The Function Of MAPK Cascade And TCF12 In Oocyte Maturation And Preimplantational Development

During mammalian follicular growth,the transcription and translation of oocytes are active,massive maternal m RNAs and proteins accumulate in oocytes.As chromatin condenses into chromosome,full-blown oocytes become transcriptionally silenced.The process of meiotic resumption,meiotic maturation and preimplantational development before zygotic genome activation(ZGA)are all regulated by maternal products stored in the cytoplasm.However,the molecular mechanism underlying the regulation and function of maternal transcripts are not well known.In mammalian species,both the maturation promoting factor(MPF)and mitogen-activated protein kinase(MAPK)cascades play critical roles in modulating oocyte meiotic cell-cycle progression.MPF is a critical heterodimer that is composed of CDK1(cyclin-dependent kinase-1)and cyclin B1.MAPK cascade is composed of MOS(moloney sarcoma oncogene),MEK1/2(MAPK-ERK kinase 1 and 2)and ERK1/2(Extracellular signal-regulated kinases 1/2).Activation of MPF and ERK1/2 requires the activation of maternal Ccnb1(coding cyclin B1)and Mos m RNAs translation,respectively.The phosphorylation and degradation of CPEB1(cytoplasmic polyadenylation element binding protein 1)that triggered by ERK1/2 is a principal mechanism in activating maternal m RNA translation.However,the activation of ERK1/2 is later than MPF,and the accumulation of MOS protein around GVBD(germinal vesical breakdown)is a prerequisite for the activation of the MAPK cascade.In addition,MPF and MAPK activities intimately affect each other during oocyte meiosis.For these reasons,the interplay of the MPF and MAPK cascades in regulating Ccnb1 and Mos m RNA translation are not be concluded.On the other hand,the basic helix-loop-helix(b HLH)transcription factors are widespread in eukaryotes,controling growth and development.E protein is an important derivation of b HLH family,comprising TCF4(transcription factor 4)and TCF3(transcription factor3),as well as TCF12(transcription factor 12).It was reported that Tcf12 mutation homozygotes are female infertile.In this study,we applied mice as model organism to investigate the mechanism by which the MAPK cascade regulated the translation activation,and the role of the transcriptional accumulation by TCF12 in female reproduction.Our research found that translational activation of Ccnb1 transcripts containing a long 3?-UTR during meiotic resumption works in an ERK1/2-dependent way.A low level of ERK1/2 activation was detected prior to meiotic resumption.Precocious activation of MAPK signaling in germinal vesicle stage oocytes promoted the translation of Ccnb1 m RNAs and meiotic maturation.Inhibition or precocious activation of CDK1 activity had an appreciable effect on the translation of Ccnb1 m RNA,suggesting that both kinases are required for Ccnb1 m RNA translational activation.CDK1 triggered phosphorylation,but not degradation,of CPEB1 in oocytes;the degradation of CPEB1 was only triggered by ERK1/2.Moreover,the translational activation of Mos m RNA was regulated by ERK1/2 and cytoplasmic polyadenylation elements too.Taken together,the cooperation and positive feedback activation of ERK1/2 and CDK1 lead to the fine-tuning of m RNA translation and cell-cycle progression during mouse oocyte maturation.Moreover,we applied oocyte-expressed Gdf9-Cre in primordial follicles to specifically delete Tcf12 to study the function of maternal accumulation in female fertility.We proved evidence that TCF12 is a transcription factor abundantly expressed in the nucleus of oocytes during growing phase.TCF12 recognizes and binds target genes through functional domains to moderate the transcriptional activity of target genes.Maternal deletion of Tcf12 led to female sterility.Although Tcf12 does not participate in oocyte maturation,it is essential for fertilization and preimplantational development.On the one hand,Tcf12 maintained the competence of fertilization by controling the expression of Astl,which influencing the proper location of cortical granules(CG).On the other hand,Tcf12 probably preserved the phosphatase activity by regulating the expression of maternal Arpp19.Zygotes without TCF12 had a prolonged mitosis cell cycle upon a drop of PP2 A activity inhibition.Oocyte-specific deletion of Tcf12 led to ZGA failure at the 2-cell stage,with excessive H3K27me3 modifications.In the subsequent process of development,maternal knockout embryos gradually lost their development potential.These observations clarifies the maternal effect induced by Tcf12 ensures preimplantational development.Our research not only revealed the interplay of kinases in mediating translational activation of cell-cycle related m RNAs during oocyte maturation,but also explored the function of maternal accumulation induced by transcriptional factors in maintaining female fertility.We systematically studied the regulation of transcriptome during oocyte growth,maturation and early embryonic development,and clarified the significant roles of these regulations on female reproduction,providing a new theoretical basis for clinical infertility problems and the advancement of assisted reproductive technology.