miR-424(322)-5p targets to inhibit the proliferation and differentiation of myoblasts.

The proliferation and differentiation of myoblasts are considered the key biological processes in muscle development and muscle-related diseases, in which the miRNAs involved remain incompletely understood. Previous research reported that miR-424(322)-5p is highly expressed in mouse skeletal muscle. Therefore, C2C12 cells are used as a model to clarify the effect of miR-424(322)-5p on the proliferation and differentiation of myoblasts. The data show that miR-424(322)-5p exhibits a decreasing trend upon myogenic differentiation. Overexpression of miR-424(322)-5p inhibits the proliferation of myoblasts, manifested by downregulation of proliferation marker genes ( , , and ), decreased percentage of EdU cells, and reduced cell viability. In contrast, these phenotypes are promoted in myoblasts treated with an inhibitor of miR-424(322)-5p. Interestingly, its gain of function inhibits the expression of myogenic regulators, including MyoD, MyoG, MyHC, and Myf5. Additionally, immunofluorescence staining of MyHC and MyoD shows that overexpression of miR-424(322)-5p reduces the number of myotubes and decreases the myotube fusion index. Consistently, inhibition of its function mediated by an inhibitor promotes the expressions of myogenic markers and myotube fusion. Mechanistically, gene prediction and dual-luciferase reporter experiments confirm that enhancer of zeste homolog 1 ( ) is one of the targets of miR-424(322)-5p. Furthermore, knockdown of inhibits the proliferation and differentiation of myoblasts. Compared with NC and inhibitor treatment, inhibitor+si- treatment rescues the phenotypes of proliferation and differentiation mediated by the miR-424(322)-5p inhibitor. Taken together, these data indicate that miR-424(322)-5p targets to negatively regulate the proliferation and differentiation of myoblasts.