Identification and functional annotation of m6A methylation modification in granulosa cells during antral follicle development in pigs.

The N-methyladenosine (m6A) derivative has the capacity for ubiquitous epigenetic modification of messenger RNA (mRNA) that regulates gene expression through post-transcriptional mRNA modifications. Findings with mapping of m6A methylomes have indicated there are potential functions of this derivative in different cell types of several species. A profile of m6A methylomes and potential functions in granulosa cells of pigs during antral follicle development, however, has not yet occurred. In the present study, there was profiling of an epitranscriptome-wide map of m6A methylation in granulosa cells of pigs derived from small and large follicles using methylated RNA immunoprecipitation techniques, next-generation sequencing and further annotation of the potential functions of m6A utilizing bioinformatic analyses procedures. The m6A modification is abundant in granulosa cells of pigs, and there are dynamic changes in m6A methylomes during the developmental transition from small (< 3 mm) to large (> 5 mm) sized follicles. In particular, there was a prevalence of 7289 and 6882 m6A in granulosa cells from follicles of two different sizes. There was an increased prevalence of m6A in close proximity to the 5' or 3'-untranslated coding regions and a shared conserved consensus motif. Results from further analysis indicated there was significant enrichment of differentially expressed m6A methylated genes in several signaling pathways associated with steroidogenesis, granulosa cell proliferation and follicular development. When considered as a whole, these results indicate there are differential m6A modifications in granulosa cells of pigs during follicle development that are potentially associated with steroidogenesis and folliculogenesis.