Activation of the transcription factor NF-κB involves multiple forms of ubiquitylation and the degradation of proteins that inhibit its activity in the unstimulated condition. A20 is a protein with deubiquitylating activity, ubiquitin linkage-modifying activity, and the ability to inhibit NF-κB activation. Verhelst et al. and Tokunaga et al. both identified the seventh zinc finger (ZF7) motif as critical for inhibition of NF-κB activation in response to overexpression of components of LUBAC, a complex that contributes to NF-κB activation by adding linear ubiquitin chains to the adaptor proteins NEMO and RIP1. Both groups reported that, in response to tumor necrosis factor–α (TNF-α), A20 coimmunoprecipitated with LUBAC and NEMO, and this interaction was compromised by mutation of ZF7. A20 with mutations in ZF7 was less effective at inhibition, and introduction of ZF7 coupled to green fluorescent protein was sufficient to inhibit NF-κB activation in response to overexpression of LUBAC. In vitro binding studies indicated that ZF7 preferentially bound to linear ubiquitin chains. Tokunaga et al. also crystallized ZF7 with linear diubiquitin or linear tetraubiquitin and identified residues critical for the interaction. Indeed, mutations that are associated with human disease involved residues that participated in the ZF7–linear ubiquitin interaction, and A20 with these mutations was less effective at inhibiting LUBAC-induced NF-κB activity. Thus, the A20 ZF7 motif defines a linear ubiquitin-binding domain (LUBID), and interactions mediated by the LUBID with NEMO appear to serve as a mechanism by which A20 inhibits activation of NF-κB.
K. Verhelst, I. Carpentier, M. Kreike, L. Meloni, L. Verstrepen, T. Kensche, I. Dikic, R. Beyaert, A20 inhibits LUBAC-mediated NF-κB activation by binding linear polyubiquitin chains via its zinc finger 7. EMBO J. 31, 3845–3855 (2012). [PubMed]
F. Tokunaga, H. Nishimasu, R. Ishitani, E. Goto, T. Noguchi, K. Mio, K. Kamei, A. Ma, K. Iwai, O. Nureki, Specific recognition of linear polyubiquitin by A20 zinc finger 7 is involved in NF-κB regulation. EMBO J. 31, 3856–3870 (2012). [PubMed]