FER-1 is required for fusion of specialized vesicles, called membranous organelles, with the sperm plasma membrane during Caenorhabditis elegans spermiogenesis. To investigate its role in membranous organelle fusion, we examined ten fer-1 mutations and found that they all cause the same defect in membrane fusion. FER-1 and the ferlin protein family are membrane proteins with four to seven C2 domains. These domains commonly mediate Ca 2+ -dependent lipid-processing events. Most of the fer-1 mutations fall within these C2 domains, showing that they have distinct, non-redundant functions. We found that membranous organelle fusion requires intracellular Ca 2+ and that C2 domain mutations alter Ca 2+ sensitivity. This suggests that the C2 domains are involved in Ca 2+ sensing and further supports their independent function. Using two immunological approaches we found three FER-1 isoforms, two of which might arise from FER-1 by proteolysis. By both light and electron microscopy, these FER-1 proteins were found to be localized to membranous organelle membranes. Dysferlin, a human homologue of FER-1 involved in muscular dystrophy, is required for vesicle fusion during Ca 2+ -induced muscle membrane repair. Our results suggest that the ferlin family members share a conserved mechanism to regulate cell-type-specific membrane fusion.