Tumor necrosis factor-associated factor 6 (TRAF6) is an essential adaptor protein for IL-1R or TLR-mediated NF-kappa B signaling pathway activation In previous work we have found NUMBL interacts with TAB2 and negatively regulates NF-kappa B signaling pathway. Here, we report that NUMBL directly binds to TRAF6 in vivo and in vitro NUMBL down-regulates TRAF6 protein level and shortens its half-life. Furthermore. knockdown of NUMBL significantly increases endogenous TRAF6 protein level in the cultured cortical neurons In vivo ubiquitination assays indicate that NUMBL promotes the assembly of K48-linked polyubiquitination chains on TRAF6, but has no significant effect oil its K63-linked polyubiquitination Our results collectively reveal that NUMBL Interacts with TRAF6 and promotes the degradation of TRAF6 in vivo. leading to the inhibition of NF-kappa B signaling pathway (C) 2010 Elsevier Inc All rights reserved