaseisrequiredbetweenRIPandNEMO.Unravelingthemoleculardetailsofhowtheenzymeslikecaspase-8andtheIKKcomplexbecomeactivatedwithintheTNF-R1complexwillbekeytoafullunderstandingofthedynamicnatureofTNFsignaling.Finally,themolecularbasisforcrosstalkbetweenTNF-mediatedapoptosis,NF- B,andJNKsignalingpathwaysisnotwellunderstood.Decipheringthesepuzzleswillgreatlyhelpinterprethowaspecificout-comeofTNFsignalingisachievedindis-tinctbiologicalcontexts.

1.G.Chen,D.V.Goeddel,TNFPathway,Science’sSTKE(ConnectionsMap,asseenMay2002),www.1mpi.com.

2.R.M.Locksley,N.Killeen,M.J.Lenardo,Cell104,487(2001).

3.M.Feldmann,R.N.Maini,Annu.Rev.Immunol.19,163(2001).

4.S.Ghosh,M.Karin,Cell109,S81(2002).

References

TheFasSignalingPathway:MoreThanaParadigm

HaraldWajant*

Apoptosisandrelatedformsofcelldeathhavecentralimportanceindevelopment,homeostasis,tumorsurveillance,andthefunctionoftheimmunesystem.Apopto-sisisinitiatedbytwoprincipalpathways.Theintrinsicpathwayemergesfrommitochondria,whereastheextrinsicpathwayisactivatedbytheligationofdeathreceptors.ThisViewpointintroducesthebasicmechanismsoftheextrinsicpath-way,usingtheexampleoftheprototypicaldeathreceptorFasanditsroleinapoptosis,butitalsopointsouttheincreasinglyunderstoodimportanceofthisreceptorasanon-apoptoticsignaltransducer.

Fas(alsocalledApo-1orCD95)isadeathdomain–containingmemberofthetumornecro-sisfactorreceptor(TNFR)superfamily.Ithasacentralroleinthephysiologicalregulationofprogrammedcelldeathandhasbeenimplicatedinthepathogenesisofvariousmalignanciesanddiseasesoftheimmunesystem(1,2)[seeFasSignalingPathway,www.1mpi.com(3)andFasSignalingPath-wayinCardiomyocytes,www.1mpi.com(4)].AlthoughtheFasligand(FasL)–Fassystemhasbeenappreciatedmainlywithrespecttoitsdeath-inducingfunc-tion,italsotransducesproliferativeandactivat-ingsignalsthroughpathwaysthatarestillpoorlydefined(1,2).

InstituteofCellBiologyandImmunology,Allman-dring31,UniversityofStuttgart,70569Stuttgart,Germany.E-mail:harald.wajant@po.uni-stuttgart.de

Intheabsenceofmembrane-boundligand,inactivecomplexesofFasareformedbythepre–ligand-bindingassemblydomainofthemol-ecule(2).Interactionwithmembrane-boundFasL(oragonisticantibodies)reorganizesthesecomplexesandallowstheformationofadeath-inducingsignalingcomplex(DISC).TheFasDISCcontainstheadaptorproteinFas-associat-eddeathdomainprotein(FADD)andcaspases8and10,whichcaninitiatetheprocessofapopto-sis.FasL-inducedclusteringofFas,FADD,andcaspase-8or-10withintheDISCresultsinautoproteolyticprocessingofthesecaspasesbyinducedproximityandinreleaseofthepro-cessedactiveproteases(Fig.1).IntypeIcells,processedcaspase-8issufficienttodirectlyac-tivateothermembersofthecaspasefamily,whoseactionondefinedsubstratespavesthewaytotheexecutionphaseofapoptosis(1).IntypeIIcells,properactivationofeffector

caspasesbyFasdependsonanamplificationloopthatreliesoncaspase-8–mediatedcleavageofthepro-apoptoticBcl-2familymemberBidandsubsequentreleaseofmitochondrialpro-apoptoticfactors[forexample,cytochromecandsecondmitochondria-derivedactivatorofcaspases(SMAC,alsocalledDiablo)]todrivetheformationofthecaspase-9–activatingap-optosome.Activecaspase-9activatestheexe-cutionercaspase-3,whichinturnactivatescaspase-8outsidetheFasDISC,therebycom-pletingapositivefeedbackloop(1).

EachstepinFas-mediatedapoptosiscanbeatargetofregulatorymechanismsenablingcellstoshowflexibleresponsestostimulationbyFas.CorrespondingtothehierarchyofeventsinFas-mediatedapoptosis,theseregulatorymecha-nismscanbespecificforFasorcommontodeathreceptors,ortheycanaffecttheapoptoticcoremachineryofthecell.TheFasLgeneistranscriptionallyinactiveinmostcells.Thus,regulationofFasLexpressionitself,forexample,bythetranscriptionfactorsnuclearfactorkappaB(NF- B),activatingprotein1(AP1),ornucle-arfactorofactivatedTcells(NF-AT),regulatesFasL/Fas-mediatedeffects,suchasthoseofac-tivation-inducedcelldeathofCD4 Tcells(5).Toalesserextent,regulationofFasexpressionisalsousedtocontrolFasresponses,forexample,

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