Subcellular distribution and function of Rab3A-D in pancreatic acinar AR42J cells

Members of the Rab3 subfamily have been linked to the regulation of exocyto sis in secretory cells. We have recently shown by Northern blot analysis th at pancreatic acinar-like AR42J cells express all four Rab3 isoforms (Rab3A -D). In the present study, we examined the subcellular distribution of endo genously expressed Rab3 proteins and their relation to the amylase-containi ng secretory compartment in dexamethasone-differentiated AR42J cells. Rab3A and Rab3C were enriched in the cytosol, Rab3B and Rab3D in the membrane fr action. Accordingly, confocal immunocytochemistry revealed that Rab3B and R ab3D were located in a compartment close to the plasma membrane, whereas an ti-Rab3A and Rab3C mainly stained the cytosol. Sucrose density gradient cen trifugation showed overlapping, but distinct localization of each Rab3 isof orm. The order of banding from lighter to more dense fractions was Rab3C < Rab3A < Rab3B < Rab3D. All Rab3 proteins at least partially colocalized wit h amylase immunoreactivity. Transient overexpression of Rab3 proteins showe d that Rab3A inhibited cholecystokinin (CCK)-induced amylase secretion, whe reas overexpression of other Rab3 isoforms had no significant effect. In co nclusion, our data indicate that the different Rab3 proteins show distinct subcellular distribution, suggesting different impact on exocrine secretory response in dexamethasone-differentiated AR42J cells. (C) 2001 Academic Pr ess.