title: IkappaB is a sensitive target for oxidation by cell-permeable chloramines: inhibition of NF-kappaB activity by glycine chloramine through methionine oxidation.
abstract: Hypochlorous acid (HOCl) is produced by the neutrophil enzyme, myeloperoxidase, and reacts with amines to generate chloramines. These oxidants react readily with thiols and methionine and can affect cell-regulatory pathways. In the present study, we have investigated the ability of HOCl, glycine chloramine (Gly-Cl) and taurine chloramine (Tau-Cl) to oxidize IkappaBalpha, the inhibitor of NF-kappaB (nuclear factor kappaB), and to prevent activation of the NF-kappaB pathway in Jurkat cells. Glycine chloramine (Gly-Cl) and HOCl were permeable to the cells as determined by oxidation of intracellular GSH and inactivation of glyceraldehyde-3-phosphate dehydrogenase, whereas Tau-Cl showed no detectable cell permeability. Both Gly-Cl (20-200 muM) and HOCl (50 microM) caused oxidation of IkappaBalpha methionine, measured by a shift in electrophoretic mobility, when added to the cells in Hanks buffer. In contrast, a high concentration of Tau-Cl (1 mM) in Hanks buffer had no effect. However, Tau-Cl in full medium did modify IkappaBalpha. This we attribute to chlorine exchange with other amines in the medium to form more permeable chloramines. Oxidation by Gly-Cl prevented IkappaBalpha degradation in cells treated with TNFalpha (tumour necrosis factor alpha) and inhibited nuclear translocation of NF-kappaB. IkappaBalpha modification was reversed by methionine sulphoxide reductase, with both A and B forms required for complete reduction. Oxidized IkappaBalpha persisted intracellularly for up to 6 h. Reversion occurred in the presence of cycloheximide, but was prevented if thioredoxin reductase was inhibited, suggesting that it was due to endogenous methionine sulphoxide reductase activity. These results show that cell-permeable chloramines, either directly or when formed in medium, could regulate NF-kappaB activation via reversible IkappaBalpha oxidation.
authors.affiliation: Free Radical Research Group, Department of Pathology, Christchurch School of Medicine and Health Sciences, P.O. Box 4345, Christchurch, New Zealand. robyn.midwinter@chmeds.ac.nz
authors.forename: Robyn G
authors.initials: RG
authors.lastname: Midwinter
authors.forename: Fook-Choe
authors.initials: FC
authors.lastname: Cheah
authors.forename: Jackob
authors.initials: J
authors.lastname: Moskovitz
authors.forename: Margret C
authors.initials: MC
authors.lastname: Vissers
authors.forename: Christine C
authors.initials: CC
authors.lastname: Winterbourn
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000975:Antioxidants / Q000494:pharmacology; D002463:Cell Membrane Permeability; D002700:Chloramines / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism / Q000494:pharmacology; D005998:Glycine / Q000031:analogs & derivatives* / Q000494:pharmacology; D006801:Humans; D006997:Hypochlorous Acid / Q000494:pharmacology; D020672:I-kappa B Proteins / Q000737:chemistry / Q000187:drug effects*; D019169:Jurkat Cells; D008715:Methionine / Q000378:metabolism*; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism; D009504:Neutrophils / Q000378:metabolism; D016877:Oxidants / Q000494:pharmacology*; D010084:Oxidation-Reduction; D013654:Taurine / Q000031:analogs & derivatives / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: D000975:Antioxidants; D002700:Chloramines; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D009363:Neoplasm Proteins; D016877:Oxidants; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha; D013654:Taurine; C012826:glycine chloramine; C043410:N-chlorotaurine; D006997:Hypochlorous Acid; D008715:Methionine; D005978:Glutathione; D005998:Glycine