title: Cholecystokinin-regulated exocytosis in rat pancreatic acinar cells is inhibited by a C-terminus truncated mutant of SNAP-23.
abstract: BACKGROUND
  Exocytosis is thought to result from the fusion of vesicle and plasma membranes caused by the formation of a trans-complex between proteins of the vesicle-associated membrane protein (VAMP) family on the vesicle with members of the syntaxin and synaptosomal-associated protein of 25 kd (SNAP-25) families on the plasma membrane. In the pancreatic acinar cell, synaptosomal-associated protein of 23 kd (SNAP-23) is the major SNAP-25 isoform expressed in pancreatic acinar cells, but its role in acinar cell exocytosis has not been determined.
  
  
  OBJECTIVE
  To examine the role of SNAP-23 in regulated exocytosis in acinar cells, we subcloned into adenoviral vectors SNAP-23, SNAP-25, and dominant negative mutants in which the C-terminal domains corresponding to the botulinum neurotoxin A cleavage sites are deleted.
  
  
  RESULTS
  High-efficiency infection of rat pancreatic acini in culture with these adenoviruses by subcellular fractionation showed that the overexpressed SNAP-23, SNAP-25, and their truncated mutant proteins were uniformly targeted to the zymogen granules and plasma membrane. To maximally stimulate apical exocytosis from these infected acini, we used the cholecystokinin-phenylethyl ester analog (CCK-OPE), which does not show inhibition of secretion from maximal levels at high doses. CCK-OPE-stimulated amylase release from adenovirus-cytomegalovirus (AdCMV)-SNAP-23 or AdCMV-SNAP-25-infected acini to the same extent as from acini infected with the empty vector. In contrast, CCK-OPE-evoked enzyme secretion from AdCMV-SNAP-23deltaC8- and AdCMV-SNAP-25(1-197)-infected acini were inhibited by 60% and 40%, respectively. The identical targeting of the mutant SNAP-23 and SNAP-25 proteins to the same membrane compartments as SNAP-23 suggests that the inhibition of secretion was a result of their competition against endogenous SNAP-23. This is supported by the fact that this inhibition by the mutant proteins was partially reversed or rescued when the AdCMV-SNAP-25AC8- or AdCMV-SNAP-25(1-197)-infected acini were co-infected with wild-type SNAP-23 or SNAP-25.
  
  
  CONCLUSIONS
  From these results, we conclude that SNAP-23 plays a role in CCK-evoked regulated exocytosis in the acinar cells.
authors.affiliation: Department of Medicine and Physiology, University of Toronto, Ontario, Canada.
authors.forename: X
authors.initials: X
authors.lastname: Huang
authors.forename: L
authors.initials: L
authors.lastname: Sheu
authors.forename: Y
authors.initials: Y
authors.lastname: Tamori
authors.forename: W S
authors.initials: WS
authors.lastname: Trimble
authors.forename: H Y
authors.initials: HY
authors.lastname: Gaisano
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000681:Amylases / Q000378:metabolism; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics* / Q000502:physiology*; D002766:Cholecystokinin / Q000502:physiology*; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000502:physiology; D010179:Pancreas / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D010446:Peptide Fragments / Q000494:pharmacology; D050766:Qb-SNARE Proteins; D050767:Qc-SNARE Proteins; D051381:Rats; D017207:Rats, Sprague-Dawley; D017384:Sequence Deletion; D012844:Sincalide / Q000031:analogs & derivatives* / Q000494:pharmacology; D050825:Synaptosomal-Associated Protein 25
chemical_list: D002352:Carrier Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D050766:Qb-SNARE Proteins; D050767:Qc-SNARE Proteins; C100493:SNAP23 protein, human; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; C067408:cholecystokinin (26-32), Tyr-Gly-Nle(28,31) phenethyl ester-; D002766:Cholecystokinin; D000681:Amylases; D012844:Sincalide