Transcriptome Studies Reveal the N(6)-Methyladenosine Differences in Testis of Yaks at Juvenile and Sexual Maturity Stages

SIMPLE SUMMARY: The methylation level of testicular tissue in young yaks and adult yaks was determined, and the N(6)-methyladenosine (m(6)A) level was significantly increased after sexual maturity. Functional enrichment analysis of differentially methylated mRNAs screened between the two groups after sequencing showed that they mainly regulated cell development, spermatogenesis, and testicular endocrine function. Functional analysis of the differentially expressed genes screened between the two groups showed that they were involved in the biological processes of mitosis, meiosis, and flagellated sperm motility. Finally, several genes related to spermatogenesis were screened for further verification. ABSTRACT: Studying the mechanism of spermatogenesis is key to exploring the reproductive characteristics of male yaks. Although N(6)-methyladenosine (m(6)A) RNA modification has been reported to regulate spermatogenesis and reproductive function in mammals, the molecular mechanism of m(6)A in yak testis development and spermatogenesis remains largely unknown. Therefore, we collected testicular tissue from juvenile and adult yaks and found that the m(6)A level significantly increased after sexual maturity in yaks. In MeRIP-seq, 1702 hypermethylated peaks and 724 hypomethylated peaks were identified. The hypermethylated differentially methylated RNAs (DMRs) (CIB2, AK1, FOXJ2, PKDREJ, SLC9A3, and TOPAZ1) mainly regulated spermatogenesis. Functional enrichment analysis showed that DMRs were significantly enriched in the adherens junction, gap junction, and Wnt, PI3K, and mTOR signaling pathways, regulating cell development, spermatogenesis, and testicular endocrine function. The functional analysis of differentially expressed genes showed that they were involved in the biological processes of mitosis, meiosis, and flagellated sperm motility during the sexual maturity of yak testis. We also screened the key regulatory factors of testis development and spermatogenesis by combined analysis, which included BRCA1, CREBBP, STAT3, and SMAD4. This study indexed the m(6)A characteristics of yak testicles at different developmental stages, providing basic data for further research of m(6)A modification regulating yak testicular development.