Ca2+ Regulates the Interaction between Synaptotagmin and Syntaxin 1

While there is compelling evidence that the synaptic vesicle protein synaptotagmin serves as the major Ca2+ sensor for regulated exocytosis, it is not known how Ca2+ binding initiates membrane fusion. Here we report that Ca2+ increases the affinity, by approximately 2 orders of magnitude, between synaptotagmin and syntaxin 1, a component of the synaptic fusion apparatus. This effect is specific for divalent cations which can stimulate exocytosis of synaptic vesicles (Ca2+ > Ba2+, Sr2+ >> Mg2+). The Ca(2+)-dependence of the interaction was composed of two components with EC50 values of 0.7 and 180 microM Ca2+. The interaction is mediated by the carboxyl-terminal region of syntaxin 1 (residues 194-288) and is regulated by a novel Ca(2+)-binding site(s) which does not require phospholipids and is not disrupted by mutations that abolish Ca(2+)-dependent phospholipid binding to synaptotagmin. We propose that this interaction constitutes an essential step in excitation-secretion coupling.