title: Alpha-SNAP functions in insulin exocytosis from mature, but not immature secretory granules in pancreatic beta cells.
abstract: To explore alpha-SNAP function in insulin exocytosis from either immature or mature secretory granules in pancreatic beta cells, we studied the effects of overexpression of adenovirus-mediated wild-type alpha-SNAP and C-terminally deleted alpha-SNAP mutant (1-285) on newly synthesized proinsulin and insulin release by rat islets and MIN6 cells. Rat islets overexpressing alpha-SNAP and mutant alpha-SNAP were pulse-chased. Exocytosis from immature and mature insulin secretory granules was measured as fractional (%) labeled-proinsulin release immediately after the pulse-labeling and percentage labeled-insulin release after a 3-h chase period, respectively. There was no difference in percentage labeled-proinsulin release between the control and alpha-SNAP or mutant alpha-SNAP-overexpressed islets. Although percentage labeled-insulin release after a 3-h chase period was significantly increased in alpha-SNAP-overexpressed islets, it was decreased in mutant alpha-SNAP-overexpressed islets. Thus, the results demonstrated that alpha-SNAP overexpression in rat islets primarily increased exocytosis from mature, but not immature insulin secretory granules. On the other hand, in MIN6 cells, alpha-SNAP overexpression scarcely affected glucose-stimulated insulin release; therefore, we examined the effect of mutant alpha-SNAP overexpression as the dominant-negative inhibitor on the newly synthesized proinsulin/insulin release using the same protocol as in the rat islet experiments. alpha-SNAP mutant (1-285) overexpression in MIN6 cells decreased the percentage labeled insulin release from mature secretory granules, but not percentage labeled proinsulin release from immature secretory granules. Thus, our data demonstrate that alpha-SNAP functions mainly in the mature insulin secretory granules in pancreatic beta cells.
authors.affiliation: Department of Biochemistry, Kyorin University School of Medicine, Mitaka, Tokyo, 181-8611, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Nakamichi
authors.forename: S
authors.initials: S
authors.lastname: Nagamatsu
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D002352:Carrier Proteins / Q000502:physiology*; D002460:Cell Line; D003594:Cytoplasmic Granules / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005089:Exocytosis; D005947:Glucose / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism*; D007515:Islets of Langerhans / Q000378:metabolism* / Q000648:ultrastructure; D008297:Male; D008565:Membrane Proteins / Q000502:physiology*; D051381:Rats; D017208:Rats, Wistar; D050600:SNARE Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D033921:Vesicular Transport Proteins
chemical_list: D002352:Carrier Proteins; D007328:Insulin; D008565:Membrane Proteins; D050600:SNARE Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D033921:Vesicular Transport Proteins; D005947:Glucose