GRAF1 promotes ferlin-dependent myoblast fusion

•GRAF1 deficient mice exhibit a reduction in skeletal myofiber size.•GRAF1 plays an important role in muscle fusion in vivo.•GRAF1 regulates vesicle trafficking to promote myoblast fusion in vitro.•GRAF1 regulates trafficking of EHD1, myoferlin and Fer1L5 to pre-fusion complexes.•GRAF2 also plays a role in myoblast fusion and EHD1 membrane recruitment.

Myoblast fusion (a critical process by which muscles grow) occurs in a multi-step fashion that requires actin and membrane remodeling; but important questions remain regarding the spatial/temporal regulation of and interrelationship between these processes. We recently reported that the Rho-GAP, GRAF1, was particularly abundant in muscles undergoing fusion to form multinucleated fibers and that enforced expression of GRAF1 in cultured myoblasts induced robust fusion by a process that required GAP-dependent actin remodeling and BAR domain-dependent membrane sculpting. Herein we developed a novel line of GRAF1-deficient mice to explore a role for this protein in the formation/maturation of myotubes in vivo. Post-natal muscles from GRAF1-depleted mice exhibited a significant and persistent reduction in cross-sectional area, impaired regenerative capacity and a significant decrease in force production indicative of lack of efficient myoblast fusion. A significant fusion defect was recapitulated in isolated myoblasts depleted of GRAF1 or its closely related family member GRAF2. Mechanistically, we show that GRAF1 and 2 facilitate myoblast fusion, at least in part, by promoting vesicle-mediated translocation of fusogenic ferlin proteins to the plasma membrane.

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