Multiple Autonomous Cell Death Suppression Strategies Ensure Cytomegalovirus Fitness

Abstract

Programmed cell death pathways eliminate infected cells and regulate infection-associated inflammation during pathogen invasion. Cytomegaloviruses encode several distinct suppressors that block intrinsic apoptosis, extrinsic apoptosis, and necroptosis, pathways that impact pathogenesis of this ubiquitous herpesvirus. Here, we expanded the understanding of three cell autonomous suppression mechanisms on which murine cytomegalovirus relies: (i) M38.5-encoded viral mitochondrial inhibitor of apoptosis (vMIA), a BAX suppressor that functions in concert with M41.1-encoded viral inhibitor of BAK oligomerization (vIBO), (ii) M36-encoded viral inhibitor of caspase-8 activation (vICA), and (iii) M45-encoded viral inhibitor of RIP/RHIM activation (vIRA). Following infection of bone marrow-derived macrophages, the virus initially deflected receptor-interacting protein kinase (RIPK)3-dependent necroptosis, the most potent of the three cell death pathways. This process remained independent of caspase-8, although suppression of this apoptotic protease enhances necroptosis in most cell types. Second, the virus deflected TNF-mediated extrinsic apoptosis, a pathway dependent on autocrine TNF production by macrophages that proceeds independently of mitochondrial death machinery or RIPK3. Third, cytomegalovirus deflected BCL-2 family protein-dependent mitochondrial cell death through combined TNF-dependent and -independent signaling even in the absence of RIPK1, RIPK3, and caspase-8. Furthermore, each of these cell death pathways dictated a distinct pattern of cytokine and chemokine activation. Therefore, cytomegalovirus employs sequential, non-redundant suppression strategies to specifically modulate the timing and execution of necroptosis, extrinsic apoptosis, and intrinsic apoptosis within infected cells to orchestrate virus control and infection-dependent inflammation. Virus-encoded death suppressors together hold control over an intricate network that upends host defense and supports pathogenesis in the intact mammalian host.

Differential effects of CMV infection on the viability of cardiac cells

“…Our results show that MCMV infection promotes the viability of cardiomyocytes by increasing mitochondrial biogenesis, reducing ROS accumulation and inflammation, and inhibiting necroptotic and apoptotic signaling pathways. MCMV is known to use multiple strategies to evade the host cell’s programmed cell death mechanisms [ 50 ]. In cardiomyocytes, MCMV utilizes these established mechanisms to promote the survival of cardiomyocytes, although other unrecognized mechanisms may play additional roles.…”

“…MCMV mutants M45 mut RHIM, ∆M36, and ∆M38.5/41.1 have been previously described [ 29 , 50 , 57 ]. Wild type (K181-bac) and mutant viral stocks were generated, clarified, concentrated, and tittered by plaque assay as previously described using NIH 3T3 fibroblasts (ATCC CRL-1658) [ 58 ].…”

The role of RHIM in necroptosis

“…Specifically, the Betaherpesvirinae, a family of large DNA viruses, utilize an elaborate system to evade several host immune response mechanisms and persist as latent infection [ 45 ]. Their mouse-specific member murine cytomegalovirus (MCMV), probably the best studied viral RHIM protein, expresses four different proteins to sequentially block caspase-8-mediated extrinsic apoptosis, RIPK3-mediated necroptosis, and BCL2 family-mediated mitochondrial cell death [ 42 ]. By interacting with ZBP1, RIPK1, and RIPK3, MCMV abolishes regulated cell death and TNF-mediated activation of NF-κB and MAPK [ 46 , 47 ].…”

“…Necroptosis plays a prominent role in the clearance of infected cells and appears to have developed as a backup mechanism to execute cell death when apoptosis fails. However, viral pathogens have co-evolved with the host-regulated cell death machinery and developed mechanisms to ensure cell survival during infection [19,[41][42][43]. In particular, the primate genes encoding RIPK3 and MLKL are in an evolutionary arms race with viral-encoded pathogenicity factors containing functional RHIM domains [34,44].…”

TAT–RHIM: a more complex issue than expected

“…RHIM-containing proteins also play a special role in the defence against viral infections. Infection of a cell by various viruses leads to RHIM-mediated cell death, in particular necroptosis [ 5 , 18 , 19 ]. To ensure their own survival or lifelong infection in the host, specific viruses carry out sophisticated molecular strategies to evade these host cell death responses.…”