Comprehensive Analysis of the Transcriptome-Wide m(6)A Methylome in Shaziling Pig Testicular Development

RNA N(6)-methyladenosine (m(6)A) modification is one of the principal post-transcriptional modifications and plays a dynamic role in testicular development and spermatogenesis. However, the role of m(6)A in porcine testis is understudied. Here, we performed a comprehensive analysis of the m(6)A transcriptome-wide profile in Shaziling pig testes at birth, puberty, and maturity. We analyzed the total transcriptome m(6)A profile and found that the m(6)A patterns were highly distinct in terms of the modification of the transcriptomes during porcine testis development. We found that key m(6)A methylated genes (AURKC, OVOL, SOX8, ACVR2A, and SPATA46) were highly enriched during spermatogenesis and identified in spermatogenesis-related KEGG pathways, including Wnt, cAMP, mTOR, AMPK, PI3K-Akt, and spliceosome. Our findings indicated that m(6)A methylations are involved in the complex yet well-organized post-transcriptional regulation of porcine testicular development and spermatogenesis. We found that the m(6)A eraser ALKBH5 negatively regulated the proliferation of immature porcine Sertoli cells. Furthermore, we proposed a novel mechanism of m(6)A modification during testicular development: ALKBH5 regulated the RNA methylation level and gene expression of SOX9 mRNA. In addition to serving as a potential target for improving boar reproduction, our findings contributed to the further understanding of the regulation of m(6)A modifications in male reproduction.