title: Phosphorylation of Nox1 regulates association with NoxA1 activation domain.
abstract: BACKGROUND
  Activation of Nox1 initiates redox-dependent signaling events crucial in the pathogenesis of vascular disease. Selective targeting of Nox1 is an attractive potential therapy, but requires a better understanding of the molecular modifications controlling its activation.
  
  
  OBJECTIVE
  To determine whether posttranslational modifications of Nox1 regulate its activity in vascular cells.
  
  
  RESULTS
  We first found evidence that Nox1 is phosphorylated in multiple models of vascular disease. Next, studies using mass spectroscopy and a pharmacological inhibitor demonstrated that protein kinase C-beta1 mediates phosphorylation of Nox1 in response to tumor necrosis factor-α. siRNA-mediated silencing of protein kinase C-beta1 abolished tumor necrosis factor-α-mediated reactive oxygen species production and vascular smooth muscle cell migration. Site-directed mutagenesis and isothermal titration calorimetry indicated that protein kinase C-beta1 phosphorylates Nox1 at threonine 429. Moreover, Nox1 threonine 429 phosphorylation facilitated the association of Nox1 with the NoxA1 activation domain and was necessary for NADPH oxidase complex assembly, reactive oxygen species production, and vascular smooth muscle cell migration.
  
  
  CONCLUSIONS
  We conclude that protein kinase C-beta1 phosphorylation of threonine 429 regulates activation of Nox1 NADPH oxidase.
authors.affiliation: From the Departments of Internal Medicine (B.M.S., S.J., B.S., W.H.T., F.J.M.), Microbiology (J.C.D.H.), Anatomy and Cell Biology (J.S.), Biochemistry (L.G.), and Protein Crystallography Facility (L.G.), University of Iowa, Iowa City; and Veterans Affair Medical Center, Iowa City, IA (F.J.M.).
authors.forename: Jennifer
authors.initials: J
authors.lastname: Streeter
authors.affiliation: From the Departments of Internal Medicine (B.M.S., S.J., B.S., W.H.T., F.J.M.), Microbiology (J.C.D.H.), Anatomy and Cell Biology (J.S.), Biochemistry (L.G.), and Protein Crystallography Facility (L.G.), University of Iowa, Iowa City; and Veterans Affair Medical Center, Iowa City, IA (F.J.M.).
authors.forename: Brandon M
authors.initials: BM
authors.lastname: Schickling
authors.affiliation: From the Departments of Internal Medicine (B.M.S., S.J., B.S., W.H.T., F.J.M.), Microbiology (J.C.D.H.), Anatomy and Cell Biology (J.S.), Biochemistry (L.G.), and Protein Crystallography Facility (L.G.), University of Iowa, Iowa City; and Veterans Affair Medical Center, Iowa City, IA (F.J.M.).
authors.forename: Shuxia
authors.initials: S
authors.lastname: Jiang
authors.affiliation: From the Departments of Internal Medicine (B.M.S., S.J., B.S., W.H.T., F.J.M.), Microbiology (J.C.D.H.), Anatomy and Cell Biology (J.S.), Biochemistry (L.G.), and Protein Crystallography Facility (L.G.), University of Iowa, Iowa City; and Veterans Affair Medical Center, Iowa City, IA (F.J.M.).
authors.forename: Bojana
authors.initials: B
authors.lastname: Stanic
authors.affiliation: From the Departments of Internal Medicine (B.M.S., S.J., B.S., W.H.T., F.J.M.), Microbiology (J.C.D.H.), Anatomy and Cell Biology (J.S.), Biochemistry (L.G.), and Protein Crystallography Facility (L.G.), University of Iowa, Iowa City; and Veterans Affair Medical Center, Iowa City, IA (F.J.M.).
authors.forename: William H
authors.initials: WH
authors.lastname: Thiel
authors.affiliation: From the Departments of Internal Medicine (B.M.S., S.J., B.S., W.H.T., F.J.M.), Microbiology (J.C.D.H.), Anatomy and Cell Biology (J.S.), Biochemistry (L.G.), and Protein Crystallography Facility (L.G.), University of Iowa, Iowa City; and Veterans Affair Medical Center, Iowa City, IA (F.J.M.).
authors.forename: Lokesh
authors.initials: L
authors.lastname: Gakhar
authors.affiliation: From the Departments of Internal Medicine (B.M.S., S.J., B.S., W.H.T., F.J.M.), Microbiology (J.C.D.H.), Anatomy and Cell Biology (J.S.), Biochemistry (L.G.), and Protein Crystallography Facility (L.G.), University of Iowa, Iowa City; and Veterans Affair Medical Center, Iowa City, IA (F.J.M.).
authors.forename: Jon C D
authors.initials: JC
authors.lastname: Houtman
authors.affiliation: From the Departments of Internal Medicine (B.M.S., S.J., B.S., W.H.T., F.J.M.), Microbiology (J.C.D.H.), Anatomy and Cell Biology (J.S.), Biochemistry (L.G.), and Protein Crystallography Facility (L.G.), University of Iowa, Iowa City; and Veterans Affair Medical Center, Iowa City, IA (F.J.M.). francis-miller@uiowa.edu.
authors.forename: Francis J
authors.initials: FJ
authors.lastname: Miller
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001011:Aorta / Q000166:cytology; D001665:Binding Sites; D002465:Cell Movement; D002478:Cells, Cultured; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism; D009154:Mutation; D032389:Myocytes, Smooth Muscle / Q000378:metabolism / Q000502:physiology; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000074624:NADPH Oxidase 1; D010766:Phosphorylation; D011485:Protein Binding; D064546:Protein Kinase C beta / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C472286:Noxa1 protein, mouse; D011506:Proteins; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D009247:NADH, NADPH Oxidoreductases; D000074624:NADPH Oxidase 1; C000614711:NOX1 protein, mouse; D064546:Protein Kinase C beta
keywords: NADPH oxidase; PKCβ; TNF-α; atherosclerosis; neointima; vascular smooth muscle cells