Figure 2.
Myoferlin is concentrated at sites of membrane fusion. (A) Confocal images of differentiating myoblasts, where myoferlin (red) appeared at the membrane, concentrated at sites of cell-cell contact (arrow and boxed area). (B) Regions of enhanced myoferlin immunoreactivity (red; and arrowhead) corresponded with membrane, as they were also positive for caveolin 3 immunoreactivity (green; arrowhead). Scale bar: 20 μm. (C) The C2A domain of myoferlin was generated as a GST-fusion protein and tested for calcium-sensitive lipid binding to 3H-labeled vesicles containing 50% phosphotidylcholine and 50% phosphotidylserine. Binding to C2A required the presence of calcium. Myoferlin C2A engineered with the mutation I67D showed no phospholipid binding in the presence of calcium. This mutation is predicted to disrupt the hydrophobic packing of the β-strands of the myoferlin C2A domain.