Study On The Role Of MiRNA-383and MiRNA-320in Mouse Follicular Development

Our previous studies have indicated that microRNA-383(miR-383) and microRNA-320(miR-320) are two of the most down-regulated miRNAs in TGF-β1-treated mouse ovarian granulosa cells (GC). However, the roles and mechanisms of miR-383and miR-320in GC functions during follicular development remain unknown. In the first study of this paper, miR-383was mainly expressed in the GC and oocytes of mouse ovarian follicles in different development stage. Overexpression of miR-383increased estradiol production in GC by binding to the3’UTR of RNA binding motif, single stranded interacting protein1(RBMS1). miR-383suppressed the expression of RBMS1by regulating its mRNA stability, which subsequently inhibited the expression of c-Myc (a downstream gene of RBMS1). Overexpression of RBMS1or c-Myc inhibited miR-383-mediated steroidogenesis-promoting effects. Silence of the transcription factor steroidogenic factor-1(SF-1) significantly inhibited the expression of Sarcoglycan zeta (SGCZ)(the host gene of miR-383), primary and mature miR-383in GC. These indicated that miR-383was transcriptionally promoted by SF-1. Luciferase assays and chromatin immunoprecipitation assays showed that SF-1specifically bound to the promoter region of SGCZ and then transactivated the expression of miR-383and SGCZ. In addition, SF-1participated in the regulation of miR-383-and RBMS1/c-Myc-mediated estradiol production in the GC. These results reveal that miR-383promotes steroidogenesis by targeting RBMS1, partially through inactivation of c-Myc. SF-1plays positive roles in the regulation of miR-383processing and function in the GC.Treatment of21PND female mice with PMSG resulted in the decreased expression of miR-320in a time-dependent manner. miR-320was mainly expressed in GC and oocytes of mouse ovarian follicles in the follicular development. There is negative correlation between the expression of miR-320and the development stage of mouse ovaries. The expression of miR-320is down-regulated in FSH-treated mouse GC in a time-and does-dependent manner. miR-320inhibited estradiol synthetize and proliferation of GC through targeting E2F1and SF-1. Overexpression or knockdown of E2F1or SF-1rescued and enhanced miR-320-mediated the suppression of GC proliferation and steroidogenesis-promoting effects. The expression of SF-1and E2F1was negatively correlated with the expression of miR-320in the development stage of mouse ovaries and PMSG-treated mouse ovary. These results indicates that miR-320 promtes the proliferation and steroids production through targeting E2F1and SF-1in GC. Injection of miR-320into the ovarian bursa of mice promoted the production of testosterone and progesterone, but inhibited estradiol release in the serum. These data demonstrates that miR-320plays an important role in the follicular development.By summarizing the above studies, miRNA-383and miRNA-320regulate the GC proliferation and steroid production by targeting different genes in the follicular development. Understanding of the regulation of miRNA biogenesis and function in the follicular development will potentiate the usefulness of miRNA in the treatment of reproduction and some steroid-related disorders.