The role of CA2+-dependent binding residues of synaptotagmin in neurotransmission

Abstract

Synaptic vesicle fusion and the concomitant release of neurotransmitter has long been a field of intense study. Synaptotagmin I is the Ca2+ sensor for this synaptic vesicle fusion, but the mechanism by which it triggers this fusion is still unclear. Synaptotagmin is composed of an intravesicular domain, a transmembrane domain through the synaptic vesicle and two tandem, cytosolic C2 domains that both bind Ca2+. Multiple Ca2+-dependent interactions have been mapped to these C2 domains, including those with SNAREs, the proposed fusion machinery, and the phospholipids present in both the vesicular and synaptic membranes. I have tested the importance for synaptic transmission of several residues that mediate phospholipid binding of both C2 domains using site directed mutagenesis in Drosophila third instar larvae. These larvae all showed deficits of various severity in their evoked release consistent with the hypothesis that Ca2+-dependent phospholipid binding by these residues is required for efficient synaptic transmission. I also demonstrate that Ca2+-dependent phospholipid binding by C2B is of greater physiological significance than that of C2A. Taken together, these results suggest that Ca2+-dependent phospholipid binding by the C2 domains mediate the majority synaptotagmin's role as the Ca sensor for evoked release.