Transcriptome-wide Dynamics of m⁶A mRNA Methylation During Porcine Spermatogenesis

Spermatogenesis is a continual process that occurs in the testes, in which diploid spermatogonial stem cells (SSCs) differentiate and generate haploid spermatozoa. This highly efficient and intricate process is orchestrated at multiple levels. N-6-methyladenosine (m(6)A), an epigenetic modification prevalent in mRNAs, is implicated in the transcriptional regulation during spermatogenesis. However, the dynamics of m(6)A modification in non-rodent mammalian species remains unclear. Here, we systematically investigated the profile and role of m(6)A during spermatogenesis in pigs. By analyzing the transcriptomic distribution of m(6)A in spermatogonia, spermatocytes, and round spermatids, we identified a globally conserved m(6)A pattern between porcine and murine genes with spermatogenic function. We found that m(6)A was enriched in a group of genes that specifically encode the metabolic enzymes and regulators. In addition, transcriptomes in porcine male germ cells could be subjected to the m(6)A modification. Our data show that m(6)A plays the regulatory roles during spermatogenesis in pigs, which is similar to that in mice. Illustrations of this point are three genes (SETDB1, FOXO1, and FOXO3) that are crucial to the determination of the fate of SSCs. To the best of our knowledge, this study for the first time uncovers the expression profile and role of m(6) Aduring spermatogenesis in large animals and provides insights into the intricate transcriptional regulation underlying the lifelong male fertility in non-rodent mammalian species.