Abstract
Cytomegaloviruses all encode the viral inhibitor of caspase-8-induced apoptosis (vICA). After binding to this initiator caspase, vICA blocks caspase-8 proteolytic activity and ability to activate caspase-3 and/or caspase-7. In this manner, vICA has long been known to prevent apoptosis triggered via tumor necrosis factor (TNF) family death receptor-dependent extrinsic signaling. Here, we employ fully wild-type murine cytomegalovirus (MCMV) and vICA-deficient MCMV (∆M36) to investigate the contribution of TNF signaling to apoptosis during infection of different cell types. ∆M36 shows the expected ability to kill mouse splenic hematopoietic cells, bone marrow-derived macrophages (BMDM), and dendritic cells (BMDC). Antibody blockade or genetic elimination of TNF protects myeloid cells from death, and caspase-8 activation accompanies cell death. Interferons, necroptosis, and pyroptotic gasdermin D (GSDMD) do not contribute to myeloid cell death. Human and murine fibroblasts or murine endothelial cells (SVEC4-10) normally insensitive to TNF become sensitized to ∆M36-induced apoptosis when treated with TNF or TNF-containing BMDM-conditioned medium. We demonstrate that myeloid cells are the natural source of TNF that triggers apoptosis in either myeloid (autocrine) or non-myeloid cells (paracrine) during ∆M36 infection of mice. Caspase-8 suppression by vICA emerges as key to subverting innate immune elimination of a wide variety of infected cell types.
Highlights
Programmed cell death pathways provide well-orchestrated defense mechanisms leading to clearance of unnecessary, injured, or infected cells during development, disease, and pathogen invasion.Apoptosis is an evolutionarily conserved programmed cell death pathway, either mediated by external factors or by intracellular stress induced by DNA damage, hypoxia, organelle dysfunction, infection, and metabolic inhibition [1]
These observations suggest that RIPK1, CASP8, and cFLIPL all contribute to the cell autonomous signaling network associated with pathogen sensing such that infection induces their expression, thereby priming cells to die unless suppressed by viral inhibitor of caspase-8-induced apoptosis (vICA)
Ripk3−/− murine embryonic fibroblasts (MEF) succumbed but Casp8−/− Ripk3−/− cells completely resisted this apoptosis, establishing the requirement for CASP8 during ∆M36-induced death in permissive non-myeloid cell types. ∆M36-induced death in human fibroblasts indicated that the murine cytomegalovirus (MCMV) M36-encoded vICA acts in a species-independent fashion, similar to what we found in studies of M45mutRHIM-induced necroptosis [8]
Summary
Programmed cell death pathways provide well-orchestrated defense mechanisms leading to clearance of unnecessary, injured, or infected cells during development, disease, and pathogen invasion. Consistent with the expected mechanism of action [23,28], bone marrow-derived macrophages (BMDMs) from mice with deficiency in CASP8 and necroptotic kinase RIPK3 are not susceptible to murine vICA-deficient virus (∆M36)-induced apoptosis [33]. When infected, these mice exhibit normalized levels of dissemination that have been attributed to the inability of infected macrophages to support extrinsic apoptosis. When expressed independently of virus infection, vICA binds to the pro-domain of CASP8 [23] and prevents TNF-, Fas-, or RIPK3 inhibitor-dependent apoptosis in fibroblasts [30,44]. In all CMV-infected cells, TNF signaling may eliminate infected cells unless CASP8 proteolytic activity is suppressed by vICA
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