IKKγ/NEMO facilitates the recruitment of the IκB Proteins into the IκB kinase complex

Abstract IKKγ/NEMO is an essential regulatory component of the IκB kinase complex that is required for NF-κB activation in response to various stimuli including tumor necrosis factor-α and interleukin-1β. To investigate the mechanism by which IKKγ/NEMO regulates the IKK complex, we examined the ability of IKKγ/NEMO to recruit the IκB proteins into this complex. IKKγ/NEMO binding to wild-type, but not to a kinase-deficient IKKβ protein, facilitated the association of IκBα and IκBβ with the high molecular weight IKK complex. Following tumor necrosis factor-α treatment of HeLa cells, the majority of the phosphorylated form of endogenous IκBα was associated with the high molecular weight IKK complex in HeLa cells and parental mouse embryo fibroblasts but not in IKKγ/NEMO-deficient cells. Finally, we demonstrate that IKKγ/NEMO facilitates the association of the IκB proteins and IKKβ and leads to increases in IKKβ kinase activity. These results suggest that an important function of IKKγ/NEMO is to facilitate the association of both IKKβ and IκB in the high molecular weight IKK complex to increase IκB phosphorylation.