Biochemical studies of mammalian oogenesis: Synthesis and stability of various classes of RNA during growth of the mouse oocyte in vitro☆

[摘要] ThesynthesisofvariousclassesofRNAinmouseoocytesatdifferentstagesofgrowthhasbeenexaminedafterincubatingfolliclesinmediumcontainingradiolabeleduridine.Afterfractionationonpoly(U)-SepharoseofradiolabeledoocyteRNA,ofwhichabout83%isassociatedwiththenucleusaftera5-hrlabelingperiod,revealedthatabout40–50%oftheradiolabeledRNAbehavedaspoly(A)-containingRNA.Thisvalueremainedfairlyconstantduringtheperiodofoocytegrowthinwhichoocytediameterincreasedfromabout35toabout55μm.Aftera5-hrlabeling,thepercentageofradiolabeledpoly(A)-containingRNAineitherthefullygrowndictyateoocyte,metaphaseIIoocyte,orone-cellembryowasabout20%.Aftera5-hrlabeling,agarosegelelectrophoreticanalysisoftheradiolabeledspeciesofoocyteRNAobtainedafterfractionationonpoly(U)-SepharoserevealedthepresenceofaputativeribosomalRNAprecursor,ribosomal(28and18S)RNA,transferplus5SRNAandheterodispersepoly(A)-containingRNA.AsignificantfractionoftheradiolabeledRNAspecieswasquitelarge(>40S).TheratiosoftherelativeproportionsoftheradiolabeledribosomalRNAsandtransferplus5SRNAremainedessentiallyconstantduringoocytegrowth.ThestabilityofvariousclassesofRNAwasexaminedbyincubatingfollicleswithradiolabeleduridine,washingthefolliclesfreeofradioactivityandculturingthefolliclesunderconditionswhichsupportoocytegrowthinvitro(Eppig,1977).Undertheseconditions,totaloocyteradiolabeledRNAwasquitestableasdeterminedbyretentionofacid-insolubleradioactivematerial(t12=28days).However,underconditionsinwhichoocytesareviablebutdonotgrow,thehalf-lifeoftotalRNAwasabout4.5days.Poly(A)-containingRNAwasalsoverystable;after8daysinculture,about50%oftheradiolabeledpoly(A)-containingRNApresentafter5hroflabelingwasstillpresent.AgarosegelelectrophoreticanalysisofradiolabeledRNAinoocytesafter4daysofcultureandafterfractionationonpoly(U)-Sepharoserevealedthepresenceofribosomal(28and18S)RNA,transferplus5SRNA,andheterodispersepoly(A)-containingRNA.Atthistime,theseRNAsarelocatedintheoocytecytoplasm.Inaddition,themolecularweightdistributionofpoly(A)-containingRNAwassignificantlylowerthanthatafter5hroflabeling.TheratiosoftherelativeproportionsofradiolabeledribosomalRNAsandtransferplus5SRNAwerequitesimilartothoseafter5hroflabeling.

[学科分类] 生物科学（综合）