Disruption of NF-kappa B signaling and chemokine gene activation by retroviral mediated expression of IKK gamma/NEMO mutants

Phosphorylation Of I kappa Bs-the cytoplasmic inhibitors of the NF-kappaB t ranscription factors-is the key event which triggers activation of the NF-k appaB cascade. Signal-mediated phosphorylation Of I kappaB alpha is mediate d by a multiprotein complex, the I kappaB kinase (IKK) complex, which is co mposed of at least three identified subunits. Two of these polypeptides, IK K alpha and IKK beta, also known as IKK1 and IKK2, are the catalytic subuni ts of the kinase complex and phosphorylate I kappaB alpha and I kappaB beta . The third component, NEMO/IKK gamma, does not exhibit kinase activity, bu t rather constitutes a regulatory subunit, In the present study, C-terminal truncated forms of IKK-gamma-DeltaC-IKK gamma 306 and DeltaC-IKK gamma 261 - were stably expressed in the myeloid calf line U937 by retroviral-mediat ed gene transfer, Overexpression of DeltaC-IKK gamma resulted in a reductio n in IKK kinase activity in vitro, a subsequent decrease in NF-kappaB DNA b inding activity, and inhibition of chemokine gene induction in response to TNF alpha stimulation or paramyxovirus infection. This study demonstrates t he efficacy of DeltaC-IKK gamma as a repressor of IKK signaling and NF-kapp aB activation and suggests a potential gene therapy approach to limit chron ic inflammation due to chemokine hyperactivation. (C) 2001 Academic Press.