Gene expression profiling in conjunction with physiological rescues of IKKα-null cells with wild type or mutant IKKα reveals distinct classes of IKKα/NF-κB-dependent genes

Cellular responses to stress-like stimuli require the I kappa B kinase (IKK) signalsome (IKK alpha, IKK beta, and NEMO/IKK gamma) to activate NF-kappa B-dependent genes. IKK beta and NEMO/IKK gamma are required to release NF-kappa B p65/p50 heterodimers from I kappa B alpha, resulting in their nuclear migration and sequence-specific DNA binding; but IKK alpha was found to be dispensable for this initial phase of canonical NF-kappa B activation. Nevertheless, IKK alpha(-/-) mouse embryonic fibroblasts (MEFs) fail to express NF-kappa B targets in response to proinflammatory stimuli, uncovering a nuclear role for IKK alpha in NF-kappa B activation. However, it remains unknown whether the global defect in NF-kappa B-dependent gene expression of IKK alpha(-/-) cells is caused by the absence of IKK alpha kinase activity. We show by gene expression profiling that rescue of near physiological levels of wild type IKK alpha in IKK alpha(-/-) MEFs globally restores expression of their canonical NF-kappa B target genes. To prove that the kinase activity of IKK alpha was required on a genomic scale, the same physiological rescue was performed with a kinase-dead, ATP binding domain IKK alpha mutant (IKK alpha(K44M)). Remarkably, the IKK alpha(K44M) protein rescued similar to 28% of these genes, albeit in a largely stimulus-independent manner with the notable exception of several genes that also acquired tumor necrosis factor-alpha responsiveness. Thus the IKK alpha-containing signalsome unexpectedly functions in the presence and absence of extracellular signals in both kinase-dependent and -independent modes to differentially modulate the expression of five distinct classes of IKK alpha/NF-kappa B-dependent genes.