Study On The Relationship Between The Initiation Of Meiosis In Female Germ Cells And The Electronic Respiratory Chain

To generate haploid gametes by meiosis is a unique property of germ cells and is critical for sexual reproduction.Abnormal meiosis can cause aneuploidy,eventually led to the early embryonic developmental abnormalities,embryonic death and birth defects.Although the development of mammalian embryonic gonads has been studied for over half a century,a number of genes are known to be important for meiotic initiation,our understanding of the regulation of meiotic initiation remains fragmentary,especially at protein level.To advance our understanding of female germ cells meiosis initiation at protein level,in our previous work,we construct a comparative proteome profile of female mouse gonads at specific time points(E11.5,E12.5 and E13.5),spanning a critical window in female germ cells initiating meiosis.We identified 3666 proteins,including 473 significantly differentially expressed.Further bioinformatics analysis showed that these differentially expressed proteins were enriched in mitochondrion,especially in electron transport chain.It should be noted that among the proteins which contributed electron transport chain complex I,23.8% proteins were the differentially expressed proteins,which indicated that the electron transport chain was tightly related to meiosis initiation of female germ cells.Thus,this study mainly focused on the relationship between female meiosis initiation and the electron transport chain in mouse.By adding rotenone to E11.5 female gonads cultured-in vitro,we found no obvious difference in germ cell number via counting DDX4(DEAD Asp-Glu-AlaAsp box polypeptide 4,the germ cell marker)postive cells,the proportion of ?H2AX(The phosphorylated form of Histone H2 AX,the meiotic cell marker)positive germ cells had significantly decreased 40% in the culture under 0.1?M rotenone compared with control groups,while the proportion of OCT4(POU domain,class 5,transcription factor 1,the oogonia marker)positive germ cells obviously increased 50%,and GATA4(Transcription factor GATA-4,the somatic cell marker)positive cells had no obvious difference.In further study,by adding a well-known antioxidant,CoQ10,to 0.1?M rotenone group could rescue the phenotype.To explore the reason for the occurrence of meiosis arrest,we observed the 2C germ cells obviously increased in the culture under 0.1?M rotenone compared to normal and control group,female germ cells could not complete premeiotic DNA replication.Further through the detection of ATP and ROS levels,we found the level of ATP decreased 30% in the culture under 0.1?M rotenone,and the ROS level increased about 40% in the culture under 0.1?M rotenone compared to normal and control group.During the exploration of the mechanism of female germ cell meiosis arrest,we observed in the culture under 0.1?M rotenone,the expression of two important factors,CyclinE and P-RB1,were obviously decreased during the process of G1-S transition compared with normal and control group.This result further confirmed that female germ cells were arrested and the oogonia could not complete the premeiotic DNA replication.Taken together,the study proved suppressing electron transport chain by rotenone might inhibit female germ cells meiosis initation.Characterization of the quantitative proteome of female gonads and verification of the relationship between the electron transport chain and meiosis initiation help us to better understand the mechanismof meiosis initiation.This work may provide new ideas for the study of generation and differentiation of germ cells and more potential molecular targets for clinical genetic screening gamete in assisted reproductive technology.