Ryanodine receptor RyR1-mediated elevation of Ca2+ concentration is required for the late stage of myogenic differentiation and fusion

Background: Cytosolic CA²⁺ plays vital roles in myogenesis and muscle development. As a major CA²⁺ release channel of endoplasmic reticulum (ER), ryanodine receptor 1 (RyR1) key mutations are main causes of severe congenital myopathies. The role of RyR1 in myogenic differentiation has attracted intense research interest but remains unclear.

Results: In the present study, both RyR1-knockdown myoblasts and CRISPR/Cas9-based RyR1-knockout myoblasts were employed to explore the role of RyR1 in myogenic differentiation, myotube formation as well as the potential mechanism of RyR1-related myopathies. We observed that RyR1 expression was dramatically increased during the late stage of myogenic differentiation, accompanied by significantly elevated cytoplasmic CA²⁺ concentration. Inhibition of RyR1 by siRNA-mediated knockdown or chemical inhibitor, dantrolene, significantly reduced cytosolic CA²⁺ and blocked multinucleated myotube formation. The elevation of cytoplasmic CA²⁺ concentration can effectively relieve myogenic differentiation stagnation by RyR1 inhibition, demonstrating that RyR1 modulates myogenic differentiation via regulation of CA²⁺ release channel. However, RyR1-knockout-induced CA²⁺ leakage led to the severe ER stress and excessive unfolded protein response, and drove myoblasts into Apoptosis.

Conclusions: Therefore, we concluded that CA²⁺ release mediated by dramatic increase in RyR1 expression is required for the late stage of myogenic differentiation and fusion. This study contributes to a novel understanding of the role of RyR1 in myogenic differentiation and related congenital myopathies, and provides a potential target for regulation of muscle characteristics and meat quality.