α-SNAP Functions in Insulin Exocytosis from Mature, but Not Immature Secretory Granules in Pancreatic β Cells

Abstract

To explore α-SNAP function in insulin exocytosis from either immature or mature secretory granules in pancreatic β cells, we studied the effects of overexpression of adenovirus-mediated wild-type α-SNAP and C-terminally deleted α-SNAP mutant (1–285) on newly synthesized proinsulin and insulin release by rat islets and MIN6 cells. Rat islets overexpressing α-SNAP and mutant α-SNAP were pulse-chased. Exocytosis from immature and mature insulin secretory granules was measured as fractional (%) labeled-proinsulin release immediately after the pulse-labeling and percentage labeled-insulin release after a 3-h chase period, respectively. There was no difference in percentage labeled-proinsulin release between the control and α-SNAP or mutant α-SNAP-overexpressed islets. Although percentage labeled-insulin release after a 3-h chase period was significantly increased in α-SNAP-overexpressed islets, it was decreased in mutant α-SNAP-overexpressed islets. Thus, the results demonstrated that α-SNAP overexpression in rat islets primarily increased exocytosis from mature, but not immature insulin secretory granules. On the other hand, in MIN6 cells, α-SNAP overexpression scarcely affected glucose-stimulated insulin release; therefore, we examined the effect of mutant α-SNAP overexpression as the dominant-negative inhibitor on the newly synthesized proinsulin/insulin release using the same protocol as in the rat islet experiments. α-SNAP mutant (1–285) overexpression in MIN6 cells decreased the percentage labeled insulin release from mature secretory granules, but not percentage labeled proinsulin release from immature secretory granules. Thus, our data demonstrate that α-SNAP functions mainly in the mature insulin secretory granules in pancreatic β cells.