VAMP‐Specific Zymogen Granules (ZG) Differentially Regulate Basal and Ca2+‐Stimulated Secretory Pathways in Pancreatic Acinar Cells

SNARE proteins are the minimal essential machinery for membrane fusion events. In this study, we investigated the SNARE complexes in pancreatic acinar cells that regulate digestive enzyme secretion. We previously reported that two isoforms of VAMPs: VAMP 2 and VAMP 8, as well as SNAP 23 and SNAP 29 are expressed on ZG membranes. VAMP 8 strongly interacts with SNAP 29, however, VAMP 2 was found to specifically complex with SNAP 23, but not with SNAP 29. Significantly, quantitative immunofluorescence of intact acini and purified ZGs showed minimal overlap between VAMP 2 and VAMP 8, indicating there are likely different populations of ZGs. CoIP and GST‐pull‐down assays were conducted to identify complete SNARE complexes in acinar cells. Syntaxin (SYN) 4 is present on both acinar basolateral and apical membrane and interacts with VAMP 8 and SNAP 29 following cholecystokinin (CCK)‐stimulation of cells. VAMP 2 was found to interact with SYN 2, a SNARE present exclusively at the apical membrane. Finally, permeabilized acinar cells incubated with recombinant cytosolic VAMP 2 or 8 showed a 50% maximum decrease in Ca 2+ ‐stimulated exocytosis, whereas VAMP 1 and VAMP 1 truncated peptides showed little or no effects. Incubation of acini with cytosolic construct of SYN 4 induced a 50% decrease in stimulated exocytosis, whereas SYN 2 and 3 only had marginal effects. Both SYN 2 and 4 each significantly inhibited basal secretion. These data suggest there are different VAMP 2 and 8 carrying populations of ZGs in acinar cells. SYN 4 is the major isoform regulating stimulated secretion, whereas SYN 2 and 4 modulate basal secretion, likely through their interactions with VAMP‐specific populations of ZGs.