RESEARCHARTICLESUMMARY◥SIGNALTRANSDUCTIONInvivobrainGPCRsignalingelucidatedbyphosphoproteomicsJeffreyJ.Liu,KirtiSharma,LucaZangrandi,ChongguangChen,SeanJ.Humphrey,Yi-TingChiu,MarianaSpetea,Lee-YuanLiu-Chen,ChristophSchwarzer*,MatthiasMann*INTRODUCTION:TheGprotein–coupledre-ceptor(GPCR)superfamilyisamajordrugtargetforneurologicaldiseases.FunctionallyselectiveagonistsactivateGPCRs,suchasthekappaopioidreceptor(KOR),inapathway-specificmannerthatmayleadtodrugswithfewersideeffects.Forexample,KORagoniststhattriggerbeneficialantinociceptive,anti-pruritic,andanticonvulsanteffectswhilecausingminimalornoundesirabledysphoric,aversive,orpsychotomimeticeffectswouldbeinvaluableinlightofthecurrentopioidepidemic.How-ever,functionalselectivityobservedinvitrofrequentlyhaslittlepredictivevalueforbehav-ioraloutcomes.RATIONALE:ObtainingasystemsviewofGPCRsignalinginthebrainmayovercomethegapbetweeninvitropharmacologyandinvivotesting.Recentbreakthroughsinmassspectrometry–basedproteomicshaveenabledustoquantifytensofthousandsofphosphoryl-ationeventssimultaneouslyinahigh-throughputfashion.UsingtheKORasaGPCRmodel,weappliedthistechnologytoachieveaglobalover-viewofthearchitectureofbrainphosphopro-teomeinfivemousebrainregions,inwhichweexaminedsignalinginducedbystructurallyandbehaviorallydiverseagonists.RESULTS:Throughthequantificationof50,000differentphosphosites,thisapproachyieldedabrainregion–specificsystemsviewofthephosphoproteome,providingacontexttoun-derstandKORsignalinginvivo.WeobservedstrongregionalspecificityofKORsignalingattributabletodifferencesinprotein-proteininteractionnetworks,neuronalcontacts,andthedifferenttissuesinneuronalcircuitries.Agonistswithdistinctsignalingprofileseliciteddifferentialdynamicphosphorylationofsynap-ticproteins,therebylinkingGPCRsignalingtothemodulationofbrainfunctions.Themostprominentchangesoccurredonsynapticpro-teinsassociatedwithdopaminergic,glutama-tergic,andg-aminobutyricacid–mediated(GABAergic)signalingandsynapticvesiclerelease.Thelarge-scaledepho...
