Tumor necrosis factor (TNF) signaling, but not TWEAK (TNF-like weak inducer of apoptosis)-triggered cIAP1 (cellular inhibitor of apoptosis protein 1) degradation, requires cIAP1 RING dimerization and E2 binding.
Cellular inhibitor of apoptosis (cIAP) proteins, cIAP1 and cIAP2, are important regulators of tumor necrosis factor (TNF) superfamily (SF) signaling and are amplified in a number of tumor types. They are targeted by IAP antagonist compounds that are undergoing clinical trials. IAP antagonist compounds trigger cIAP autoubiquitylation and degradation. The TNFSF member TWEAK induces lysosomal degradation of TRAF2 and cIAPs, leading to elevated NIK levels and activation of non-canonical NF-kappaB. To investigate the role of the ubiquitin ligase RING domain of cIAP1 in these pathways, we used cIAP-deleted cells reconstituted with cIAP1 point mutants designed to interfere with the ability of the RING to dimerize or to interact with E2 enzymes. We show that RING dimerization and E2 binding are required for IAP antagonists to induce cIAP1 degradation and protect cells from TNF-induced cell death. The RING functions of cIAP1 are required for full TNF-induced activation of NF-kappaB, however, delayed activation of NF-kappaB still occurs in cIAP1 and -2 double knock-out cells. The RING functions of cIAP1 are also required to prevent constitutive activation of non-canonical NF-kappaB by targeting NIK for proteasomal degradation. However, in cIAP double knock-out cells TWEAK was still able to increase NIK levels demonstrating that NIK can be regulated by cIAP-independent pathways. Finally we show that, unlike IAP antagonists, TWEAK was able to induce degradation of cIAP1 RING mutants. These results emphasize the critical importance of the RING of cIAP1 in many signaling scenarios, but also demonstrate that in some pathways RING functions are not required.
Feltham
Rebecca
R
Department of Biochemistry, La Trobe University, Victoria 3086, Australia.
Moulin
Maryline
M
Vince
James E
JE
Mace
Peter D
PD
Wong
Wendy Wei-Lynn
WW
Anderton
Holly
H
Day
Catherine L
CL
Vaux
David L
DL
Silke
John
J
D000818:Animals; D017209:Apoptosis; D000074049:Cytokine TWEAK; D019281:Dimerization; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000378:metabolism*; D051379:Mice; D008958:Models, Molecular; D008968:Molecular Conformation; D016328:NF-kappa B / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding; D025941:Protein Interaction Mapping; D015398:Signal Transduction; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D048069:Tumor Necrosis Factors / Q000378:metabolism*
D000074049:Cytokine TWEAK; D051034:Inhibitor of Apoptosis Proteins; D016328:NF-kappa B; C491645:Tnfsf12 protein, mouse; D014409:Tumor Necrosis Factor-alpha; D048069:Tumor Necrosis Factors