GENOTOXIC STRESS Sumoylating NEMO

Huang et al. investigated activation of the transcription factor nuclear factor-B (NF-B) and implicated two distinct signaling pathways in the response to genotoxic stress. NF-B, which mediates cellular responses to stress, is sequestered in the cytoplasm through interactions with its inhibitors, IBα and IBβ. Stimulation by proinflammatory cytokines, or other extracellular mediators of stress responses, leads to activation of IB kinase (IKK) [composed of two catalytic subunits and the NF-B essential modulator (NEMO) regulatory subunit]. IKK activation leads to the proteasomal degradation of IB and the release of NF-B, which translocates to the nucleus and activates target genes. NF-B activation following damage to DNA also depends on modulation of NEMO to activate IKK; however, the mechanisms underlying the nuclear-to-cytoplasmic signaling pathway remain poorly understood. Huang et al. used Western analysis in combination with immunoprecipitation to show that, in mouse 1.3E2 pre-B cells transfected with NEMO, DNA-damaging chemotherapeutic agents induced first NEMO sumoylation and then NEMO ubiquitination. Sumoylation, which was required for DNA damage-dependent activation of IKK and NF-B, led to nuclear accumulation of NEMO that was not associated with IKK. Small interfering RNA knockdown of the ataxia telangiectasia mutant (ATM) kinase, a nuclear kinase that has been implicated in DNA-damage pathways, indicated that ATM was required for NEMO ubiquitination but not sumoylation. Experiments in which NF-B-IBα and NF-B-IBβ were respectively localized to the nucleus and cytoplasm by inhibiting nuclear export indicated that NEMO activation of IKK and NF-B following DNA damage occurred in the cytoplasm. Thus, NF-B activation in response to genotoxic agents involves ATM-independent NEMO sumoylation, which leads to NEMO's accumulation in the nucleus, followed by ATM-dependent ubiquitination, and finally by IKK activation in the cytoplasm.

T. T. Huang, S. M. Wuerberger-Davis, Z.-H. Wu, S. Miyamoto, Sequential modification of NEMO/IKK by SUMO-1 and ubiquitin mediates NF-B activation by genotoxic stress. Cell 115, 565-576 (2003). [Online Journal]