title: Tomosyn-1 is involved in a post-docking event required for pancreatic beta-cell exocytosis.
abstract: Although the assembly of a ternary complex between the SNARE proteins syntaxin-1, SNAP25 and VAMP2 is known to be crucial for insulin exocytosis, the mechanisms controlling this key event are poorly understood. We found that pancreatic beta-cells express different isoforms of tomosyn-1, a syntaxin-1-binding protein possessing a SNARE-like motif. Using atomic force microscopy we show that the SNARE-like domain of tomosyn-1 can form a complex with syntaxin-1 and SNAP25 but displays binding forces that are weaker than those observed for VAMP2 (237+/-13 versus 279+/-3 pN). In pancreatic beta-cells tomosyn-1 was found to be concentrated in cellular compartments enriched in insulin-containing secretory granules. Silencing of tomosyn-1 in the rat beta-cell line INS-1E by RNA interference did not affect the number of secretory granules docked at the plasma membrane but led to a reduction in stimulus-induced exocytosis. Replacement of endogenous tomosyn-1 with mouse tomosyn-1, which differs in the nucleotide sequence from its rat homologue and escapes silencing, restored a normal secretory rate. Taken together, our data suggest that tomosyn-1 is involved in a post-docking event that prepares secretory granules for fusion and is necessary to sustain exocytosis of pancreatic beta-cells in response to insulin secretagogues.
authors.affiliation: Department of Cell Biology and Morphology, Rue du Bugnon 9, University of Lausanne, 1005 Lausanne, Switzerland.
authors.forename: Séverine
authors.initials: S
authors.lastname: Cheviet
authors.forename: Paola
authors.initials: P
authors.lastname: Bezzi
authors.forename: Rosita
authors.initials: R
authors.lastname: Ivarsson
authors.forename: Erik
authors.initials: E
authors.lastname: Renström
authors.forename: David
authors.initials: D
authors.lastname: Viertl
authors.forename: Sandor
authors.initials: S
authors.lastname: Kasas
authors.forename: Stefan
authors.initials: S
authors.lastname: Catsicas
authors.forename: Romano
authors.initials: R
authors.lastname: Regazzi
mesh_terms: D000818:Animals; D019556:COS Cells; D020013:Calcium Signaling; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D015536:Down-Regulation / Q000235:genetics; D005089:Exocytosis; D020868:Gene Silencing; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000166:cytology* / Q000378:metabolism*; D051379:Mice; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D050683:R-SNARE Proteins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D022142:Secretory Vesicles / Q000378:metabolism; D050825:Synaptosomal-Associated Protein 25 / Q000378:metabolism; D050827:Syntaxin 1 / Q000378:metabolism; D013816:Thermodynamics; D015854:Up-Regulation / Q000235:genetics
chemical_list: D007328:Insulin; D009419:Nerve Tissue Proteins; D050683:R-SNARE Proteins; D012333:RNA, Messenger; C484769:Stxbp5 protein, rat; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; C112618:tomosyn protein, mouse