IκB kinase, a molecular target for inhibition by 4-hydroxy-2-nonenal

In unstimulated cells, transcription factor NF-kappaB is retained in the cytoplasm by interaction with the inhibitory protein, I kappaB alpha. Appropriate cellular stimuli inactivate I kappaB alpha by phosphorylation, ubiquination, and proteolytic degradation, which allows NF-kappaB to translocate to the nucleus and modulate gene expression. 4-Hydroxy-2-nonenal (HNE), a major lipid peroxidation product, inhibits activation of NF-kappaB in the human colorectal carcinoma cell line (RKO) and human lung carcinoma cell line (H1299), Pretreatment of cells with HNE dose-dependently suppresses tetradecanoylphorbol acetate (TPA)/ionomycin (IM)-induced NF-kappaB DNA binding activity and transactivation of luciferase-based reporter constructs. HNE pretreatment has no affect on TPA/IM-induced AP-1 DNA binding activity. HNE inhibits TPA/ IM-induced degradation of I kappaB alpha in both H1299 and Jurkat T cells. The accumulation of I kappaB alpha parallels the inhibition of its phosphorylation. At doses that inhibit I kappaB alpha degradation, HNE inhibits I kappaB kinase (IKK) activity by direct reaction with IKK, Covalent adducts of HNE to IKK are detected on Western blots using antibodies against IKK or HNE-protein conjugates, Addition of dithiothreitol prevents HNE modification of IKK. Thus, HNE is an endogenous inhibitor of NF-kappaB activation that acts by preventing IKK activation and subsequent I kappaB alpha degradation.