RNA Sequencing Reveals the Regulation of Betaine on Chicken Myogenesis

Simple Summary Betaine is a healthy source of methyl and glycine and a widely used feed additive to promote animal growth. Previous studies mainly focused on its anti-osmotic pressure, anti-inflammatory, and growth-promoting effect in vitro. Still, poultry's growth-promoting mechanism and regulation of muscle cells remain unknown. In this study, we found that a low concentration of betaine could inhibit myoblast cell proliferation but promote myotube formation, suggesting that the growth-promoting effect of betaine was achieved through the differentiation and fusion of myotubes. In addition, RNA sequencing found a series of betaine-affected genes, which could provide a theoretical basis for explaining betaine's regulation on chicken myogenesis in vitro. Betaine is trimethylglycine and a universal methyl donor which could provide methyl and glycine for cells and animals. As a new star in epigenetics, N6-Methyladenosine has been reported to regulate multiple biological activities, but the regulatory mechanism of betaine on N6-Methyladenosine as well as myogenesis was little studied. In this study, we treated chicken primary myoblast cells with different concentrations of betaine (0, 10, 25, and 50 mmol/L) and found that myoblast cell proliferation was inhibited, although the cell cycle was promoted in the S phase by betaine, where the myotube area was increased as well as the differentiation marker genes MyoD, MyoG, MyHC, Myomarker, and Ckm. RNA sequencing obtained a total of 61 differentially expressed genes (DEGs); DEGs caused by 50 mmol/L betaine were mainly enriched in the regulation of skeletal muscle tissue regeneration and some amino acid metabolic processes. The gene expression pattern trends of all DEGs were mainly clustered into 2 profiles, with the increase in betaine concentration, the gene expression pattern either increased or decreased continuously. Overall, a low concentration betaine can increase the N6-Methyladenosine modification level and myotube area but depresses myoblast cell proliferation in vitro.