Targeted Demethylation of the TGFβ1 mRNA Promotes Myoblast Proliferation via Activating the SMAD2 Signaling Pathway

Recent evidence suggested that N6-methyladenosine (m(6)A) methylation can determine m(6)A-modified mRNA fate and play an important role in skeletal muscle development. It was well known that transforming growth factor beta 1 (TGFβ1) is involved in a variety of cellular processes, such as proliferation, differentiation, and apoptosis. However, little is known about the m(6)A-mediated TGFβ1 regulation in myogenesis. Here, we observed an increase in endogenous TGFβ1 expression and activity during myotube differentiation. However, the knockdown of TGFβ1 inhibits the proliferation and induces cell apoptosis of myoblast. Moreover, we found that m(6)A in 5′-untranslated regions (5′UTR) of TGFβ1 promote its decay and inhibit its expression, leading to the blockage of the TGFβ1/SMAD2 signaling pathway. Furthermore, the targeted specific demethylation of TGFβ1 m(6)A using dCas13b-FTO significantly increased the TGFβ1-mediated activity of the SMAD2 signaling pathway, promoting myoblast proliferation. These findings suggest that TGFβ1 is an essential regulator of myoblast growth that is negatively regulated by m(6)A. Overall, these results highlight the critical role of m(6)A-mediated post-transcriptional regulation in myogenesis.