MicroRNA-499-5p regulates skeletal myofiber specification via NFATc1/MEF2C pathway and Thrap1/MEF2C axis

This study aimed to investigate the role of microRNA-499-5p (miR-499-5p) in the regulation of skeletal myofiber specification and its underlying mechanisms.

Mouse C2C12 cells were used in this study. Cyclosporin A and siRNA targeting Thrap1 (si-Thrap1) were used to inhibit NFATc1/MEF2C pathway and knockdown Thrap1, respectively. The expressions of miR-499-5p and genes were evaluated by real-time quantitative PCR and western blot analysis.

Overexpression of miR-499-5p promoted oxidative fiber gene expression and repressed glycolytic fiber gene expression, affecting several factors associated with fiber specification including NFATc1/MEF2C pathway, PGC-1α, FoxO1 and Wnt5a. Inhibition of NFATc1/MEF2C pathway partly reduced the effect of miR-499-5p overexpression on muscle fiber gene expression. MiR-499-5p targeted Thrap1 in proliferating and differentiating C2C12 cells. Knockdown of Thrap1 showed a parallel function with miR-499-5p overexpression on muscle fiber gene expression and NFATc1/MEF2C pathway, accompanied by an increase of miR-499-5p level. The effects of miR-499-5p inhibitor on muscle fiber type specific gene expression and NFATc1/MEF2C pathway were effectively reversed by Thrap1 knockdown.

MiR-499-5p regulated skeletal myofiber specification and affected several factors associated with fiber specification. MiR-499-5p regulated muscle gene expression partly through NFATc1/MEF2C pathway. We also showed a clue that miR-499-5p regulates skeletal muscle fiber specification in C2C12 cells through targeting Thrap1, thereby, promoting NFATc1/MEF2C pathway and then triggering a series of oxidative muscle fiber gene expression.