title: S-nitrosylation of syntaxin 1 at Cys(145) is a regulatory switch controlling Munc18-1 binding.
abstract: Exocytosis is regulated by NO in many cell types, including neurons. In the present study we show that syntaxin 1a is a substrate for S-nitrosylation and that NO disrupts the binding of Munc18-1 to the closed conformation of syntaxin 1a in vitro. In contrast, NO does not inhibit SNARE {SNAP [soluble NSF (N-ethylmaleimide-sensitive fusion protein) attachment protein] receptor} complex formation or binding of Munc18-1 to the SNARE complex. Cys(145) of syntaxin 1a is the target of NO, as a non-nitrosylatable C145S mutant is resistant to NO and novel nitrosomimetic Cys(145) mutants mimic the effect of NO on Munc18-1 binding in vitro. Furthermore, expression of nitrosomimetic syntaxin 1a in living cells affects Munc18-1 localization and alters exocytosis release kinetics and quantal size. Molecular dynamic simulations suggest that NO regulates the syntaxin-Munc18 interaction by local rearrangement of the syntaxin linker and H3c regions. Thus S-nitrosylation of Cys(145) may be a molecular switch to disrupt Munc18-1 binding to the closed conformation of syntaxin 1a, thereby facilitating its engagement with the membrane fusion machinery.
authors.affiliation: The Physiological Laboratory, School of Biomedical Sciences, University of Liverpool, Crown St, Liverpool L69 3BX, U.K.
authors.forename: Zoë J
authors.initials: ZJ
authors.lastname: Palmer
authors.forename: Rory R
authors.initials: RR
authors.lastname: Duncan
authors.forename: James R
authors.initials: JR
authors.lastname: Johnson
authors.forename: Lu-Yun
authors.initials: LY
authors.lastname: Lian
authors.forename: Luciane V
authors.initials: LV
authors.lastname: Mello
authors.forename: David
authors.initials: D
authors.lastname: Booth
authors.forename: Jeff W
authors.initials: JW
authors.lastname: Barclay
authors.forename: Margaret E
authors.initials: ME
authors.lastname: Graham
authors.forename: Robert D
authors.initials: RD
authors.lastname: Burgoyne
authors.forename: Ian A
authors.initials: IA
authors.lastname: Prior
authors.forename: Alan
authors.initials: A
authors.lastname: Morgan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003198:Computer Simulation; D003545:Cysteine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005089:Exocytosis; D006367:HeLa Cells; D006801:Humans; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D051938:Munc18 Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding; D050600:SNARE Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D050827:Syntaxin 1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013816:Thermodynamics
chemical_list: D051938:Munc18 Proteins; D050600:SNARE Proteins; D050827:Syntaxin 1; D009569:Nitric Oxide; D003545:Cysteine