Disruption of NF-κB signaling and chemokine gene activation by retroviral mediated expression of IKKγ/NEMO mutants

Abstract

Phosphorylation of IκBs - the cytoplasmic inhibitors of the NF-KB transcription factors - is the key event which triggers activation of the NF-KB cascade. Signal-mediated phosphorylation of IκBα is mediated by a multiprotein complex, the IKB kinase (IKK) complex, which is composed of at least three identified subunits. Two of these polypeptides, IKKα and IKKβ, also known as IKK1 and IKK2, are the Catalytic subunits of the kinase complex and phosphorylate IκBα and IκBβ. The third component, NEMO/IKKγ, does not exhibit kinase activity, but rather constitutes a regulatory subunit. In the present study, C-terminal truncated forms of IKK-γ - ΔC-IKKγ 306 and ΔC-IKKγ 261 - were stably expressed in the myeloid cell line U937 by retroviral-mediated gene transfer. Overexpression of ΔC-IKKγ resulted in a reduction in IKK kinase activity in vitro, a subsequent decrease in NF-KB DNA binding activity, and inhibition of chemokine gene induction in response to TNFα stimulation or paramyxovirus infection. This study demonstrates the efficacy of ΔC-IKKγ as a repressor of IKK signaling and NF-KB activation and suggests a potential gene therapy approach to limit chronic inflammation due to chemokine hyperactivation. © 2001 Academic Press.