The transcription factor NF-kappaB is critical for setting the cellular sensitivities to apoptotic stimuli, including DNA damaging anticancer agents. Central to NF-kappaB signaling pathways is NEMO/IKKgamma, the regulatory subunit of the cytoplasmic IkappaB kinase (IKK) complex. While NF-kappaB activation by genotoxic stress provides an attractive paradigm for nuclear-to-cytoplasmic signaling pathways, the mechanism by which nuclear DNA damage modulates NEMO to activate cytoplasmic IKK remains unknown. Here, we show that genotoxic stress causes nuclear localization of IKK-unbound NEMO via site-specific SUMO-1 attachment. Surprisingly, this sumoylation step is ATM-independent, but nuclear localization allows subsequent ATM-dependent ubiquitylation of NEMO to ultimately activate IKK in the cytoplasm. Thus, genotoxic stress induces two independent signaling pathways, SUMO-1 modification and ATM activation, which work in concert to sequentially cause nuclear targeting and ubiquitylation of free NEMO to permit the NF-kappaB survival pathway. These SUMO and ubiquitin modification pathways may serve as anticancer drug targets.