自噬与核糖体的关系

known(PowersandWalter1999;Kuruvillaetal.2001;Gelperinetal.2002;Ramirez-Valleetal.2008).WefoundthattheturnoverofeIF4GIandeRF3ishighlyselectiveduringnitrogenstarvation,sincenodecreaseinabundanceoftheinitiationfactorseIF2αandeIF5B,ortheelongationfactoreEF3wereobservedundertheseconditions.Previousstudiesfoundthattwootherinitiationfactors,eIF4EandeIF4A,alsoremainstableinbothyeastandmammaliancellsfollowingnutrientlimitationorrapamycinaddition(Bersetetal.1998;PowersandWalter1999;Kuruvillaetal.2001;Ramirez-Valleetal.2008).WhilethedegradationofeIF4GIandeRF3sharessomecomponentsoftheribophagymachinery,theturnoverrateoftheseproteinsismuchfasterthanribophagy,andtheoverallextentofproteinturnoverisalsomuchgreater.Theseresultsindicatethatselectiveautophagyofspecifictranslationfactorsoccursfollowingtheonsetofnitrogenstarvationinamannerthatcomple-mentstheribophagyprocess.

ThebalancebetweenmRNAsynthesisanddegradationcontrolsthesteady-statelevelsofcellularmRNAs.RegulationofmRNAsynthesisrateshaslongbeenappreciatedasakeymechanismthatcontrolsgeneexpression,whiletheimpor-tanceofregulatedmRNAdecayisgenerallylessappreciated.WefoundthatthedegradationofDcp2p(anmRNAdecap-pingfactor)andPop2p(acomponentoftheCCR4/Pop2pmRNAdeadenylasecomplex)duringnitrogenstarvationwasextremelyrapidanddependentuponproteasomefunc-tion(Fig.3)andtheribophagydeubiquitinase,Ubp3p(Fig.4).Similarly,apreviousstudyfoundthatUbp3pinteractswiththeproteasometostimulatetheselectivedegradationofaprotein(Rad4p)tosuppressnucleotideexcisionrepair(MaoandSmerdon2010).Thedegradationweobservedwasselective,asotherproteinsinvolvedinmRNAdecay,suchasTpa1pandLsm1p,werestableunderthesamecon-ditions(Fig.1B).Twootherproteins,thepoly(A)bindingproteinPab1panddecappingenhancer,Pat1p,werealsode-pletedbyautophagyatdifferentrates(Fig.1C–E).Whentak-entogether,theseresultsindicatethatspecifictranslationandRNAmetabolismfactorsareselectivelydegradedbyeithertheproteasomeorautophagyduringnitrogenstarvation.Itispossiblethatthedifferentialdegradationoftheseproteinshelpscellsaltertheirgeneexpressionprofiletoadapttostar-vationconditions.

PreviousstudiesfoundthatdepletionofeithereIF4GIoreRF3reducesgrowthratesandinhibitsTORsignalinginmammaliancells(Chauvinetal.2007;Ramirez-Valleetal.2008).Thesesimilareffectssuggestthattheseproteinsmayshareacommonfunction,suchasasharedroleintranslation.Inthisregard,botheIF4GIandeRF3participateintheforma-tionofmRNAclosed-loopstructuresrequiredforefficientinitiationofcap-dependenttranslation(Amranietal.2008).BasedonourfindingthatthedeubiquitinaseUbp3panditsbindingpartnerBre5parerequiredforefficientdegradationofeIF4GIandeRF3(Fig.4A,B),theautophagymachineryandtheUbp3pcomplexmayfacilitateeRF3andeIF4GIdeg-Selectiveproteindepletionduringnitrogenstarvation

radationinordertoreprogramtranslationinitiationduringadaptationtonitrogenstarvationconditions.

Previousstudieshaveshownthatselectiveautophagyexistsinorganismsrangingfromyeasttomammals.Targetsofthisprocesscanrangefromspecificproteinstoproteinaggre-gates,ribosomes,variousorganelles,andbacteria.Whilethemechanismofselectiveautophagyremainspoorlyunder-stood,aroleforubiquitinisacommontheme.Forexample,degradationofvarioussubstratesbyautophagyinmammali-ancellsrequiresubiquitination,aswellasubiquitin-bindingreceptorssuchasp62(Kraftetal.2010).Similarly,theselec-tivedegradationofribosomesfollowingnitrogenstarvationrequiresthedeubiquitinaseactivityofUbp3panditspartner,Bre5p(Kraftetal.2008).Recently,theUbp3p/Bre5pcom-plexwasshowntointeractwithbothCdc48p,whichplaysaroleintheproteasomalpathway,aswellasUfd3p,aubiq-uitin-bindingadaptorofCdc48(Ossareh-Nazarietal.2010).TheseUbp3ppartnersarerequiredforefficientribophagybutarenotrequiredforbulkautophagy.Sinceinhibitionofproteasomalactivitydoesnotblockribophagy,itwaspro-posedthatCdc48pfunctionsasamolecularplatformonwhichubiquitinatedribosomesarerecognizedanddeubiqui-tinatedbeforedeliverytothevacuolarfordegradation.

Inthecurrentstudy,wefoundthatUbp3pitselfisdegradedduringnitrogenstarvation.Thereducedsteady-stateabun-danceoffull-lengthUbp3pisaccompaniedbytheaccumula-tionofacarboxy-terminalfragmentthatcorrespondstotheUCHcatalyticdomainofUbp3pthatlackstheBre5pbindingdomain(Fig.6;Lietal.2007).Thelossoffull-lengthUbp3pandappearanceoftheUbp3pfragmentareinfluencedbyboththeproteasomeandautophagy,sinceinhibitionofeitherpathwaymoderatesthedecreaseinfull-lengthUbp3pandtheappearanceofthedegradationfragment(Fig.6B,C).TheseresultssuggestthatUbp3p,possiblyincomplexwithknownbindingpartnerssuchasCdc48pandUfd3p(Ossareh-Nazarietal.2010),mayberegulatedbytheactivitiesoftheproteasomeandautophagyduringnitrogenstarvationtolimitthesedegradativeprocesses.Previousstudieshavede-scribedaprocesstermedRegulatedUbiquitinProteasome-dependentprocessing(RUP)toyieldpolypeptidefragmentswithbiologicalactivity(RapeandJentsch2004).ItispossiblethataccumulationofaUbp3pfragmentthatretainstheUCHcatalyticdomainbutlackstheBre5pbindingdomaincoulddown-regulateitsdeubiquitinaseactivityandpossiblyallowittoswitchfunctions(Cohenetal.2003;Lietal.2007).Basedonthisinformationandourcurrentresults,wepro-posethemodelpresentedinFigure8.Duringnitrogenexcess(Fig.8A),ubiquitinationofDcp2p,Pop2p,eRF3,andeIF4GImaybeblockedbyTOR1activity(step1).Alternatively,thesefactorsmaybecomeubiquitinated,buttheirUbp3p-depen-dentturnoverisinhibitedbyTOR1activity(step2).Followingtheonsetofnitrogenstarvation(Fig.8B),TOR1activityisrepressed.ThisallowsDcp2p,Pop2p,eRF3,andeIF4GItobeubiquitinatedandsubsequentlyprocessedbytheUbp3pcomplex(consistingofUbp3p–Cdc48p–Ufd3p)

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