title: Proteomic analysis of the role of S-nitrosoglutathione reductase in lipopolysaccharide-challenged mice.
abstract: S-Nitrosoglutathione reductase (GSNOR) is a key regulator of protein S-nitrosylation, the covalent modification of cysteine residues by nitric oxide that can affect activities of many proteins. We recently discovered that excessive S-nitrosylation from GSNOR deficiency in mice under inflammation inactivates the key DNA repair protein O(6) -alkylguanine-DNA alkyltransferase and promotes both spontaneous and carcinogen-induced hepatocellular carcinoma. To explore further the mechanism of tumorigenesis due to GSNOR deficiency, we compared the protein expression profiles in the livers of wild-type and GSNOR-deficient (GSNOR(-/-) ) mice that were challenged with lipopolysaccharide to induce inflammation and expression of inducible nitric oxide synthase (iNOS). Two-dimensional difference gel electrophoresis analysis identified 38 protein spots of significantly increased intensity and 31 protein spots of significantly decreased intensity in the GSNOR(-/-) mice compared to those in the wild-type mice. We subsequently identified 19 upregulated and 19 downregulated proteins in GSNOR(-/-) mice using mass spectrometry. Immunoblot analysis confirmed in GSNOR(-/-) mice a large increase in the expression of the pro-inflammatory mediator S100A9, a protein previously implicated in human liver carcinogenesis. We also found a decrease in the expression of multiple members of the protein disulfide-isomerase (PDI) family and an alteration in the expression pattern of the endoplasmic reticulum (ER) chaperones in GSNOR(-/-) mice. Furthermore, altered expression of these proteins from GSNOR deficiency was prevented in mice lacking both GSNOR and iNOS. In addition, we detected S-nitrosylation of two members of the PDI protein family. These results suggest that S-nitrosylation resulting from GSNOR deficiency may promote carcinogenesis under inflammatory conditions in part through the disruption of inflammatory and ER stress responses.
authors.affiliation: World-Leading Drug Discovery Research Center, Kyoto University, Kyoto, Japan. k.ozawa@pharm.kyoto-u.ac.jp
authors.forename: Kentaro
authors.initials: K
authors.lastname: Ozawa
authors.forename: Hiroki
authors.initials: H
authors.lastname: Tsumoto
authors.forename: Wei
authors.initials: W
authors.lastname: Wei
authors.forename: Chi-Hui
authors.initials: CH
authors.lastname: Tang
authors.forename: Akira T
authors.initials: AT
authors.lastname: Komatsubara
authors.forename: Hiroto
authors.initials: H
authors.lastname: Kawafune
authors.forename: Kazuharu
authors.initials: K
authors.lastname: Shimizu
authors.forename: Limin
authors.initials: L
authors.lastname: Liu
authors.forename: Gozoh
authors.initials: G
authors.lastname: Tsujimoto
mesh_terms: D000426:Alcohol Dehydrogenase; D000818:Animals; D015153:Blotting, Western; D015180:Electrophoresis, Gel, Two-Dimensional; D059865:Endoplasmic Reticulum Stress / Q000235:genetics; D005260:Female; D005980:Glutathione Reductase / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000633:toxicity*; D008099:Liver / Q000737:chemistry / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D019704:Protein Disulfide-Isomerases / Q000378:metabolism; D020543:Proteome / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D040901:Proteomics
chemical_list: D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D020543:Proteome; C062477:endoplasmic reticulum glycoprotein p72; C483598:Adh5 protein, mouse; D000426:Alcohol Dehydrogenase; D005980:Glutathione Reductase; D019704:Protein Disulfide-Isomerases