Inducible phosphorylation of NF-κB p65 at serine 468 by T cell costimulation is mediated by IKKε

Here we identify IKK epsilon as a novel NF-kappa B p65 kinase that mediates inducible phosphorylation of Ser(468) and Ser(536) in response to T cell costimulation. In addition, the kinase activity of IKK epsilon contributes to the control of p65 nuclear uptake. Serines 468 and 536 are evolutionarily conserved, and the surrounding amino acids display sequence homology. Down-regulation of IKK epsilon levels by small interfering RNA does not affect inducible phosphorylation of Ser(536) but largely prevents Ser(468) phosphorylation induced by T cell costimulation. Ser(536)-phosphorylated p65 is found predominantly in the cytosol. In contrast, the Ser(468) phosphorylated form of this transcription factor occurs mainly in the nucleus, suggesting a function for transactivation. Reconstitution of p65(-/-) cells with either wild type p65 or point-mutated p65 variants showed that inducible phosphorylation of Ser(468) serves to enhance p65-dependent transactivation. These results also provide a mechanistic link that helps to explain the relevance of IKK epsilon for the expression of a subset of NF-kappa B target genes without affecting cytosolic I kappa B degradation.