The Zinc Finger Protein A20 Inhibits TNF-induced NF- k B–dependent Gene Expression by Interfering with an RIP- or TRAF2-mediated Transactivation Signal and Directly Binds to a Novel NF- k B–inhibiting Protein ABIN

The zinc finger protein A20 is a tumor necrosis factor (TNF)– and interleukin 1 (IL-1)-inducible protein that negatively regulates nuclear factor-kappa B (NFk B)–dependent gene expression. However, the molecular mechanism by which A20 exerts this effect is still unclear. We show that A20 does not inhibit TNFinduced nuclear translocation and DNA binding of NFk B, although it completely prevents the TNFinduced activation of an NFk B–dependent reporter gene, as well as TNF-induced IL-6 and granulocyte macrophage–colony stimulating factor gene expression. Moreover, NFk B activation induced by overexpression of the TNF receptor–associated proteins TNF receptor–associated death domain protein (TRADD), receptor interacting protein (RIP), and TNF receptor–associated factor 2 (TRAF2) was also inhibited by expression of A20, whereas NFk B activation induced by overexpression of NFk B–inducing kinase (NIK) or the human T cell leukemia virus type 1 (HTLV-1) Tax was unaffected. These results demonstrate that A20 inhibits NFk B–dependent gene expression by interfering with a novel TNF-induced and RIPor TRAF2-mediated pathway that is different from the NIK–I k B kinase pathway and that is specifically involved in the transactivation of NFk B. Via yeast two-hybrid screening, we found that A20 binds to a novel protein, ABIN, which mimics the NFk B inhibiting effects of A20 upon overexpression, suggesting that the effect of A20 is mediated by its interaction with this NFk B inhibiting pro-