The conserved redox-sensitive cysteine residue of the DNA-binding region in the c-Rel protein is involved in the regulation of the phosphorylation ofthe protein

The DNA-binding activity of the transcription nuclear factor kappaB (NF-kap paB) is regulated by a redox-control mechanism involving the reduction of a disulphide bond from a specific cysteine residue conserved in all members of the NF-kappaB family. Thioredoxin is involved in this redox control. DNA binding and transactivating capacity of NF-kappaB are up-regulated by indu cible phosphorylation. Here we demonstrate that the conserved redox cystein e in the c-Rel protein is involved in the phosphorylation regulation of the protein. When this cysteine residue is mutated to an aspartic acid residue , the mutant protein loses its capacity to be phosphorylated and its DNA-bi nding activity. In addition, our results suggest that, when the conserved r edox cysteine is chemically modified by N-ethylmaleimide and 2-chloro-1,3-d initrobenzene, the protein c-Rel cannot be phosphorylated. In contrast, the protein in which the cysteine residue was replaced by a serine residue, cr eating a potential phosphorylation site, is highly phosphorylated and binds kappa sequences. The protein could loose the redox regulation of the phosp horylation when the residue replacing the cysteine can be itself phosphoryl ated. We also show that specific inhibitors of thioredoxin reductases impai r the phosphorylation of the c-Rel protein, suggesting that the redox regul ation of the protein controls its phosphorylation.