title: Phosphorylation of SNAP-25 on serine-187 is induced by secretagogues in insulin-secreting cells, but is not correlated with insulin secretion.
abstract: The tSNARE (the target-membrane soluble NSF-attachment protein receptor, where NSF is N -ethylmaleimide-sensitive fusion protein) synaptosomal-associated protein of 25 kDa (SNAP-25) is implicated in regulated insulin secretion. In pheochromocytoma PC12 cells, SNAP-25 is phosphorylated at Ser(187), which lies in a region that is important for its function. The aims of the present study were to determine whether SNAP-25 is phosphorylated at Ser(187) in insulin-secreting cells and, if so, whether this is important for regulated insulin secretion. The major findings are: (i) SNAP-25 is rapidly and reversibly phosphorylated on Ser(187) in both rat insulinoma INS-1 cells and rat islets in response to the phorbol ester, PMA; (ii) less than 35% of SNAP-25 in INS-1 cells is phosphorylated in response to PMA, and phosphorylation is limited to plasma-membrane-associated SNAP-25; (iii) both SNAP-25 isoforms (a and b) are phosphorylated, with 1.8-fold greater phosphorylation for SNAP-25b in response to PMA; (iv) in rat islets, Ser(187) phosphorylation is stimulated by glucose or carbachol, albeit to a lesser extent than by PMA, but not by cAMP; (v) insulin secretion from botulinum neurotoxin E-treated hamster insulinoma tumour (HIT) cells, transfected with toxin-resistant Ser(187)-->Ala or Ser(187)-->Asp mutant SNAP-25, was similar to that of wild-type HIT cells. Furthermore, in rat islets no correlation was found between the extent of SNAP-25 phosphorylation at Ser(187) in response to secretagogues and stimulation of insulin release; (vi) use of protein kinase C (PKC) inhibitors suggests that glucose stimulates SNAP-25 phosphorylation via conventional and non-conventional PKC isoforms. In summary, although SNAP-25 phosphorylation at Ser(187) occurs in insulin-secreting cells and is mediated by PKC, it does not appear to play a major role in regulated insulin secretion.
authors.affiliation: Laboratoires de Recherche Louis Jeantet, Centre Médical Universitaire, rue Michel Servet, CH-1211 Geneva 4, Switzerland. carmen.gonelle@medecine.unige.ch
authors.forename: Carmen
authors.initials: C
authors.lastname: Gonelle-Gispert
authors.forename: Maria
authors.initials: M
authors.lastname: Costa
authors.forename: Masami
authors.initials: M
authors.lastname: Takahashi
authors.forename: Karin
authors.initials: K
authors.lastname: Sadoul
authors.forename: Philippe
authors.initials: P
authors.lastname: Halban
mesh_terms: D000818:Animals; D001905:Botulinum Toxins / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D004789:Enzyme Activation; D005089:Exocytosis; D005947:Glucose / Q000378:metabolism; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism*; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D016716:PC12 Cells; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D050600:SNARE Proteins; D012694:Serine / Q000378:metabolism*; D050825:Synaptosomal-Associated Protein 25; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D033921:Vesicular Transport Proteins
chemical_list: D007328:Insulin; D007527:Isoenzymes; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D050600:SNARE Proteins; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; D033921:Vesicular Transport Proteins; D012694:Serine; D002217:Carbachol; D011493:Protein Kinase C; D001905:Botulinum Toxins; D005947:Glucose; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; C096326:botulinum toxin type E