Study On The Expression And Function Of MARF1 In Hamster Oocytes

Oocyte quality is a key factor in determining women’s fertility and reproductive health.The quality of oocytes largely depends on whether they can recover and finish meiosis,whether they can fertilize and support embryonic development,and the integrity of their genome.Although this field has been widely studied,the specific molecular mechanism is far from being clarified.Recent series of studies have shown that MARF1(Meiosis regulator and mRNA stability factor 1)is a key factor that can specifically regulate the development and maturation of oocytes and play a decisive role in the quality of oocytes.Its gene mutation lead to female infertility without affecting male reproductive ability.The mutant female mice discharged immature oocytes because their oocytes could not resume meiosis;At the same time,the number of DNA double strand breaks and the expression level of retrotransposon in the oocytes of mutant female mice increased,and the transcriptome and cytoplasmic maturation were defective.Therefore,MARF1 is an important factor that can specifically regulate meiosis and genomic integrity of oocytes at the same time.Recent studies in Drosophila have also found that MARF1 deletion leads to the arrest of meiotic maturation of its oocytes,indicating that MARF1 is evolutionarily conservative,and it may also play an equally important role in the development and maturation of human oocytes.However,due to ethical constraints,the research on MARF1 regulating female reproduction cannot be carried out easily.Therefore,it is necessary to use species similar to the regulation mode of human oocyte development to carry out relevant research.Golden hamster has been widely used in the research fields of novel coronavirus,oncology,reproductive physiology and other research fields due to its higher similarity with humans in some characteristics of physiology,pathology and immunology,and the advantages of short reproductive cycle,large number of offspring,simple feeding and low cost.The latest research show that hamster oocytes are closer to human oocytes in the regulation of PIWI-piRNA pathway than mouse.Our previous studies suggest that the function of MARF1 in mouse oocytes may be similar to that of PIWI-piRNA pathway in male mouse germ cells.This suggests that hamster may be a more ideal animal model for studying MARF1 regulation of human oocyte development and maturation.Therefore,in this paper,the expression and localization of MARF1 in hamster oocytes and its role in oocyte maturation and genome integrity maintenance were studied by using the new model of hamster oocyte in vitro maturation and in vivo Marfl knock out(KO),with a view to providing useful insights into the regulatory role of MARF1 in human reproduction.Through sequence alignment analysis and cloning and sequencing analysis of Marfl cDNA,we found that the key domain of MARF1 protein may be highly conserved in different species,and found that Marf1 expressed in hamster oocytes is a unique splicing variant different from somatic and reference sequences;Its mRNA and protein are highly expressed in oocytes.Immunofluorescence staining show that MARF1 has different localization and distribution before and after meiotic recovery of oocytes,suggesting that MARF1 may play different functions through different mechanisms before and after meiotic recovery of oocytes.Nearly half(47.14%)of the oocytes discharged from Marf-KO female hamsters were arrested in GV phase,while more than half of the other half recovered meiosis but had serious meiosis defects,which further indicated that MARF1 not only regulated meiosis recovery of hamster oocytes but also participated in the regulation of meiosis process of oocytes.Different from mice,the mRNA level of Ppp2cb in Marf-KO hamster oocytes is not up-regulated,but the mRNA level of Cdc25b was significantly decreased.This indicates that MARF1 may promote the activation of MPF and further regulate the maturation process of oocytes by promoting the expression of Cdc25b in hamster oocytes;At the same time,severe DNA damage and specific retrotransposon activation occurred in Marf-KO hamster oocytes.These results suggest that MARF1 may regulate the integrity of oocyte genome through different mechanisms in hamsters.In conclusion,this study revealed the specific expression and localization of MARF1 during the development and maturation of hamster oocytes and its specific regulatory function for meiosis and genomic integrity of oocytes,which provided a reference for discussing the regulation of MARF 1 on the development and maturation of human oocytes and its role in female reproduction.