title: Glutathione S-transferase P1 suppresses iNOS protein stability in RAW264.7 macrophage-like cells after LPS stimulation.
abstract: Glutathione S-transferase P1 (GSTP1) is a ubiquitous expressed protein which plays an important role in the detoxification and xenobiotics metabolism. Previous studies showed that GSTP1 was upregulated by the LPS stimulation in RAW264.7 macrophage-like cells and GSTP1 overexpression downregulated lipopolysaccharide (LPS) induced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression. Here we show that GSTP1 physically associates with the oxygenase domain of iNOS by the G-site domain and decreases the protein level of iNOS dimer. Both overexpression and RNA interference (RNAi) experiments indicate that GSTP1 downregulates iNOS protein level and increases S-nitrosylation and ubiquitination of iNOS. The Y7F mutant type of GSTP1 physically associates with iNOS, but shows no effect on iNOS protein content, iNOS S-nitrosylation, and changes in iNOS from dimer to monomer, suggesting the importance of enzyme activity of GSTP1 in regulating iNOS S-nitrosylation and stability. GSTM1, another member of GSTs shows no significant effect on regulation of iNOS. In conclusion, our study reveals the novel role of GSTP1 in regulation of iNOS by affecting S-nitrosylation, dimerization, and stability, which provides a new insight for analyzing the regulation of iNOS and the anti-inflammatory effects of GSTP1.
authors.affiliation: a Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, College of Life Science, Nanjing Normal University , Nanjing, 210046, Jiangsu , P R China.
authors.forename: Xiang
authors.initials: X
authors.lastname: Cao
authors.affiliation: a Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, College of Life Science, Nanjing Normal University , Nanjing, 210046, Jiangsu , P R China.
authors.forename: Xiuqin
authors.initials: X
authors.lastname: Kong
authors.affiliation: a Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, College of Life Science, Nanjing Normal University , Nanjing, 210046, Jiangsu , P R China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: a Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, College of Life Science, Nanjing Normal University , Nanjing, 210046, Jiangsu , P R China.
authors.forename: Lei
authors.initials: L
authors.lastname: Lan
authors.affiliation: b State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University , Nanjing 210093, Jiangsu , P R China.
authors.forename: Lan
authors.initials: L
authors.lastname: Luo
authors.affiliation: a Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, College of Life Science, Nanjing Normal University , Nanjing, 210046, Jiangsu , P R China.
authors.forename: Zhimin
authors.initials: Z
authors.lastname: Yin
mesh_terms: D000818:Animals; D051549:Glutathione S-Transferase pi / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D055550:Protein Stability; D000067996:RAW 264.7 Cells; D034741:RNA, Small Interfering; D060888:Real-Time Polymerase Chain Reaction; D014162:Transfection
chemical_list: D008070:Lipopolysaccharides; D034741:RNA, Small Interfering; D052247:Nitric Oxide Synthase Type II; D051549:Glutathione S-Transferase pi
keywords: S-nitrosylation; dimerization; glutathione S-transferase P1 (GSTP1); inducible nitric oxide synthase (iNOS); ubiquitination