Synaptotagmin III/VII isoforms mediate Ca2+-induced insulin secretion in pancreatic islet beta -cells.

Synaptotagmins (Syt) play important roles in Ca(2+)-induced neuroexocytosis. Insulin secretion of the pancreatic beta-cell is dependent on an increase in intracellular Ca(2+); however, Syt involvement in insulin exocytosis is poorly understood. Reverse transcriptase-polymerase chain reaction studies showed the presence of Syt isoforms III, IV, V, and VII in rat pancreatic islets, whereas Syt isoforms I, II, III, IV, V, VII, and VIII were present in insulin-secreting betaTC3 cell. Syt III and VII proteins were identified in rat islets and betaTC3 and RINm5F beta-cells by immunoblotting. Confocal microscopy showed that Syt III and VII co-localized with insulin-containing secretory granules. Two-fold overexpression of Syt III in RINm5F beta-cell (Syt III cell) was achieved by stable transfection, which conferred greater Ca(2+) sensitivity for exocytosis, and resulted in increased insulin secretion. Glyceraldehyde + carbachol-induced insulin secretion in Syt III cells was 2.5-fold higher than control empty vector cells, whereas potassium-induced secretion was 6-fold higher. In permeabilized Syt III cells, Ca(2+)-induced and mastoparan-induced insulin secretion was also increased. In Syt VII-overexpressing RINm5F beta-cells, there was amplification of carbachol-induced insulin secretion in intact cells and of Ca(2+)-induced and mastoparan-induced insulin secretion in permeabilized cells. In conclusion, Syt III/VII are located in insulin-containing secretory granules, and we suggest that Syt III/VII may be the Ca(2+) sensor or one of the Ca(2+) sensors for insulin exocytosis of the beta-cell.