Transient exposure to hydrogen peroxide inhibits the ubiquitination of phosphorylated IκBα in TNFα-stimulated HEK293 cells.
During ischemia-reperfusion injury, brief pre-exposure to oxidative stress renders organs resistant to subsequent severe damage. NF-κB is a transcription factor that is involved in reperfusion-induced inflammatory and immune responses. The activity of NF-κB has been shown to be modulated by oxidative stress in various cell types through different pathways. We studied the effect of pre-exposure to oxidative stress on subsequent NF-κB activation in TNFα-stimulated HEK293 cells. The cells were transiently exposed to 0.5 mM H(2)O(2) for 20 min, prior to stimulation with TNFα, and the subsequent expression of NF-κB-dependent genes and the levels of NF-κB signaling molecules were measured. Pre-exposure to H(2)O(2) significantly delayed the TNFα-induced expression of an NF-κB reporter gene and inflammatory proteins (intercellular adhesion molecule-1 and IL-1β). The degradation of inhibitor of NF-κB α (IκBα) and the nuclear translocation of NF-κB were also delayed by H(2)O(2) treatment, whereas IκBα phosphorylation and IκB kinase activity were not changed. When we examined the ubiquitin/proteosome pathway in H(2)O(2)-treated cells, we could not detect significant changes in proteosomal peptidase activities, but we were able to detect a delay of IκBα poly-ubiquitination. Our results suggest that transient exposure to oxidative stress temporally inhibits NF-κB-dependent gene expression by suppressing the poly-ubiquitination of phosphorylated IκBα in HEK293 cells.
Department of Biochemistry, College of Medicine, The Catholic University of Korea, Seoul 137-701, Korea.
Yeji
Y
Lee
Jin
J
Choi
Kyung-Ho
KH
Ha
Dae-Myung
DM
Jue
D021581:Active Transport, Cell Nucleus; D002467:Cell Nucleus / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D057809:HEK293 Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D051550:I-kappa B Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D021381:Protein Transport; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D054875:Ubiquitination / Q000187:drug effects*
D014409:Tumor Necrosis Factor-alpha; D006861:Hydrogen Peroxide; D051550:I-kappa B Kinase