Tyrosine Phosphorylation of I B Activates NF B through a Redox-regulated and c-Src-dependent Mechanism Following Hypoxia/Reoxygenation*

NF B is a critical transcription factor involved in modulating cellular responses to environmental injuries. Tyrosine 42 phosphorylation of I B has been shown to mediate NF B activation following hypoxia/ reoxygenation (H/R) or pervanadate treatment. This pathway differs from the canonical proinflammatory pathways, which mediate NF B activation through serine phosphorylation of I B by the IKK complex. In the present study, we investigated the involvement of c-Src in the redox activation of NF B following H/R or pervanadate treatment. Our results demonstrate that pervanadate or H/R treatment leads to tyrosine phosphorylation of I B and NF B transcriptional activation independent of the IKK pathway. In contrast, inhibition of c-Src by pp2 treatment or in c-Src ( / ) knockout cell lines, demonstrated a significant reduction in I B tyrosine phosphorylation and NF B activation following pervanadate or H/R treatment. Overexpression of glutathione peroxidase-1 or catalase, but not Mn-SOD or Cu,Zn-SOD, significantly reduced both NF B activation and tyrosine phosphorylation of I B . In vitro kinase assays further demonstrated that immunoprecipitated c-Src has the capacity to directly phosphorylate GSTI B and that this I B kinase activity is significantly reduced by Gpx-1 overexpression. These results suggest that c-Src-dependent tyrosine phosphorylation of I B and subsequent activation of NF B is controlled by intracellular H2O2 and defines an important redox-regulated pathway for NF B activation following H/R injury that is independent of the IKK complex.