title: Hsp90 regulates NADPH oxidase activity and is necessary for superoxide but not hydrogen peroxide production.
abstract: The goal of this study was to identify whether heat-shock protein 90 (Hsp90) regulates the production of superoxide and other reactive oxygen species from the NADPH oxidases (Nox). We found that pharmacological and genetic inhibition of Hsp90 directly reduced Nox5-derived superoxide without secondarily modifying signaling events. Coimmunoprecipitation and bioluminescence resonance energy transfer studies suggest that the C-terminus of Nox5 binds to Hsp90. Long-term Hsp90 inhibition reduced Nox5 expression and provides further evidence that Nox5 is an Hsp90 client protein. Inhibitors of Hsp90 also reduced superoxide from Nox1, Nox2 (neutrophils), and Nox3. However, Nox4, which emits only hydrogen peroxide, was unaffected by Hsp90 inhibitors. Hydrogen peroxide production from the other Nox enzymes was not affected by short-term inhibition of Hsp90, but long-term inhibition reduced production of all reactive oxygen species coincident with loss of enzyme expression. Expression of chimeric Nox enzymes consisting of N-terminal Nox1 or Nox3 and C-terminal Nox4 resulted in only hydrogen peroxide formation that was insensitive to Hsp90 inhibitors. We conclude that Hsp90 binds to the C-terminus of Noxes1-3 and 5 and is necessary for enzyme stability and superoxide production. Hsp90 does not bind to the C-terminus of Nox4 and is not required for hydrogen peroxide formation.
authors.affiliation: Department of Forensic Sciences, Xi'an Jiaotong University School of Medicine, Xi'an Shaanxi, P.R. China.
authors.forename: Feng
authors.initials: F
authors.lastname: Chen
authors.forename: Deepesh
authors.initials: D
authors.lastname: Pandey
authors.forename: Ahmed
authors.initials: A
authors.lastname: Chadli
authors.forename: John D
authors.initials: JD
authors.lastname: Catravas
authors.forename: Teng
authors.initials: T
authors.lastname: Chen
authors.forename: David J R
authors.initials: DJ
authors.lastname: Fulton
mesh_terms: D000818:Animals; D016227:Benzoquinones / Q000494:pharmacology; D019556:COS Cells; D002522:Chlorocebus aethiops; D057075:Enzyme Assays; D057809:HEK293 Cells; D018841:HSP90 Heat-Shock Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D047468:Immunoprecipitation; D047029:Lactams, Macrocyclic / Q000494:pharmacology; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism; D000074664:NADPH Oxidase 5; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism*
chemical_list: D016227:Benzoquinones; D018841:HSP90 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D008565:Membrane Proteins; D011994:Recombinant Proteins; D013481:Superoxides; C112765:tanespimycin; D006861:Hydrogen Peroxide; D000074664:NADPH Oxidase 5; D019255:NADPH Oxidases; C431571:NOX5 protein, human