Autophagy and mTORC1 regulate the stochastic phase

ARTICLESAutophagyandmTORC1regulatethestochasticphaseofsomaticcellreprogrammingYasongWu1,10,YuanLi1,2,10,HuiZhang1,YinghuaHuang1,PingZhao1,YujiaTang1,XiaohuiQiu1,YueYing1,WenLi3,4,SuNi1,MengZhang1,5,LongqiLiu1,YanXu1,QiangZhuang1,ZhiweiLuo1,ChristinaBenda1,HongSong1,BaohuaLiu6,LiangxueLai1,XingguoLiu1,Hung-FatTse3,7,8,XichenBao1,Wai-YeeChan9,MiguelA.Esteban1,8,BaomingQin1,8,11andDuanqingPei1,8,11Wedescriberobustinductionofautophagyduringthereprogrammingofmousefibroblaststoinducedpluripotentstemcellsbyfourreprogrammingfactors(Sox2,Oct4,Klf4andc-Myc),henceforth4F.Thisprocessoccursindependentlyofp53activation,andismediatedbythesynergisticdownregulationofmechanistictargetofrapamycincomplex1(mTORC1)andtheinductionofautophagy-relatedgenes.The4FcoordinatelyrepressmTORC1,butbifurcateintheirregulationofautophagy-relatedgenes,withKlf4andc-MycinducingthembutSox2andOct4inhibitingthem.Ononehand,inhibitionofmTORC1facilitatesreprogrammingbypromotingcellreshaping(mitochondrialremodellingandcellsizereduction).Ontheotherhand,mTORC1paradoxicallyimpairsreprogrammingbytriggeringautophagy.Autophagydoesnotparticipateincellreshapinginreprogrammingbutinsteaddegradesp62,whoseaccumulationinautophagy-deficientcellsfacilitatesreprogramming.OurresultsthusrevealacomplexsignallingnetworkinvolvingmTORC1inhibitionandautophagyinductionintheearlyphaseofreprogramming,whosedelicatebalanceultimatelydeterminesreprogrammingefficiency.Somaticcellreprogrammingbydefinedfactorsshowcasestheplasticityofcellsandthepoweroftranscriptionfactorstospecifyaparticularfate1.Despitetheprospectofgeneratingpatient-specificinducedpluripotentstemcells(iPSCs)forregenerativemedicine2,3,thepracticeofreprogrammingbydefinedfactorsremainslengthyandhighlyvariableineciencyandquality4.Thesedrawbackshighlighttheneedtounderstandbettertheunderlyingmechanismstoimprovethetechnologyfurther5.Inthisregard,recentstudiesusingbothpopulation-basedandsingle-cellanalysishaveshownthatreprogrammingcanbedividedinto3phases:initiation,maturationandstabilization6,7.Theinitiationphaseisstochasticandmostcellsfailtoactivategenescorrespondingtothe2laterphases,suggestingthatsomeoftheinitialchangesarenegativeandresponsibleforderailingtheprocess.Abettercharacterizationofthisinitiationphasemaythusprovidemuchneededrationalestofacilitatetheprogressionofthereprogrammingprocessinadeterministicmanner.Thiswouldensurenotonlynear100%reprogrammingeciency8,9withoutadditionalgeneticmanipulationbutpotentiallyalsohigherfidelityoftheconversion.Autophagyisacatabolicmechanismresponsibleformaintainingcellularhomeostasis10.Bysequesteringcellularcomponents(forexample,membranes,organelles,cytoskeletalcomponentsorproteincomplexes)intoautophagosomesanddegradingthemthroughthelysosomes,autophagyallowscellstorestoreenergygenerationandsurviveundermetabolicstressconditions,suchasstarvation,orameliorategenotoxicstresses,suchasDNAdamage.Asanintegratedstressresponse,autophagyisinvolvedinavarietyofphysiologicalandpathologicalprocesses,suchasembryonicdevelopment,ageingandcancer11.Notably,aseriesofautophagy-related(Atg)geneshavebeenidentifiedthatcontrolthemembranedynamicsduring1KeyLaboratoryofRegenerativeBiologyoftheChineseAcademyofSciencesandGuangdongProvincialKeyLaboratoryofStemCellsandRegenerativeMedicine,SouthChinaInstituteforStemCellBiologyandRegenerativeMedicine,GuangzhouInstitutesofBiomedicineandHealth,ChineseAcademyofSciences,Guangzhou510530,China.2SchoolofLifeSciences,UniversityofScienceandTechnologyofChina,Hefei,Anhui230027,China.3CardiologyDivision,DepartmentofMedicine,QueenMaryHospital,TheUniversityofHongKong,HongKongSAR,China.4DepartmentofAnatomy,LiKaShingFacultyofMedicine,TheUniversityofHongKong,HongKongSAR,China.5SchoolofLifeSciences,AnhuiUniversity,Hefei230601,China.6HealthScienceCenter,ShenzhenUniversity,Shenzhen518060,China.7ShenzhenInstitutesofResearchandInnovation,theUniversityofHongKong,HongKongSAR,China.8HongKong-GuangdongJointLaboratoryonStemCellandRegenerativeMedicine,theUniversityofHongKongandGuangzhouInstitutesofBiomedicineandHealth,China.9CUHK-GIBHJointResearchLaboratoryofStemCellsandRegenerativeMedicine,SchoolofBiomedicalSciences,FacultyofMedicine,theChineseUniversityofHongKong,HongKong,China.10Theseauthorscontributedequallytothiswork.11CorrespondenceshouldbeaddressedtoB.Q.orD.P.(e-mail:[e-mail][e-mail])Received23May2014;accepted27March2015;publishedonline18May2015;DOI:10.1038/ncb3172NATURECELLBIOLOGYADVANCEONLINEPUBLICATION1©2015MacmillanPublishersLimited.AllrightsreservedARTICLESautophagosomeformationandfusionwithlysosomesinasequentialmanner(initiation,nucleation,elongationandexpansion).TheinitiationphaseofautophagyissuppressedbymTORC1,whichintegratesdierentupstreamnutrientandstresssignalsandalsopromotesbiosynthesis12,13.Notably,throughitsroleinbiosynthesis,mTORC1regulatescellcytoarchitecture(includingcellsizeandorganellecomposition)independentlyofautophagy.Here,wedemonstratethatthewell-adjustedinterplaybetweenmTORC1-mediatedmitochondrialregulationandautophagyisfundamentalform