OVEREXPRESSION OF RAB3D ENHANCES REGULATED AMYLASE SECRETION FROM PANCREATIC ACINI OF TRANSGENIC MICE

Rab3D, a member of the ras-related GTP-binding protein Rab family, is localized to secretory granules of various exocrine tissues such as ac inar cells of the pancreas, chief cells of the stomach, and parotid an d lacrimal secretory cells. To elucidate the function of Rab3D in exoc ytosis, we have generated transgenic mice that over-express Rab3D spec ifically in pancreatic acinar cells. Hemagglutinin-tagged Rab3D was lo calized to zymogen granules by immunohistochemistry, and was shown to be present on zymogen granule membranes by Western blotting; both resu lts are similar to previous studies of endogenous Rab3D, Secretion mea surements in isolated acinar preparations showed that overexpression o f Rab3D enhanced amylase release. Amylase secretion from intact acini of transgenic mice 5 min after 10 pM cholecystokinin octapeptide (CCK) stimulation was enhanced by 160% of control, In streptolysin-O-permea bilized acini of transgenic mice, amylase secretion induced by 100 mu M GTP-gamma-S was enhanced by 150%, and 10 mu M Ca2+-stimulated amylas e secretion was augmented by 206% of that of the control. To further e lucidate Rab3D involvement in stimulus-secretion coupling, we examined the effect of CCK on the rate of GTP binding to Rab3D, Stimulation of permeabilized acini with 10 pM CCK increased the incorporation of rad iolabeled GTP into HA-tagged Rab3D, These results indicate that overex pression of Rab3D enhances secretagogue-stimulated amylase secretion t hrough both calcium and GTP pathways, We conclude that Rab3D protein o n zymogen granules plays a stimulatory role in regulated amylase secre tion from pancreatic acini.