structure of a spliceosome remodelled for exon ligation 2017 sebastian m. fica参考.pdf

该【structure of a spliceosome remodelled for exon ligation 2017 sebastian m. fica参考 】是由【小舍儿】上传分享，文档一共【20】页，该文档可以免费在线阅读，需要了解更多关于【structure of a spliceosome remodelled for exon ligation 2017 sebastian m. fica参考 】的内容，可以使用淘豆网的站内搜索功能，选择自己适合的文档，以下文字是截取该文章内的部分文字，如需要获得完整电子版，请下载此文档到您的设备，方便您编辑和打印。:..AcceleratedArticlePreviewLETTERdoi:,ChrisOubridge,,,Xiao-ChenBai,&KiyoshiNagaiThisisaPDFfileofapeer-,,:Fica,S.,://dx./(2017).ACCELERATEDReceived15November2016;.?2017MacmillanPublishersLimited,.:..LETTERdoi:,ChrisOubridge1,?,,Xiao-ChenBai1,&KiyoshiNagai1Thespliceosomeexcisesintronsfrompre-mRNAsintwosequentialtheU2/U6snRNAsexitingtheactivesite().TheHATrepeatstrans-esterifications–branchingandexonligation1–catalysedatofClf1andSyf1togetherformalargearchwhichisrotatedconsider-asinglecatalyticmetalsiteinU6snRNA2,*-terminalendofSyf1interactsrplex4,5withthecleaved5’-exonandwithU2snRNP,thereforethisrotationdisruptstheinterfacebetweenlariat—3’--specificfactorssuchasCwc25lockthebranchhelixConsequently,thePrp8RNaseHdomainrotatesinward()andthePrp17WD40domainmovesintothebodyofthePREVIEWthe5’-,().plex,nodensityistodestabilizebranching-?cryo-electronTheRNAstructureinthecoreofthespliceosomeremainsremark-haromycescerevisiaespliceosomeablyunchangedduringtheCtoC*transition,withtheexceptionofstalledafterPrp16-(,b).TheU2/U6catalytictriplexadoptstheplex,consistentwithbiochemicalandmontobothsteps,+ionsplexCandisisobservedadjacenttothephosphateoxygenligandsforcatalyticmetalstabilizedintoanewpositionbyPrp17,Cef1,andthereorientedionsM1andM2identifiedbymetalrescuestudies(),providingfurtherevidenceforasingleactivesiteforbothore,,the5’-exonisbase-pairedwithloop3’-exondocking,andrestructuresthepairingofthe5’-icanalysisandcross-,whichexperiments16,17,andthe3’OHofthelast5’-exonnucleotide(G-1)liespromoteexonligation,,boundtoPrp8inplaceofPrp16,couldinteract3’SS(,).Prp16-inducedremodellingresultswiththe3’-exon,suggestingapossiblebasisformRNAreleaseafterinadramaticrotationofthebranchhelixbyapproximately75degreesexonligation6,,ournewaroundthehingeatA30ofU2snRNA().TheBPmovesawayentre(approximately20?),’SStodockatthecatalyticMg2+site(,c).ThismovementThespliceosomeconvertsfrombranchingtoexonligationthroughdisruptsthenon-Watson-CrickinteractionsoftheBPadenosine(A70)theATP-dependentactivityofPrp16,’-spliceactiondestabilizesthebranchingfactorsCwc25andYju2(refs4,9)site(5’SS)withtheU6snRNAACAGAGAsequence(,e).TheandcreatesstrongbindingsitesfortheexonligationfactorsSlu7andA70baseispackedagainsttheriboseofthefirstintronnucleotidePrp18(refs10,11),whilepromoting3’-(+hiletheG(+1),w1)basestackswiththebaseofU(+2).-areessentialforsplicingofpre-mRNAswithlongdistancesbetweenplex,U(+formsabase-triplewithG37ofU2snRNAandC67ofthe2)thebranchpoint(BP)andthe3’-splicesite(3’SS)12andforcorrect3’SSintron4,plexU(+2)formsanon-canonicalbase-selection13,(,e),consistentwithcross-linkingTounderstandthemechanismofPrp16-,mutationsatbothU(+2)andremodelling,,,ingroupIIintronsthenucle-pre-mRNAsubstratecontainingadeoxy-guanosineatthe3’SSUAGotideequivalenttoA51(adjacenttothetwonucleotidesinvolvedinsequenceandpurifiedthemviaanaffinity-tagonSlu7(Methods).Withtriplexformation)base-pairswiththelastnucleotideoftheintronthissubstratethespliceosomesstallafterPrp16-dependentremodel-(γ-γ’interaction)(+2)mayinteractwithlingandbeforeexonligation,plex14;thuspurifiedthelastnucleotideG(-1)-intermediate(Extendedpositions(A70,G(+1),U(+2)/A51),).Weobtaineda3.-stepdefects,arealignedtowardstheactivesite,stronglysuggestingaplex(ExtendedDataFigs1,2)ponentspathforthe3’SS().Finally,theHoogsteenbase-pairbetween(ExtendedDataTables1,2andExtendedDataFigs3,4).A(oftheintronandG50ofU6snRNA()nolongerforms+3)ACCELERATEDPrp8,Snu114andtheU5SmcoredomainformthefootdomaininC*(),icevidencethatthisinteractioninbothC*plexes(),,montobothCandC*-terminalendofClf1isanchoredbyCef1,Syf2andcore(U6snRNAISLandhelicesIaandIb)ontoPrp8,whereasthe1MRCLaboratoryofMolecularBiology,FrancisCrickAvenue,CambridgeCB20QH,UnitedKingdom.?Presentaddress:EMBLGRENOBLE,71avenuedesMartyrs,CS90181,38042GrenobleCedex9,|VOL000|NATURE|1?2017MacmillanPublishersLimited,.:..RESEARCHLETTERbranchhelixrotatessubstantiallybetweenthetwostates().paredtothecrystalstructureofthehomologousCcomplex,thebranchhelixislockedintothebranchingconforma-Prp43ATPaseintheADP-Mg-boundform29,plextionpredominantlybyCwc25,Yju2andIsy1,suchthatA70isinsertedisinanopenconformationwithawiderseparationoftheRecA1andentre()4,,Prp16promotesRecA2domains().DensityattributabletoRNAispresentinthedissociationofCwc25,Yju2andIsy1andbindingofPrp18,Slu7andPrp22activesite()andPrp22cross-links17nucleotidesdown--,entreofthespliceosomeandPrp22inourstructurecanbewhenPrp17wedgesbetweentheU2SmringandthePrp8RNaseHspannedwith16-’-exondomain(;).plex,theRNaseHemergingfromthecoretopromotemRNAreleaseanddissociationofdomainofPrp8isrotatedbyabout80°withrespecttotheLargeSlu7,(,e).Inthisorientationtheβ-fingerofOurC*mplexstructureelucidatesthestructuralconsequencescorooveofthebranchhelixandofPrp16activity(),revealsalargerotationofthebranchhelixreachesCef1(,d).Prp17bindsacrosstheinterfacebetweenthewhichcreatesaspacefor3’--f-helixbridgesexonligationconformationofthespliceosome,andprovidesastruc-thePrp8RNaseHdomainandtheCef1Mybdomainandreachestheturalframeworkforinvestigating3’splicesiteselectionandligatedC-terminusofSyf1().,alongwithanyadditionalExtendedDatadisplayitemsandrotatedpositionoftheRNaseHdomainobservedinC*wouldclashSourceData,areavailableintheonlineversionofthepaper;,explaininghowPrp16-dependentdissociationofIsy1enablestheRNaseHdomaintorotatepromotingtheexon-Received15November2016;,,.,Will,.&Lührmann,:DesignprinciplesofaWthebranchhelixbindingface().,701–718(2009).plex,,-,229–234(2013).interactswithPrp18,andlatchestheRNaseHdomainontotheendo-,.&Guthrie,-pairinginteractionbetweenU2andnucleasedomainofPrp8,thusstabilizingtherotatedconformationofU6snRNAssuggestsamechanismforthecatalyticactivationofthetheRNaseHdomainandthebindingofPrp18inC*(,803–817(1992).,-).,197–201(2016).,R.,Yan,C.,Bai,R.,Huang,G.&Shi,-?,895–904(2016).,’-exonforPrp22-,743–754(2008).,.,Blanco,.,Walter,.&Staley,’-exonatthecatalyticmetalbind-DEAH-BoxATPasesRemodelPre-,985–998(2016).,B.&Guthrie,-dependentATPasethatinteractsACAGAGAsequenceandthe5’--,494–499(1991).,.,Liu,.&Cheng,-boxATPasePrp16hasdualrolesincouldpositionthe3’,145–154(2010).,S.-A.,Turner,W.&Schwer,().Ifsecondstepofyeastpre-,1068–1077(2002).thisisthecase,itispossiblethatthepenultimatenucleotideA(-2),-mRNAitsprecedingnucleotideY(-3),902–915(2013).,A.&Schwer,-mRNAsplicingisnucleotideG(+1)andthebranchpointadenosine(A70).Aninterac-dictatedbythedistancebetweenthebranchpointandthe3',tionbetweenthe3’SSUAGsequence,theACAGAGAsequenceandthe707–717(1996).5’SSintronsequenceforexon-,K.&Reed,'splice-,207–210(1999).However,mutationalstudiesofthe5’SSand3’,.&Sharp,-specificmodificationofpre-mRNA:thebase-pairingscheme19,25suggestingthattheseinteractionsmayinvolve2'-,992–997(1992).non-canonicalbase--pairingbetweenthe5’-,.,Mefford,.,irilli,.&Staley,-,loop1ofU5snRNA()placesthe3’OHgroupofthe5’-exonclose464–471(2014).,.&Norman,'andatthe3’SSisboundtoM1andM2(ExtendedDataFigure6;refs2,26).3',743–754(1992).,.&Steitz,,1989–1996(1993).betw