title: Inhibition of apoptosis signal-regulating kinase 1 by nitric oxide through a thiol redox mechanism.
abstract: Nitric oxide is an endogenous thiol-reactive molecule that modulates the functions of many regulatory proteins by a thiol-redox mechanism. NO has now been shown to inhibit the activation of apoptosis signal-regulating kinase 1 (ASK1) in murine fibrosarcoma L929 cells through such a mechanism. Exposure of L929 cells to interferon-gamma resulted in the endogenous production of NO and in inhibition of the activation of ASK1 by hydrogen peroxide. The interferon-gamma-induced inhibition of ASK1 activity was blocked by N(G)-nitro-l-arginine, an inhibitor of NO synthase. Furthermore, the NO donor S-nitro-N-acetyl-dl-penicillamine (SNAP) inhibited ASK1 activity in vitro, and this inhibition was reversed by thiol-reducing agents such as dithiothreitol and beta-mercaptoethanol. SNAP did not inhibit the kinase activities of MKK3, MKK6, or p38 in vitro. The inhibition of ASK1 by interferon-gamma was not changed by 1H- (1,2,4)oxadiazolo[4,3-alpha]quinoxalin-1-one, an inhibitor of guanylyl cyclase nor was it mimicked by 8-bromo-cyclic GMP. Site-directed mutagenesis revealed that replacement of cysteine 869 of ASK1 by serine rendered this protein resistant to the inhibitory effects both of interferon-gamma in intact cells and of SNAP in vitro. Co-immunoprecipitation data showed that NO production inhibited a binding of ASK1, but not ASK1(C869S), to MKK3 or MKK6. Moreover, interferon-gamma induced the S-nitrosylation of endogenous ASK1 in L929 cells. Together, these results suggest that NO mediates the interferon-gamma-induced inhibition of ASK1 in L929 cells through a thiolredox mechanism.
authors.affiliation: National Creative Research Initiative Center for Cell Death and School of Life Science and Biotechnology, Korea University, Seoul, 136-701, Korea.
authors.forename: Hee-Sae
authors.initials: HS
authors.lastname: Park
authors.forename: Je-Wook
authors.initials: JW
authors.lastname: Yu
authors.forename: Jun-Ho
authors.initials: JH
authors.lastname: Cho
authors.forename: Mi-Sung
authors.initials: MS
authors.lastname: Kim
authors.forename: Sung-Ho
authors.initials: SH
authors.lastname: Huh
authors.forename: Kanghyun
authors.initials: K
authors.lastname: Ryoo
authors.forename: Eui-Ju
authors.initials: EJ
authors.lastname: Choi
mesh_terms: D000818:Animals; D002460:Cell Line; D003545:Cysteine; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005354:Fibrosarcoma; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007371:Interferon-gamma / Q000494:pharmacology; D048848:MAP Kinase Kinase Kinase 5; D020930:MAP Kinase Kinase Kinases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009569:Nitric Oxide / Q000494:pharmacology*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D019335:Nitroarginine / Q000494:pharmacology; D010084:Oxidation-Reduction; D010396:Penicillamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D013329:Structure-Activity Relationship; D013438:Sulfhydryl Compounds / Q000378:metabolism*; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D004791:Enzyme Inhibitors; D020030:Nitric Oxide Donors; C110881:S-nitro-N-acetylpenicillamine; D013438:Sulfhydryl Compounds; D019335:Nitroarginine; D009569:Nitric Oxide; D007371:Interferon-gamma; D006861:Hydrogen Peroxide; D019001:Nitric Oxide Synthase; D048848:MAP Kinase Kinase Kinase 5; D020930:MAP Kinase Kinase Kinases; C482858:MAP3K5 protein, human; C482859:Map3k5 protein, mouse; D010396:Penicillamine; D003545:Cysteine