p53 Regulates Hematopoietic Stem Cell Quiescence

p53RegulatesHematopoieticStemCellQuiescenceYanLiu1,ShannonE.Elf1,YasuhikoMiyata1,GoroSashida1,YuhuiLiu2,GangHuang1,SilvanaDiGiandomenico1,JenniferM.Lee1,AnthonyDeblasio1,SilviaMenendez1,JackAntipin3,BorisReva3,AndrewKoff2,andStephenD.Nimer1,*1MolecularPhamacologyandChemistryProgram,Sloan-KetteringInstitute,MemorialSloan-KetteringCancerCenter,NewYork,NewYork100212MolecularBiologyProgram,Sloan-KetteringInstitute,MemorialSloan-KetteringCancerCenter,NewYork,NewYork100213ComputationalBiologyProgram,Sloan-KetteringInstitute,MemorialSloan-KetteringCancerCenter,NewYork,NewYork10021SUMMARYTheimportanceofthep53proteininthecellularresponsetoDNAdamageiswellknown,butitsfunctionduringsteady-statehematopoiesishasnotbeenestablished.Wehavedefinedacriticalroleofp53inregulatinghematopoieticstemcellquiescence,especiallyinpromotingtheenhancedquiescenceseeninHSCsthatlacktheMEF/ELF4transcriptionfactor.TranscriptionprofilingofHSCsisolatedfromwildtypeandp53nullmiceidentifiedGfi-1andNecdinasp53targetgenesandusinglentiviralvectorstoupregulateorknockdowntheexpressionofthesegenes,weshowtheirimportanceinregulatingHSCquiescence.Establishingtheroleofp53(anditstargetgenes)incontrollingthecellcycleentryofHSCsmayleadtotherapeuticstrategiescapableofeliminatingquiescentcancer(stem)cells.Keywordsp53;HSCquiescence;Mef;Necdin;Gfi-1INTRODUCTIONHematopoieticstemcellscanremainquiescentortheycanenterthecellcycleandeitherself-renewordifferentiateintomultiplelineages.Althoughrelativelyquiescent,theHSCpopulationmustgiverisetoahierarchyofdifferentiatingprogenitorcellpopulationsthatcanreplenishthebloodsystemeachday(AttarandScadden,2004).Bloodcellproductionmustalsorespondefficientlytohematologicstresses,suchasbloodloss,infection,orexposuretocytotoxicagents,viaexpansionoftheHSCand/orprogenitorcellpopulations(Passeguéetal.,2005).Theseprocessesmustoccurwithoutdepletingthestem-cellpool(Veneziaetal.,2004;Forsbergetal.,2005).©2009llPress.Allrightsreserved.*Correspondingauthor:e-mailaddress,[e-mail],Phonenumber:646-888-3040,FAXnumber:646-422-0246.Publisher'sDisclaimer:ThisisaPDFfileofanuneditedmanuscriptthathasbeenacceptedforpublication.Asaservicetoourcustomersweareprovidingthisearlyversionofthemanuscript.Themanuscriptwillundergocopyediting,typesetting,andreviewoftheresultingproofbeforeitispublishedinitsfinalcitableform.Pleasenotethatduringtheproductionprocesserrorsmaybediscoveredwhichcouldaffectthecontent,andalllegaldisclaimersthatapplytothejournalpertain.NIHPublicAccessAuthorManuscriptCellStemCell.Authormanuscript;availableinPMC2010March16.Publishedinfinaleditedformas:CellStemCell.2009January9;4(1):37–48.doi:10.1016/j.stem.2008.11.006.NIH-PAAuthorManuscriptNIH-PAAuthorManuscriptNIH-PAAuthorManuscriptHSCquiescenceislikelycontrolledbybothHSC-intrinsicmechanismsandbonemarrowmicroenvironmentalfactors(WilsonandTrumpp,2006).SeveraltranscriptionfactorshavebeenshowntoplaykeyrolesinHSCcellfatedecisions.Gfi-1hasbeenshowntorestrictHSCproliferationandpreserveHSCfunctionalintegrity(Hocketal.,2004;Zengetal.,2004).HOXB4andGATA-2regulateHSCself-renewal(Krosletal.,2003;Lingetal.,2004),whereastheEtstranscriptionfactorMEF/ELF4regulatesbothHSCself-renewalandquiescence(Lacorazzaetal.,2006).MefnullmiceexhibitgreaternumbersofHSCs(i.e.LSKcells)andMefnullLSKcellsaremorequiescentthannormal.Earlystudiessuggestedaroleforp21inrestrictingHSCentryintothecellcycleandregulatingHSCpoolsizeandHSCexhaustionunderstress(Chengetal.,2000).However,theregulationofHSCself-renewalbyp21understeady-stateconditionsmaybeminimal(vanOsetal.,2007).SeveralstudieshaveimplicatedboththeAng-1/Tie2andthethrombopoietin/MPLsignalingpathwaysinmaintainingHSCquiescence(Araietal.,2004;Yoshiharaetal.,2007;Qianetal.,2007).Thep53tumorsuppressorgenemayregulatevariousaspectsofhematopoieticcellbehavior(Wlodarskiteal.,1998;TeKippeetal.,2003;Chenetal.,2008;Alakaetal.,2008).Althoughhematopoiesisinp53knock-outmiceappearstoproceednormally,numerousstudieshaveidentifiedrolesforp53intheproliferation,differentiation,apoptosisandagingofhematopoieticcells(Shounanetal.,1996;Shaulskyetal.,1991;Kastanetal.,1991;Lotemetal.,1993;Guzmanetal.,2002;Parketal.,2003;Dumbleetal.,2007).Moreover,p53deletionsandmutationshavebeenfoundathighfrequencyinblastcrisischronicmyelogenousleukemiaandwithsomefrequencyinacuteleukemia(ProkocimerandRotter,1994).InresponsetoDNAdamage,p53caneitherelicitcell-cyclearrestorapoptosis,thetypicaloutcomeformaturehematopoieticcells(Wuetal.,2005).Butp53hasrecentlybeenshowntonegativelyregulateneuralstemcellproliferationandself-renewal(Meletisetal.,2006),andgiventhatlong-termreconstitutingHSCs(LT-HSC)expresshighlevelsofp53transcripts(Forsbergetal.,2005andourdata,seebelow),weexaminedthefunctionofp53duringsteady-statehematopoiesis.WefindanimportantinterdependencybetweenMEF/ELF4andp53onHSCquiescenceandidentifytwop53targetgenes,Gfi-1andNecdin,thatregulatequiescenceinwildtype(andMefnullHSCs).Ourfindingsidentifydistinctrolesforp53inrestingvs.cyclingcells.RESULTSMaintainingHSCquiescencebyp53WehaverecentlyfoundthatMef/Elf4nullmous