Regulation of the NF-κB activation pathway by isolated domains of FIP3/IKKγ, a component of the IκB-α kinase complex

Abstract

FIP3, isolated as a type 2 adenovirus E3-14.7-kDa interacting protein, is an essential component of the multimeric IκB-α kinase (IKK) complex and has been shown to interact with various components (Fas receptor-interacting protein, NF-κB-inducing kinase, IKKβ) of the NF-κB activation pathway. FIP3 has also been shown to repress basal and tumor necrosis factor (TNF) α- induced NF-κB activity as well as to induce cell death when overexpressed. The adenovirus E3-14.7-kDa protein (E3-14.7K) is an inhibitor of TNFα- induced cell death. In the current study, we generated deletion mutants to map the domains of FIP3, which are responsible for its various functions. The NF-κB inhibitory activity and the E3-14.7K binding domains were mapped at the carboxyl half of the FIP3 protein. We also found that the carboxyl- terminal half of FIP3 blocked TNFα-induced IκB-α phosphorylation and subsequent degradation, which suggests that the stabilization of the cytoplasmic inhibitor of NF-κB underlies the FIP3 inhibition of NF-κB activity. The amino-terminal 119 amino acids were responsible for the FIP3- IKKβ and FIP3-IKKα interaction, and the middle of the protein (amino acids 201-300) appeared to be both the FIP3 self-association domain as well as the FIP3-Fas receptor-interacting protein interaction domain. Thus, FIP3 might serve as a scaffold protein to organize the various components of the IκB-α kinase complex. Whereas the full-length protein is required for efficient cell death, the amino-terminal 200 amino acids are sufficient to cause rounding and detachment of the cells from the monolayer.