For antigen and various cytokine receptor-mediated signalling events, ubiquitination is a fundamental regulatory mechanism implicated in the different signalling pathways emanating from these receptors. Since the discovery of the importance of lysine 48-linked ubiquitin chains for NF-?B activation and the finding that the ubiquitin ligases TRAF2 and cIAP1/2 are recruited to the TNF-RSC, it became clear that the activation of TNF-induced signalling pathways depends strongly on the ubiquitin system. TNF plays a critical role in inflammatory processes and is involved in the regulation of immune responses. Depending on the cellular context, TNF initiates a complex cascade of signalling events that can result in induction of proinflammatory cytokines, cell proliferation, differentiation or cell death. Ligand-induced trimerisation of TNF-R1 leads to the formation of a multi-protein complex, the TNFR1 signalling complex (TNF-RSC). To be able to understand the multifaceted regulatory functions of the ubiquitin network and to comprehend the complex interplay of signalling pathways emanating from TNF-R1, the TNF-RSC and its composition need to be understood at the molecular level. Therefore we newly developed a modified tandem affinity purification (moTAP) procedure which allowed us to physically determine the molecular composition of the TNF-RSC in an unbiased manner. Using the moTAP approach, HOIL-1 and HOIP were identified as two novel, functional components of the native TNF-RSC. Together they were shown to form a linear ubiquitin chain assembly complex (LUBAC), catalysing the formation of linear head-to-tail ubiquitin chains. LUBAC mediates ubiquitination of NEMO with linear ubiquitin chains, required for efficient NF-?B activation following TNF stimulation. We show that the stimulation-dependent recruitment of LUBAC to the TNF-RSC is impaired in TRADD-, TRAF2-, and cIAP1/2-, but not in RIP1- and NEMO-deficient mouse embryonic fibroblast (MEF) cell lines. Furthermore, we demonstrate that the E3 ligase activity of cIAPs, but not of TRAF2, is required for HOIL-1 recruitment to the TNF-RSC. This result, together with the ability of HOIL-1 and HOIP to bind polyubiquitin chains of various linkage types, suggests that LUBAC is recruited to the TNF-RSC via cIAP-generated ubiquitin chains. LUBAC enhances NEMO interaction with the TNF-RSC, stabilises this protein complex, and is required for efficient TNF-induced activation of NF-?B and JNK, resulting in apoptosis inhibition. Finally, we show that the activity of LUBAC is required for stabilisation of the TNF-RSC, thereby adding a third form of ubiquitin linkage to the control of TNF signalling. The identification of HOIL-1 and HOIP as functional constituents of the TNF-RSC provides evidence that LUBAC is an important regulator at the apex of TNF-induced signalling cascades and increases the combinatorial complexity of ubiquitin modifications within this receptor complex. © 2011 Springer Science+Business Media, LLC.

Emmerich, C. H., Schmukle, A. C., Haas, T. L., Gerlach, B., Cordier, S. M., Rieser, E., Walczak, H., The linear ubiquitin chain assembly complex (LUBAC) forms part of the TNF-R1 signalling complex and is required for effective TNF-induced gene induction and prevents TNF-induced apoptosis, <<ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY>>, 2011; 691 (691): 115-126. [doi:10.1007/978-1-4419-6612-4_12] [http://hdl.handle.net/10807/114564]

The linear ubiquitin chain assembly complex (LUBAC) forms part of the TNF-R1 signalling complex and is required for effective TNF-induced gene induction and prevents TNF-induced apoptosis

Haas, Tobias L.;
2011

Abstract

For antigen and various cytokine receptor-mediated signalling events, ubiquitination is a fundamental regulatory mechanism implicated in the different signalling pathways emanating from these receptors. Since the discovery of the importance of lysine 48-linked ubiquitin chains for NF-?B activation and the finding that the ubiquitin ligases TRAF2 and cIAP1/2 are recruited to the TNF-RSC, it became clear that the activation of TNF-induced signalling pathways depends strongly on the ubiquitin system. TNF plays a critical role in inflammatory processes and is involved in the regulation of immune responses. Depending on the cellular context, TNF initiates a complex cascade of signalling events that can result in induction of proinflammatory cytokines, cell proliferation, differentiation or cell death. Ligand-induced trimerisation of TNF-R1 leads to the formation of a multi-protein complex, the TNFR1 signalling complex (TNF-RSC). To be able to understand the multifaceted regulatory functions of the ubiquitin network and to comprehend the complex interplay of signalling pathways emanating from TNF-R1, the TNF-RSC and its composition need to be understood at the molecular level. Therefore we newly developed a modified tandem affinity purification (moTAP) procedure which allowed us to physically determine the molecular composition of the TNF-RSC in an unbiased manner. Using the moTAP approach, HOIL-1 and HOIP were identified as two novel, functional components of the native TNF-RSC. Together they were shown to form a linear ubiquitin chain assembly complex (LUBAC), catalysing the formation of linear head-to-tail ubiquitin chains. LUBAC mediates ubiquitination of NEMO with linear ubiquitin chains, required for efficient NF-?B activation following TNF stimulation. We show that the stimulation-dependent recruitment of LUBAC to the TNF-RSC is impaired in TRADD-, TRAF2-, and cIAP1/2-, but not in RIP1- and NEMO-deficient mouse embryonic fibroblast (MEF) cell lines. Furthermore, we demonstrate that the E3 ligase activity of cIAPs, but not of TRAF2, is required for HOIL-1 recruitment to the TNF-RSC. This result, together with the ability of HOIL-1 and HOIP to bind polyubiquitin chains of various linkage types, suggests that LUBAC is recruited to the TNF-RSC via cIAP-generated ubiquitin chains. LUBAC enhances NEMO interaction with the TNF-RSC, stabilises this protein complex, and is required for efficient TNF-induced activation of NF-?B and JNK, resulting in apoptosis inhibition. Finally, we show that the activity of LUBAC is required for stabilisation of the TNF-RSC, thereby adding a third form of ubiquitin linkage to the control of TNF signalling. The identification of HOIL-1 and HOIP as functional constituents of the TNF-RSC provides evidence that LUBAC is an important regulator at the apex of TNF-induced signalling cascades and increases the combinatorial complexity of ubiquitin modifications within this receptor complex. © 2011 Springer Science+Business Media, LLC.
Inglese
Emmerich, C. H., Schmukle, A. C., Haas, T. L., Gerlach, B., Cordier, S. M., Rieser, E., Walczak, H., The linear ubiquitin chain assembly complex (LUBAC) forms part of the TNF-R1 signalling complex and is required for effective TNF-induced gene induction and prevents TNF-induced apoptosis, <<ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY>>, 2011; 691 (691): 115-126. [doi:10.1007/978-1-4419-6612-4_12] [http://hdl.handle.net/10807/114564]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/10807/114564
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