Molecular mechanism of membrane fusion in salivary exocytosis.

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Parotid acinar cells contain VAMP-2 as a v-SNARE on secretory granule membranes, but do not have brain-type t-SNAREs, such as syntaxin-1 o SNAP-25. In order to examine whether or not syntaxin-4 and SNAP-23, plausible candidates of t-SNAREs in non-neuronal cells, can act as t-SNAREs in parotid acinar cells, we studied the protein-protein interaction among those proteins. Immunoblotting analysis showed that parotid acini contain both syntaxin-4 and SNAP-23. However, when VAMP-2 was immunoprecipitated from lysates of parotid acinar cells, syntaxin-4 and SNAP-23 were not coprecipitated with VAMP-2, although syntaxin-1 and SNAP-25 were clearly coprecipitated with VAMP-2 from brain lysates. *ersely, when syntaxin-4 was immunoprecipitated from parotid lysates, SNAP-23, Munc 18c, and NSF were coprecipitated, but VAMP-2 was again undetectable. When proteins in the crude secretory granule fraction were biotinylated and then immunoprecipitated with anti- VAMP-2, some proteins were coprecipitated along with VAMP-2. These results suggest that the interaction between VAMP-2 and syntaxin-4 or SNAP-23 was very weak or hindered by other proteins in parotid acinar cells.