title: c-IAP1 and c-IAP2 are critical mediators of tumor necrosis factor alpha (TNFalpha)-induced NF-kappaB activation.
abstract: The inhibitor of apoptosis (IAP) proteins are a family of anti-apoptotic regulators found in viruses and metazoans. c-IAP1 and c-IAP2 are recruited to tumor necrosis factor receptor 1 (TNFR1)-associated complexes where they can regulate receptor-mediated signaling. Both c-IAP1 and c-IAP2 have been implicated in TNFalpha-stimulated NF-kappaB activation. However, individual c-IAP1 and c-IAP2 gene knock-outs in mice did not reveal changes in TNF signaling pathways, and the phenotype of a combined deficiency of c-IAPs has yet to be reported. Here we investigate the role of c-IAP1 and c-IAP2 in TNFalpha-stimulated activation of NF-kappaB. We demonstrate that TNFalpha-induced NF-kappaB activation is severely diminished in the absence of both c-IAP proteins. In addition, combined absence of c-IAP1 and c-IAP2 rendered cells sensitive to TNFalpha-induced cell death. Using cells with genetic ablation of c-IAP1 or cells where the c-IAP proteins were eliminated using IAP antagonists, we show that TNFalpha-induced RIP1 ubiquitination is abrogated in the absence of c-IAPs. Furthermore, we reconstitute the ubiquitination process with purified components in vitro and demonstrate that c-IAP1, in collaboration with the ubiquitin conjugating enzyme (E2) enzyme UbcH5a, mediates polymerization of Lys-63-linked chains on RIP1. Therefore, c-IAP1 and c-IAP2 are required for TNFalpha-stimulated RIP1 ubiquitination and NF-kappaB activation.
authors.affiliation: Department of Protein Engineering, Genentech, Inc., South San Francisco, California 94080, USA.
authors.forename: Eugene
authors.initials: E
authors.lastname: Varfolomeev
authors.forename: Tatiana
authors.initials: T
authors.lastname: Goncharov
authors.forename: Anna V
authors.initials: AV
authors.lastname: Fedorova
authors.forename: Jasmin N
authors.initials: JN
authors.lastname: Dynek
authors.forename: Kerry
authors.initials: K
authors.lastname: Zobel
authors.forename: Kurt
authors.initials: K
authors.lastname: Deshayes
authors.forename: Wayne J
authors.initials: WJ
authors.lastname: Fairbrother
authors.forename: Domagoj
authors.initials: D
authors.lastname: Vucic
mesh_terms: D000818:Animals; D016923:Cell Death / Q000502:physiology; D045744:Cell Line, Tumor; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D028861:Nuclear Pore Complex Proteins / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination / Q000502:physiology
chemical_list: C119724:AGFG1 protein, human; D020690:GTPase-Activating Proteins; D051034:Inhibitor of Apoptosis Proteins; D016328:NF-kappa B; D028861:Nuclear Pore Complex Proteins; D016601:RNA-Binding Proteins; C490620:Ralbp1 protein, mouse; D047888:Receptors, Tumor Necrosis Factor, Type I; C506236:Tnfrsf1a protein, mouse; D014409:Tumor Necrosis Factor-alpha