Ca. Earles et al., The tandem C2 domains of synaptotagmin contain redundant Ca2+ binding sites that cooperate to engage t-SNAREs and trigger exocytosis, J CELL BIOL, 154(6), 2001, pp. 1117-1123
Real-time voltammetry measurements from cracked PC12 cells were used to ana
lyze the role of synaptotagmin-SNARE interactions during Ca2+-triggered exo
cytosis. The isolated C2A domain of synaptotagmin I neither binds SNAREs no
r inhibits norepinephrine secretion. In contrast, two C2 domains in tandem
(either C2A-C2B or C2A-C2A) bind strongly to SNAREs, displace native synapt
otagmin from SNARE complexes, and rapidly inhibit exocytosis. The tandem C2
domains of synaptotagmin cooperate via a novel mechanism in which the disr
uptive effects of Ca2+ ligand mutations in one C2 domain can be partially a
lleviated by the presence of an adjacent C2 domain. Complete disruption of
Ca2+-triggered membrane and target membrane SNARE interactions required sim
ultaneous neutralization of Ca2+ ligands in both C2 domains of the protein.
We conclude that synaptotagmin-SNARE interactions regulate membrane fusion
and that cooperation between synaptotagmin's C2 domains is crucial to its
function.