I kappa B kinase, a molecular target for inhibition by 4-hydroxy-2-nonenal

In unstimulated cells, transcription factor NF-kappaB is retained in the cy toplasm by interaction with the inhibitory protein, I kappaB alpha. Appropr iate cellular stimuli inactivate I kappaB alpha by phosphorylation, ubiquin ation, and proteolytic degradation, which allows NF-kappaB to translocate t o the nucleus and modulate gene expression. 4-Hydroxy-2-nonenal (HNE), a ma jor lipid peroxidation product, inhibits activation of NF-kappaB in the hum an colorectal carcinoma cell line (RKO) and human lung carcinoma cell line (H1299), Pretreatment of cells with HNE dose-dependently suppresses tetrade canoylphorbol acetate (TPA)/ionomycin (IM)-induced NF-kappaB DNA binding ac tivity and transactivation of luciferase-based reporter constructs. HNE pre treatment has no affect on TPA/IM-induced AP-1 DNA binding activity. HNE in hibits TPA/ IM-induced degradation of I kappaB alpha in both H1299 and Jurk at 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, Covale nt adducts of HNE to IKK are detected on Western blots using antibodies aga inst IKK or HNE-protein conjugates, Addition of dithiothreitol prevents HNE modification of IKK. Thus, HNE is an endogenous inhibitor of NF-kappaB act ivation that acts by preventing IKK activation and subsequent I kappaB alph a degradation.