TRAF2 is a biologically important necroptosis suppressor

Abstract

Tumor necrosis factor α (TNFα) triggers necroptotic cell death through an intracellular signaling complex containing receptorinteracting protein kinase (RIPK) 1 and RIPK3, called the necrosome. RIPK1 phosphorylates RIPK3, which phosphorylates the pseudokinase mixed lineage kinase-domain-like (MLKL)-driving its oligomerization and membrane-disrupting necroptotic activity. Here, we show that TNF receptor-associated factor 2 (TRAF2)-previously implicated in apoptosis suppression-also inhibits necroptotic signaling by TNFα. TRAF2 disruption in mouse fibroblasts augmented TNFα-driven necrosome formation and RIPK3-MLKL association, promoting necroptosis. TRAF2 constitutively associated with MLKL, whereas TNFα reversed this via cylindromatosis-dependent TRAF2 deubiquitination. Ectopic interaction of TRAF2 and MLKL required the C-terminal portion but not the N-terminal, RING, or CIM region of TRAF2. Induced TRAF2 knockout (KO) in adult mice caused rapid lethality, in conjunction with increased hepatic necrosome assembly. By contrast, TRAF2 KO on a RIPK3 KO background caused delayed mortality, in concert with elevated intestinal caspase-8 protein and activity. Combined injection of TNFR1-Fc, Fas-Fc and DR5-Fc decoys prevented death upon TRAF2 KO. However, Fas-Fc and DR5-Fc were ineffective, whereas TNFR1-Fc and interferon α receptor (IFNAR1)-Fc were partially protective against lethality upon combined TRAF2 and RIPK3 KO. These results identify TRAF2 as an important biological suppressor of necroptosis in vitro and in vivo.

Necroptosis: Mechanisms and Relevance to Disease

“…Accordingly, the RIPK1-deubiquitinating enzyme cylindromatosis (CYLD) is required for optimal necroptotic responses following TNFR1 ligation (78–80), whereas the ubiquitin-modifying TNF alpha induced protein 3 (TNFAIP3; best known as A20) and LUBAC limit TNFR1-driven necroptosis by favoring RIPK1 ubiquitination (81–85). A20 also suppresses necroptosis by deubiquitinating RIPK3 at K5 (86), whereas CYLD mediates necroptotic effects by deubiquitinating TNF receptor associated factor 2 (TRAF2) (87). …”

“…Aside from being involved in the regulation of extrinsic apoptosis, TRAF2 inhibits necroptosis driven by TNFR1 and other death receptors by directly interacting with MLKL (87, 88). Accordingly, the whole-body deletion of Traf2 in adult mice is lethal, owing to an acute hepatotoxic response that can be prevented by deleting Ripk3 (87).…”

“…Accordingly, the whole-body deletion of Traf2 in adult mice is lethal, owing to an acute hepatotoxic response that can be prevented by deleting Ripk3 (87). Mice lacking Traf2 and Ripk3 , however, succumb in a delayed manner upon the hyperactivation of Casp8 in the intestinal epithelium, and this process can be completely blocked with a cocktail of monoclonal antibodies that antagonize TNFR1 and other death receptors (87).…”

“…Accordingly, the whole-body deletion of Traf2 in adult mice is lethal, owing to an acute hepatotoxic response that can be prevented by deleting Ripk3 (87). Mice lacking Traf2 and Ripk3 , however, succumb in a delayed manner upon the hyperactivation of Casp8 in the intestinal epithelium, and this process can be completely blocked with a cocktail of monoclonal antibodies that antagonize TNFR1 and other death receptors (87). Of note, whereas some authors propose that mitogen-activated protein kinase kinase kinase 7 (MAP3K7; best known as TAK1) mediates antinecroptotic effects by preventing RIPK1 signaling (77, 89, 90), others suggest that prolonged TAK1 activation (which is usually a transient consequence of TNFR1 ligation) may result in RIPK3 phosphorylation and the consequent initiation of a feedforward amplification circuitry with potent pronecroptotic activity (91).…”

Brefeldin A-Inhibited Guanine Nucleotide-Exchange Factor 1 (BIG1) Governs the Recruitment of Tumor Necrosis Factor Receptor-Associated Factor 2 (TRAF2) to Tumor Necrosis Factor Receptor 1 (TNFR1) Signaling Complexes

“…Nevertheless, TRAF2 knockout cells exhibit increased sensitivity to TNF-α-induced apoptosis, suggesting that TRAF2 is required for NF-κB-independent mechanisms that protect against TNF-α-induced apoptosis. Similar effects of TRAF2 on cell death have been demonstrated in Fas or tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor signaling [25,26]. Moreover, a recent report has demonstrated that TRAF2 suppresses the induction of apoptosis through the mechanism of K48-linked polyubiquitination of activated caspase-8, which can account for the NF-κB-independent mechanism by which TRAF2 exerts anti-apoptotic activity [27].…”