Distinct cellular locations of the syntaxin family of proteins in rat pancreatic acinar cells.

Abstract

Syntaxins are cytoplasmically oriented integral membrane soluble NEM-sensitive factor receptors (SNAREs; soluble NEM-sensitive factor attachment protein receptors) thought to serve as targets for the assembly of protein complexes important in regulating membrane fusion. The SNARE hypothesis predicts that the fidelity of vesicle traffic is controlled in part by the correct recognition of vesicle SNAREs with their cognate target SNARE partner. Here, we show that in the exocrine acinar cell of the pancreas, multiple syntaxin isoforms are expressed and that they appear to reside in distinct membrane compartments. Syntaxin 2 is restricted to the apical plasma membrane whereas syntaxin 4 is found most abundantly on the basolateral membranes. Surprisingly, syntaxin 3 was found to be localized to a vesicular compartment, the zymogen granule membrane. In addition, we show that these proteins are capable of specific interaction with vesicle SNARE proteins. Their nonoverlapping locations support the general principle of the SNARE hypothesis and provide new insights into the mechanisms of polarized secretion in epithelial cells. INTRODUCTIONThe SNARE hypothesis is a general model proposed by Rothman and Warren (1994) to explain the basic principles of membrane fusion responsible for intracellular membrane movements and secretion . This hypothesis predicts that a stepwise progression of interactions between soluble cytoplasmic proteins and membrane receptors ultimately leads to the fusion of the two membranes. The soluble factors, including the ATPase NEM-sensitive factor (NSF) and soluble NSF attachment proteins (SNAPs), are thought to bind to SNAP receptors (SNAREs), located on transport vesicles (v-SNAREs) and on target membranes (t-SNAREs), to mediate the fusion of the two membranes. A stable complex be-

Differential localization of syntaxin isoforms in polarized Madin-Darby canine kidney cells.

“…Alternatively, syntaxin 3 may participate in fusion among intracellular vesicles. Consistent with this possibility, syntaxin 3 (but not coexpressed syntaxin 2 or 4) has been localized to zymogen granules in pancreatic acinar cells (Gaisano et al, 1996), recycling tubulovesicles in gastric parietal cells (Bennett, manuscript in preparation) and secretory granules in eosinophils (Bennett, unpublished data). Recent data indicate that under certain circumstances, NSF acts solely on transport vesicles, prior to docking or fusion of these vesicles with their target (Mayer et al, 1996;Steel et al, 1996).…”

“…All syntaxin antibodies were affinity purified before use. All syntaxin antibodies were isotype specific, except for the syntaxin 2 antibody which showed a weak crossreactivity with syntaxin 1 by Western blot (Gaisano et al, 1996). Fluorescein isothiocyanate-or Texas Red-conjugated secondary antibodies were purchased from Jackson ImmunoResearch Laboratories (West Grove, PA).…”

Syntaxin-3 regulates newcomer insulin granule exocytosis and compound fusion in pancreatic beta cells

“…Syn-3-overexpressing cells showed that the number of no-dock newcomer SGs undergoing exocytosis increased by 98% and 164% in first (17.13±3.62 vs control 8.66±1. 19) and second phases (24.60±5.92 vs control 9.32 ±2.03), respectively. Short-dock newcomer SGs in Syn-3-overexpressing cells were increased, but mainly in the first phase (6.13±0.61 vs control 2.45±0.45) (Fig.…”

“…In retinal cells, SYN-3 is produced in ribbon synapses but its role in neurotransmitter release remains unknown [16,17]. In pancreatic acinar cells [18], SYN-3 was localised to zymogen granule membrane [19] and complexed with VAMP8 to mediate SG-SG fusion [20].…”

Supramaximal cholecystokinin displaces Munc18c from the pancreatic acinar basal surface, redirecting apical exocytosis to the basal membrane

“…In vitro studies of recombinant proteins indeed demonstrated stable binding of VAMP-2, SNAP-23, and syntaxin-4 (22), raising the possibility that a complex of VAMP-2/ SNAP-23/syntaxin-4 could mediate basolateral exocytosis of digestive enzymes within the pancreas. Non-neuronal Munc18 isoforms were also identified, including Munc18b and Munc18c (21)(22)(23)(24)(25)(26). Munc18c binds to syntaxin-4 and blocks its binding to SNAP-23 and VAMP-2 (24,25), and overexpression of Munc18c in adipocytes inhibits insulin-mediated GLUT-4 translocation to the plasma membrane and glucose transport (26).…”