Role of TRPC1 in Myoblasts Differentiation and Muscle Development

Abstract

Myoblasts migration is a key step in myogenesis. It allows myoblasts alignment and their fusion into myotubes. The process has been shown to involve m- or μ-calpains, two calcium-dependent cysteine proteases. Fluorometric measurements of calpain activity in cultured cells showed a peak at the beginning of the differentiation process. We also observed a concomitant and transient increase of the influx of Ca2+ and of the expression of TRPC1 protein. After repression of TRPC1 in myoblasts by siRNA and shRNA, this transient influx of calcium was significantly reduced and the concomitant peak of calpain activity was abolished. Interestingly, myoblasts fusion into myotubes was significantly slowed down, due to a reduced speed of cell migration. Accordingly, migration of control myoblasts was inhibited by 2 to 5 μM GsMT×4 toxin, an inhibitor of TRP channels or by 50 μM Z-Leu-Leu, an inhibitor of calpain. These effects were not observed in TRPC1 knocked down cells. Moreover, TRPC1 knocked down myoblasts also accumulated of myristoylated alanine-rich C-kinase substrate (MARCKS), an actin-binding protein, substrate of calpain. We therefore suggest that an entry of calcium through TRPC1 channels induces a transient activation of calpain, a subsequent proteolysis of MARCKS, allowing in its turn, myoblasts migration and fusion. The role of TRPC1 in muscle regeneration, a process involving myoblasts migration and differentiation, is under study.To further characterize the role of TRPC1 in muscle development, we compared morphological and mechanical parameters of muscles from TRPC1+/+ and TRPC1-/- mice. We observed that muscles from TRPC1-/- mice display a smaller fibre cross-sectional area and generate less force per cross section area. They do not present other major signs of myopathy but were more sensitive to muscle fatigue.