Inhibition of IκB-α Phosphorylation and Degradation and Subsequent NF-κB Activation by Glutathione Peroxidase Overexpression

• Published in: The Journal of cell biology Vol. 133; no. 5; pp. 1083 - 1093
• Main Authors: Kretz-Remy, Carole, Mehlen, Patrick, Mirault, Marc-Edouard, Arrigo, André-Patrick
• Format: Journal Article
• Language: English
• Published: United States Rockefeller University Press 01.06.1996; The Rockefeller University Press
• Subjects: Cell Line; Cell lines; Cell Nucleus - metabolism; Cells; DNA; DNA - metabolism; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - metabolism; Gene Expression; Genes, Reporter; Glutathione Peroxidase - genetics; Glutathione Peroxidase - metabolism; Humans; Hydrogen; Hydrogen Peroxide - pharmacology; I-kappa B Proteins; NF-kappa B - metabolism; NF-KappaB Inhibitor alpha; Oxidative Stress; Peroxides; Phosphorylation; Protein isoforms; Reactive oxygen species; Reactive Oxygen Species - metabolism; T lymphocytes; Transcriptional Activation; Transfection; Tumor Necrosis Factor-alpha - pharmacology
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.133.5.1083

We report here that both κB-dependent transactivation of a reporter gene and NF-κB activation in response to tumor necrosis factor (TNFα) or H2O2treatments are deficient in human T47D cell transfectants that overexpress seleno-glutathione peroxidase (GSHPx). These cells feature low reactive oxygen species (ROS) levels and decreased intracellular ROS burst in response to TNFα treatment. Decreased ROS levels and NF-κB activation were likely to result from GSHPx increment since these phenomena were no longer observed when GSHPx activity was reduced by selenium depletion. The cellular contents of the two NF-κB subunits (p65 and p50) and of the inhibitory subunit IκB-α were unaffected by GSHPx overexpression, suggesting that increased GSHPx activity interfered with the activation, but not the synthesis or stability, of NF-κB. Nuclear translocation of NF-κB as well as IκB-α degradation were inhibited in GSHPx-overexpressing cells exposed to oxidative stress. Moreover, in control T47D cells exposed to TNFα, a time correlation was observed between elevated ROS levels and IκB-α degradation. We also show that, in growing T47D cells, GSHPx overexpression altered the isoform composition of IκB-α, leading to the accumulation of the more basic isoform of this protein. GSHPx overexpression also abolished the TNFα-mediated transient accumulation of the acidic and highly phosphorylated IκB-α isoform. These results suggest that intracellular ROS are key elements that regulate the phosphorylation of IκB-α, a phenomenon that precedes and controls the degradation of this protein, and then NF-κB activation.