Regulation of IκB kinase (IKK) complex by IKKγ-dependent phosphorylation of the T-loop and C terminus of IKKβ

The mechanistic relationship of phosphorylation of the C terminus of IKK beta with phosphorylation of its T-loop kinase domain within the IKK complex remained unclear. We investigated the regulatory role of the serine cluster residing immediately adjacent to the HLH domain and of the serines in the NEMO/IKK gamma-binding domain (NBD/gamma BD) in the C-terminal portion of IKK beta in MEFs deficient in IKK beta and IKK beta and in yeast reconstitution system. We show that phosphorylation events at the C terminus of IKK beta can be divided into autophosphorylation of the serine cluster adjacent to the HLH domain and phosphorylation of the NBD/gamma BD. Autophosphorylation of the serine cluster occurs immediately after IKK activation and requires IKK gamma. In MEFs, this autophosphorylation does not have the down-regulatory function on the IKK complex that was previously described ( 1). On the other hand, phosphorylation of the NBD/gamma BD regulates IKK gamma-dependent phosphorylation of the T-loop activation domain in IKK beta and, hence, IKK complex activation. Our study suggests that, within the IKK complex, modulation of the NBD/gamma BD by IKK gamma is upstream to the T-loop phosphorylation.