miR-135a Suppresses Granulosa Cell Growth by Targeting Tgfbr1 and Ccnd2 during Folliculogenesis in Mice.

The success of female reproduction relies on high quality oocytes, which is determined by well-organized cooperation between granulosa cells (GCs) and oocytes during folliculogenesis. GC growth plays a crucial role in maintaining follicle development. Herein, miR-135a was identified as a differentially expressed microRNA in pre-ovulatory ovarian follicles between Large White and Chinese Taihu sows detected by Solexa deep sequencing. We found that miR-135a could significantly facilitate the accumulation of cells arrested at the G1/S phase boundary and increase apoptosis. Mechanically, miR-135a suppressed transforming growth factor, beta receptor I (Tgfbr1) and cyclin D2 (Ccnd2) expression by targeting their 3′UTR in GCs. Furthermore, subcellular localization analysis and a chromatin immunoprecipitation-quantitative real-time PCR (ChIP-qPCR) assay demonstrated that the TGFBR1-SMAD3 pathway could enhance Ccnd2 promoter activity and thus upregulate Ccnd2 expression. Finally, estrogen receptor 2 (ESR2) functioned as a transcription factor by directly binding to the miR-135a promoter region and decreasing the transcriptional activity of miR-135a. Taken together, our study reveals a pro-survival mechanism of ESR2/miR-135a/Tgfbr1/Ccnd2 axis for GC growth, and also provides a novel target for the improvement of female fertility.