Myomaker and ether lipids cooperate to promote fusion-competent membrane states

Cell fusion is a fundamental process for skeletal muscle development and regeneration. The muscle-specific fusogens Myomaker and Myomerger are necessary for fusion of muscle cells and together are sufficient for fusion. A mechanistic understanding of Myomaker activity is lacking. Here, we identify ether-linked phospholipids as modulators of Myomaker activity. Although Myomaker shows homology to ceramidases, we observe no ability for the protein to regulate ceramides. Treatment of myocytes with a ceramide synthase inhibitor increases fusion, and lipidomic analysis reveals an increase in cellular ether lipids. Using a lentiviral pseudotyping system to isolate Myomaker-containing membranes, we find an enrichment of ether lipids. Elevating ether lipids in Myomaker-expressing cells induces fusion, even without Myomerger, and correlates with cell surface exposure of phosphatidylethanolamine and phosphatidylserine, indicating a role for Myomaker in remodeling plasma membrane lipid distribution. These findings reveal a link between lipid asymmetry and the molecular machinery responsible for muscle cell fusion.