Building Signaling Complexes with Ubiquitin

Abstract

Signal transduction often involves the regulated interaction of proteins through specific recognition domains. Kanamaya et al. report that the adaptor proteins transforming growth factor-ß-activated kinase (TAK)-binding proteins, TAB2 and TAB3, recognize polyubiquitin chains linked through lysine 63 of ubiquitin (K63-polyUb). In contrast to polyubiquitin chains linked through lysine 48, which target proteins for proteasomal degradation, conjugation of proteins with K63-polyUb appears to contribute to regulation of signaling pathways. Recognition of K63-polyUb by TAB proteins required a C-terminal conserved zinc finger (ZnF) domain. TAB2 and TAB3 are involved in NF-κB (nuclear factor κB) and JNK (c-Jun N-terminal kinase) activation by interleukin-1 ß (IL-1ß) and tumor necrosis factor-α (TNF-α). In a reporter-gene assay to measure NF-κB activation, mutations in the ZnF domain blocked TAB2- or TAB3-mediated stimulation of NF-κB. In vitro kinase assays demonstrated that the ZnF domain is required for stimulation of IKK (IκB kinase) and TAK1, two kinases involved in NF-κB activation. The ZnF domain could be replaced with other ubiquitin-binding motifs to sustain activation of the reporter gene or stimulation of the kinases in the in vitro assay, suggesting that ubiquitin binding was crucial. In human embryonic kidney 293 (HEK293) cells, overexpressed TAB2 bound polyubiquitinated TRAF6 (tumor necrosis factor receptor-associated factor 6), a ubiquitin ligase that can couple K63-polyUb to target proteins and that participates in activation of NF-κB by IL-1ß. TRAF2 is another ubiquitin ligase that participates in regulation of NF-κB by TNF-α however, TRAF2 is not autoubiquitinated. In HeLa cells exposed to TNF-α, immunoprecipitation experiments suggested that TRAF2 ubiquitinated RIP (receptor-interacting protein), which then recruited TAB2 or TAB3. The authors suggest that K63-polyUb on TRAF6 (for IL-1ß signaling) or on RIP (for TNF-α signaling) serves to recruit TAB2 and TAB3 to signaling complexes, allowing TAK1 and IKK activation to stimulate NF-κB. A. Kanayama, R. B. Seth, L. Sun, C. K. Ea, M. Hong, A. Shaito, Y. H. Chiu, L. Deng, Z. J. Chen, TAB2 and TAB3 activate the NF-κB pathway through binding to polyubiquitin chains. Mol. Cell 15 , 535-548 (2004). [Online Journal]