Ionizing radiation-induced NF-κB activation requires PARP-1 function to confer radio-resistance

Recent reports implicate poly(ADP-ribose) polymerase-1 (PARP-1) in the activation of NF-κB. We investigated the role of PARP-1 in the NF-κB signalling cascade induced by ionizing radiation (IR). AG14361, a potent PARP-1 inhibitor, was used in two breast cancer cell lines expressing different levels of constitutively activated NF-κB, as well as mouse embryonic fibroblasts (MEFs) proficient or deficient for PARP-1 or NF-κB p65. In the breast cancer cell lines, AG14361 had no effect on IR-induced degradation of IκBα or nuclear translocation of p50 or p65. However, AG14361 inhibited IR-induced NF-κB dependent transcription of a luciferase reporter gene. Similarly, in PARP-1(-/-) MEFs, IR-induced nuclear translocation of p50 and p65 was normal, but κB binding and transcriptional activation did not occur. AG14361 sensitized both breast cancer cell lines to IR-induced cell killing, inhibited IR-induced XIAP expression and increased caspase-3 activity. However, AG14361 failed to increase IR-induced caspase activity when p65 was knocked down by siRNA. Consistent with this, AG14361 sensitized p65(+/+) but not p65(-/-) MEFs to IR. We conclude that PARP-1 activity is essential in the upstream regulation of IR-induced NF-κB activation. These data indicate that potentiation of IR-induced cytotoxicity by AG14361 is mediated soley by inhibition of NF-κB activation.