Effects of DNMT1 and TET2 on proliferation and differentiation of goose embryonic myoblasts

Shitou goose (large goose breed) and Wuzong goose (small goose breed) are important local goose breeds in Guangdong Province, exhibiting significant differences in growth performance and slaughter traits. The number of myofibers in goose skeletal muscle is determined during embryonic stage, and is subject to multiple myogenic factors and epigenetic regulation during myogenesis, among which DNA methylation is the most common and obvious epigenetic modification. In this study, we examined the expression profiles of DNA methyltransferases (DNMT1, DNMT3A, DNMT3B), DNA hydroxylases (TET1, TET2, TET3) and key myogenic factors (MYOD, MYF5, MYOG, MYHC) in embryonic and adult myoblasts, and performed DNMT1 and TET2 over-expression and interference experiments on goose embryonic myoblasts in combination with co-treatments of BC339 (TETs inhibitor) and 5AZA (DNMTs inhibitor). RT-qPCR analysis revealed Wuzong geese embryonic myoblasts exhibited significantly higher mRNA levels of TET1, TET2, and TET3 hydroxylases during early stages (E10-E22), along with elevated MYOD expression, while Shitou geese embryonic myoblasts showed increased DNMT1 and DNMT3B methyltransferases and up-regulated late-stage myogenic factors MYOG and MYHC. In myoblasts, TET2 expression peaked during proliferation, whereas DNMT1 dominated differentiation, highlighting their stage-specific regulatory roles. DNMT1 over-expression suppressed proliferation and differentiation, lower MYOD and MYHC levels, while DNMT1 interference enhanced both processes. 5AZA reversed DNMT1-mediated suppression, whereas BC339 exacerbated it. Conversely, TET2 over-expression promoted proliferation and differentiation, increased MYOD and MYHC levels, with 5AZA amplifying and BC339 attenuating these effects. TET2 interference impaired proliferation and differentiation, partially rescued by 5AZA. In this study, we investigated the relationship between DNA methylation and the proliferation and differentiation ability of goose embryonic myoblasts, which provided a theoretical basis for a systematic understanding of the molecular mechanisms underlying the differences in growth performance of goose breeds of different body sizes.