Abstract
Macrophages express TNFR1 as well as TNFR2 and are also major producers of tumor necrosis factor (TNF), especially upon contact with pathogen-associated molecular patterns. Consequently, TNF not only acts as a macrophage-derived effector molecule but also regulates the activity and viability of macrophages. Here, we investigated the individual contribution of TNFR1 and TNFR2 to TNF-induced cell death in macrophages. Exclusive stimulation of TNFR1 showed no cytotoxic effect whereas selective stimulation of TNFR2 displayed mild cytotoxicity. Intriguingly, the latter was strongly enhanced by the caspase inhibitor zVAD-fmk. The strong cytotoxic activity of TNFR2 in the presence of zVAD-fmk was reversed by necrostatin-1, indicating necroptotic cell death. TNFR1- and TNF-deficient macrophages turned out to be resistant against TNFR2-induced cell death. In addition, the cIAP-depleting SMAC mimetic BV6 also enforced TNF/TNFR1-mediated necroptotic cell death in the presence of zVAD-fmk. In sum, our data suggest a model in which TNFR2 sensitizes macrophages for endogenous TNF-induced TNFR1-mediated necroptosis by the known ability of TNFR2 to interfere with the survival activity of TRAF2-cIAP1/2 complexes.
Highlights
Tumor necrosis factor (TNF) is a pleiotropic cytokine that occurs as a type II transmembrane protein but can be released from the plasma membrane by proteolytic processing.[1]
Using macrophages genetically deficient for TNFR1, TNFR2 or TNF together with TNFR1and TNFR2-specific TNF variants, we show that TNFR2 activation sensitizes macrophages for TNFR1-mediated necroptosis triggered by autocrine produced TNF and provide evidence that this is related to TNFR2-induced depletion/ degradation of TRAF2-cIAP1/2 complexes
Cells were challenged with the TNFR1- and TNFR2-specific ligands in the presence of the pan-caspase inhibitor zVAD-fmk, which can unleash potential necroptotic activity by blocking caspase-8-mediated cleavage of RIP1.19,20 Irrespective of zVAD-fmk, human TNF-mediated TNFR1 activation had no cytotoxic effects, neither in HoxB8immortalized MPCs, nor in macrophages (Figure 1c)
Summary
Tumor necrosis factor (TNF) is a pleiotropic cytokine that occurs as a type II transmembrane protein but can be released from the plasma membrane by proteolytic processing.[1]. Several mechanisms for the cross-talk between TNFR1 and TNFR2 have been identified.[1] Besides the obvious competition for ligand binding, TNFR1 and TNFR2 can induce, for example, autocrine TNF production in a cell type-specific manner.[1] In context of TNFR1 activation by soluble TNF, subsequent induction of membrane-bound TNF results in costimulation of TNFR2, thereby converting the initially transient activation into sustained autocrine signaling. By depletion and/or degradation of TRAF2, TNFR2 is capable to modulate TNFR1 signaling.[1] TNFR2 but not TNFR1, stimulates the alternative NFκB pathway by triggering proteolytic processing of the inactive p100/RelB dimers into active p52/RelB NFκB complexes.[7] Notably, TNFR2-induced alternative NFκB signaling can be enhanced by TNFR1-mediated induction of p100 and RelB expression via the classical NFκB pathway.[7]. Using macrophages genetically deficient for TNFR1, TNFR2 or TNF together with TNFR1and TNFR2-specific TNF variants, we show that TNFR2 activation sensitizes macrophages for TNFR1-mediated necroptosis triggered by autocrine produced TNF and provide evidence that this is related to TNFR2-induced depletion/ degradation of TRAF2-cIAP1/2 complexes
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