SARS-CoV-2 spike-induced syncytia are senescent and contribute to exacerbated heart failure.

SARS-CoV-2 spike protein (SARS-2-S) induced cell-cell fusion in uninfected cells may occur in long COVID-19 syndrome, as circulating SARS-2-S or extracellular vesicles containing SARS-2-S (S-EVs) were found to be prevalent in post-acute sequelae of COVID-19 (PASC) for up to 12 months after diagnosis. Although isolated recombinant SARS-2-S protein has been shown to increase the SASP in senescent ACE2-expressing cells, the direct linkage of SARS-2-S syncytia with senescence in the absence of virus infection and the degree to which SARS-2-S syncytia affect pathology in the setting of cardiac dysfunction are unknown. Here, we found that the senescent outcome of SARS-2-S induced syncytia exacerbated heart failure progression. We first demonstrated that syncytium formation in cells expressing SARS-2-S delivered by DNA plasmid or LNP-mRNA exhibits a senescence-like phenotype. Extracellular vesicles containing SARS-2-S (S-EVs) also confer a potent ability to form senescent syncytia without de novo synthesis of SARS-2-S. However, it is important to note that currently approved COVID-19 mRNA vaccines do not induce syncytium formation or cellular senescence. Mechanistically, SARS-2-S syncytia provoke the formation of functional MAVS aggregates, which regulate the senescence fate of SARS-2-S syncytia by TNFα. We further demonstrate that senescent SARS-2-S syncytia exhibit shrinked morphology, leading to the activation of WNK1 and impaired cardiac metabolism. In pre-existing heart failure mice, the WNK1 inhibitor WNK463, anti-syncytial drug niclosamide, and senolytic dasatinib protect the heart from exacerbated heart failure triggered by SARS-2-S. Our findings thus suggest a potential mechanism for COVID-19-mediated cardiac pathology and recommend the application of WNK1 inhibitor for therapy especially in individuals with post-acute sequelae of COVID-19.

Comprehensive analytics for virus-cell and cell-cell multinucleation system.

Cell-fusion mediated generation of multinucleated syncytia represent critical feature during viral infection and in development. Efficiency of syncytia formation is usually illustrated as fusion efficiency under given condition by quantifying total number of nuclei in syncytia normalized to total number of nuclei (both within syncytia and unfused cell nuclei) in unit field of view. However heterogeneity in multinucleated syncytia sizes poses challenge in quantification of cell-fusion multinucleation under diverse conditions. Taking in-vitro SARS-CoV-2 spike-protein variants mediated virus-cell fusion model and placenta trophoblast syncytialization as cell-cell fusion model; herein we emphasize wide application of simple unbiased detailed measure of virus-cell and cell-cell multinucleation using experiential cumulative distribution function (CDF) and fusion number events (FNE) approaches illustrating comprehensive metrics for syncytia interpretation.

Human cytomegalovirus glycoprotein variants governing viral tropism and syncytium formation in epithelial cells and macrophages.

Human cytomegalovirus (HCMV) displays a broad cell tropism, and the infection of biologically relevant cells such as epithelial, endothelial, and hematopoietic cells supports viral transmission, systemic spread, and pathogenesis in the human host. HCMV strains differ in their ability to infect and replicate in these cell types, but the genetic basis of these differences has remained incompletely understood. In this study, we investigated HCMV strain VR1814, which is highly infectious for epithelial cells and macrophages and induces cell-cell fusion in both cell types. A VR1814-derived bacterial artificial chromosome (BAC) clone, FIX-BAC, was generated many years ago but has fallen out of favor because of its modest infectivity. By sequence comparison and genetic engineering of FIX, we demonstrate that the high infectivity of VR1814 and its ability to induce syncytium formation in epithelial cells and macrophages depends on VR1814-specific variants of the envelope glycoproteins gB, UL128, and UL130. We also show that UL130-neutralizing antibodies inhibit syncytium formation, and a FIX-specific mutation in UL130 is responsible for its low infectivity by reducing the amount of the pentameric glycoprotein complex in viral particles. Moreover, we found that a VR1814-specific mutation in US28 further increases viral infectivity in macrophages, possibly by promoting lytic rather than latent infection of these cells. Our findings show that variants of gB and the pentameric complex are major determinants of infectivity and syncytium formation in epithelial cells and macrophages. Furthermore, the VR1814-adjusted FIX strains can serve as valuable tools to study HCMV infection of myeloid cells.IMPORTANCEHuman cytomegalovirus (HCMV) is a major cause of morbidity and mortality in transplant patients and the leading cause of congenital infections. HCMV infects various cell types, including epithelial cells and macrophages, and some strains induce the fusion of neighboring cells, leading to the formation of large multinucleated cells called syncytia. This process may limit the exposure of the virus to host immune factors and affect pathogenicity. However, the reason why some HCMV strains exhibit a broader cell tropism and why some induce cell fusion more than others is not well understood. We compared two closely related HCMV strains and provided evidence that small differences in viral envelope glycoproteins can massively increase or decrease the virus infectivity and its ability to induce syncytium formation. The results of the study suggest that natural strain variations may influence HCMV infection and pathogenesis in humans.

High fusion and cytopathy of SARS-CoV-2 variant B.1.640.1.

SARS-CoV-2 variants with undetermined properties have emerged intermittently throughout the COVID-19 pandemic. Some variants possess unique phenotypes and mutations which allow further characterization of viral evolution and Spike functions. Around 1,100 cases of the B.1.640.1 variant were reported in Africa and Europe between 2021 and 2022, before the expansion of Omicron. Here, we analyzed the biological properties of a B.1.640.1 isolate and its Spike. Compared to the ancestral Spike, B.1.640.1 carried 14 amino acid substitutions and deletions. B.1.640.1 escaped binding by some anti-N-terminal domain and anti-receptor-binding domain monoclonal antibodies, and neutralization by sera from convalescent and vaccinated individuals. In cell lines, infection generated large syncytia and a high cytopathic effect. In primary airway cells, B.1.640.1 replicated less than Omicron BA.1 and triggered more syncytia and cell death than other variants. The B.1.640.1 Spike was highly fusogenic when expressed alone. This was mediated by two poorly characterized and infrequent mutations located in the Spike S2 domain, T859N and D936H. Altogether, our results highlight the cytopathy of a hyper-fusogenic SARS-CoV-2 variant, supplanted upon the emergence of Omicron BA.1. (This study has been registered at ClinicalTrials.gov under registration no. NCT04750720.)IMPORTANCEOur results highlight the plasticity of SARS-CoV-2 Spike to generate highly fusogenic and cytopathic strains with the causative mutations being uncharacterized in previous variants. We describe mechanisms regulating the formation of syncytia and the subsequent consequences in a primary culture model, which are poorly understood.

PIK-24 Inhibits RSV-Induced Syncytium Formation via Direct Interaction with the p85α Subunit of PI3K.

Phosphoinositide-3 kinase (PI3K) signaling regulates many cellular processes, including cell survival, differentiation, proliferation, cytoskeleton reorganization, and apoptosis. The actin cytoskeleton regulated by PI3K signaling plays an important role in plasma membrane rearrangement. Currently, it is known that respiratory syncytial virus (RSV) infection requires PI3K signaling. However, the regulatory pattern or corresponding molecular mechanism of PI3K signaling on cell-to-cell fusion during syncytium formation remains unclear. This study synthesized a novel PI3K inhibitor PIK-24 designed with PI3K as a target and used it as a molecular probe to investigate the involvement of PI3K signaling in syncytium formation during RSV infection. The results of the antiviral mechanism revealed that syncytium formation required PI3K signaling to activate RHO family GTPases Cdc42, to upregulate the inactive form of cofilin, and to increase the amount of F-actin in cells, thereby causing actin cytoskeleton reorganization and membrane fusion between adjacent cells. PIK-24 treatment significantly abolished the generation of these events by blocking the activation of PI3K signaling. Moreover, PIK-24 had an obvious binding activity with the p85α regulatory subunit of PI3K. The anti-RSV effect similar to PIK-24 was obtained after knockdown of p85α in vitro or knockout of p85α in vivo, suggesting that PIK-24 inhibited RSV infection by targeting PI3K p85α. Most importantly, PIK-24 exerted a potent anti-RSV activity, and its antiviral effect was stronger than that of the classic PI3K inhibitor LY294002, PI-103, and broad-spectrum antiviral drug ribavirin. Thus, PIK-24 has the potential to be developed into a novel anti-RSV agent targeting cellular PI3K signaling. IMPORTANCE PI3K protein has many functions and regulates various cellular processes. As an important regulatory subunit of PI3K, p85α can regulate the activity of PI3K signaling. Therefore, it serves as the key target for virus infection. Indeed, p85α-regulated PI3K signaling facilitates various intracellular plasma membrane rearrangement events by modulating the actin cytoskeleton, which may be critical for RSV-induced syncytium formation. In this study, we show that a novel PI3K inhibitor inhibits RSV-induced PI3K signaling activation and actin cytoskeleton reorganization by targeting the p85α protein, thereby inhibiting syncytium formation and exerting a potent antiviral effect. Respiratory syncytial virus (RSV) is one of the most common respiratory pathogens, causing enormous morbidity, mortality, and economic burden. Currently, no effective antiviral drugs or vaccines exist for RSV infection. This study contributes to understanding the molecular mechanism by which PI3K signaling regulates syncytium formation and provides a leading compound for anti-RSV infection drug development.

Bronchial epithelia from adults and children: SARS-CoV-2 spread via syncytia formation and type III interferon infectivity restriction.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections initiate in the bronchi of the upper respiratory tract and are able to disseminate to the lower respiratory tract, where infections can cause an acute respiratory distress syndrome with a high degree of mortality in elderly patients. We used reconstituted primary bronchial epithelia from adult and child donors to follow the SARS-CoV-2 infection dynamics. We show that, in epithelia from adult donors, infections initiate in multiciliated cells and spread within 24 to 48 h throughout the whole epithelia. Syncytia formed of ciliated and basal cells appeared at the apical side of the epithelia within 3 to 4 d and were released into the apical lumen, where they contributed to the transmittable virus dose. A small number of reconstituted epithelia were intrinsically more resistant to virus infection, limiting virus spread to different degrees. This phenotype was more frequent in epithelia derived from children versus adults and correlated with an accelerated release of type III interferon. Treatment of permissive adult epithelia with exogenous type III interferon restricted infection, while type III interferon gene knockout promoted infection. Furthermore, a transcript analysis revealed that the inflammatory response was specifically attenuated in children. Taken together, our findings suggest that apical syncytia formation is an underappreciated source of virus propagation for tissue or environmental dissemination, whereas a robust type III interferon response such as commonly seen in young donors restricted SARS-CoV-2 infection. Thus, the combination of interferon restriction and attenuated inflammatory response in children might explain the epidemiological observation of age-related susceptibility to COVID-19.

Polyploid giant cancer cells, EZH2 and Myc upregulation in mammary epithelial cells infected with high-risk human cytomegalovirus.

BACKGROUND: Human cytomegalovirus (HCMV) infection has been actively implicated in complex neoplastic processes. Beyond oncomodulation, the molecular mechanisms that might underlie HCMV-induced oncogenesis are being extensively studied. Polycomb repressive complex 2 (PRC2) proteins, in particular enhancer of zeste homolog 2 (EZH2) are associated with cancer progression. Nevertheless, little is known about EZH2 activation in the context of HCMV infection and breast oncogenesis. METHODS: Herein, we identified EZH2 as a downstream target for HCMV-induced Myc upregulation upon acute and chronic infection with high-risk strains using a human mammary epithelial model. FINDINGS: We detected polyploidy and CMV-transformed HMECs (CTH) cells harboring HCMV and dynamically undergoing the giant cells cycle. Acquisition of embryonic stemness markers positively correlated with EZH2 and Myc expression. EZH2 inhibitors curtail sustained CTH cells' malignant phenotype. Besides harboring polyploid giant cancer cells (PGCCs), tumorigenic breast biopsies were characterized by an enhanced EZH2 and Myc expression, with a strong positive correlation between EZH2 and Myc expression, and between PGCC count and EZH2/Myc expression in the presence of HCMV. Further, we isolated two HCMV strains from EZH2HighMycHigh basal-like tumors which replicate in MRC5 cells and transform HMECs toward CTH cells after acute infection. INTERPRETATION: Our data establish a potential link between HCMV-induced Myc activation, the subsequent EZH2 upregulation, and polyploidy induction. These data support the proposed tumorigenesis properties of EZH2/Myc, and allow the isolation of two oncogenic HCMV strains from EZH2HighMycHigh basal breast tumors while identifying EZH2 as a potential therapeutic target in the management of breast cancer, particularly upon HCMV infection. FUNDING: This work was supported by grants from the University of Franche-Comté (UFC) (CR3300), the Région Franche-Comté (2021-Y-08292 and 2021-Y-08290) and the Ligue contre le Cancer (CR3304) to Georges Herbein. Zeina Nehme is a recipient of a doctoral scholarship from the municipality of Habbouch. Sandy Haidar Ahmad is recipient of a doctoral scholarship from Lebanese municipality. Ranim El Baba is a recipient of a doctoral scholarship from Hariri foundation for sustainable human development.

Anti-HIV Activity of Snake Venom Phospholipase A2s: Updates for New Enzymes and Different Virus Strains.

Since the beginning of the HIV epidemic, lasting more than 30 years, the main goal of scientists was to develop effective methods for the prevention and treatment of HIV infection. Modern medicines have reduced the death rate from AIDS by 80%. However, they still have side effects and are very expensive, dictating the need to search for new drugs. Earlier, it was shown that phospholipases A2 (PLA2s) from bee and snake venoms block HIV replication, the effect being independent on catalytic PLA2 activity. However, the antiviral activity of human PLA2s against Lentiviruses depended on catalytic function and was mediated through the destruction of the viral membrane. To clarify the role of phospholipolytic activity in antiviral effects, we analyzed the anti-HIV activity of several snake PLA2s and found that the mechanisms of their antiviral activity were similar to that of mammalian PLA2. Our results indicate that snake PLA2s are capable of inhibiting syncytium formation between chronically HIV-infected cells and healthy CD4-positive cells and block HIV binding to cells. However, only dimeric PLA2s had pronounced virucidal and anti-HIV activity, which depended on their catalytic activity. The ability of snake PLA2s to inactivate the virus may provide an additional barrier to HIV infection. Thus, snake PLA2s might be considered as candidates for lead molecules in anti-HIV drug development.

Visualization of Marek's Disease Virus Genomes in Living Cells during Lytic Replication and Latency.

Visualization of the herpesvirus genomes during lytic replication and latency is mainly achieved by fluorescence in situ hybridization (FISH). Unfortunately, this technique cannot be used for the real-time detection of viral genome in living cells. To facilitate the visualization of the Marek's disease virus (MDV) genome during all stages of the virus lifecycle, we took advantage of the well-established tetracycline operator/repressor (TetO/TetR) system. This system consists of a fluorescently labeled TetR (TetR-GFP) that specifically binds to an array of tetO sequences. This tetO repeat array was first inserted into the MDV genome (vTetO). Subsequently, we fused TetR-GFP via a P2a self-cleaving peptide to the C-terminus of the viral interleukin 8 (vIL8), which is expressed during lytic replication and latency. Upon reconstitution of this vTetO-TetR virus, fluorescently labeled replication compartments were detected in the nucleus during lytic replication. After validating the specificity of the observed signal, we used the system to visualize the genesis and mobility of the viral replication compartments. In addition, we assessed the infection of nuclei in syncytia as well as lytic replication and latency in T cells. Taken together, we established a system allowing us to track the MDV genome in living cells that can be applied to many other DNA viruses.

Loss of Detection of sgN Precedes Viral Abridged Replication in COVID-19-Affected Patients-A Target for SARS-CoV-2 Propagation.

The development of prophylactic agents against the SARS-CoV-2 virus is a public health priority in the search for new surrogate markers of active virus replication. Early detection markers are needed to follow disease progression and foresee patient negativization. Subgenomic RNA transcripts (with a focus on sgN) were evaluated in oro/nasopharyngeal swabs from COVID-19-affected patients with an analysis of 315 positive samples using qPCR technology. Cut-off Cq values for sgN (Cq < 33.15) and sgE (Cq < 34.06) showed correlations to high viral loads. The specific loss of sgN in home-isolated and hospitalized COVID-19-positive patients indicated negativization of patient condition, 3-7 days from the first swab, respectively. A new detection kit for sgN, gene E, gene ORF1ab, and gene RNAse P was developed recently. In addition, in vitro studies have shown that 2'-O-methyl antisense RNA (related to the sgN sequence) can impair SARS-CoV-2 N protein synthesis, viral replication, and syncytia formation in human cells (i.e., HEK-293T cells overexpressing ACE2) upon infection with VOC Alpha (B.1.1.7)-SARS-CoV-2 variant, defining the use that this procedure might have for future therapeutic actions against SARS-CoV-2.

Enhanced fusogenicity and pathogenicity of SARS-CoV-2 Delta P681R mutation.

During the current coronavirus disease 2019 (COVID-19) pandemic, a variety of mutations have accumulated in the viral genome of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and, at the time of writing, four variants of concern are considered to be potentially hazardous to human society1. The recently emerged B.1.617.2/Delta variant of concern is closely associated with the COVID-19 surge that occurred in India in the spring of 2021 (ref. 2). However, the virological properties of B.1.617.2/Delta remain unclear. Here we show that the B.1.617.2/Delta variant is highly fusogenic and notably more pathogenic than prototypic SARS-CoV-2 in infected hamsters. The P681R mutation in the spike protein, which is highly conserved in this lineage, facilitates cleavage of the spike protein and enhances viral fusogenicity. Moreover, we demonstrate that the P681R-bearing virus exhibits higher pathogenicity compared with its parental virus. Our data suggest that the P681R mutation is a hallmark of the virological phenotype of the B.1.617.2/Delta variant and is associated with enhanced pathogenicity.

Computationally prioritized drugs inhibit SARS-CoV-2 infection and syncytia formation.

The pharmacological arsenal against the COVID-19 pandemic is largely based on generic anti-inflammatory strategies or poorly scalable solutions. Moreover, as the ongoing vaccination campaign is rolling slower than wished, affordable and effective therapeutics are needed. To this end, there is increasing attention toward computational methods for drug repositioning and de novo drug design. Here, multiple data-driven computational approaches are systematically integrated to perform a virtual screening and prioritize candidate drugs for the treatment of COVID-19. From the list of prioritized drugs, a subset of representative candidates to test in human cells is selected. Two compounds, 7-hydroxystaurosporine and bafetinib, show synergistic antiviral effects in vitro and strongly inhibit viral-induced syncytia formation. Moreover, since existing drug repositioning methods provide limited usable information for de novo drug design, the relevant chemical substructures of the identified drugs are extracted to provide a chemical vocabulary that may help to design new effective drugs.

Short-Stalk Isoforms of CADM1 and CADM2 Trigger Neuropathogenic Measles Virus-Mediated Membrane Fusion by Interacting with the Viral Hemagglutinin.

Measles virus (MeV), an enveloped RNA virus in the family Paramyxoviridae, usually causes acute febrile illness with skin rash but in rare cases persists in the brain, causing a progressive neurological disorder, subacute sclerosing panencephalitis (SSPE). MeV bears two envelope glycoproteins, the hemagglutinin (H) and fusion (F) proteins. The H protein possesses a head domain that initially mediates receptor binding and a stalk domain that subsequently transmits the fusion-triggering signal to the F protein. We recently showed that cell adhesion molecule 1 (CADM1; also known as IGSF4A, Necl-2, and SynCAM1) and CADM2 (also known as IGSF4D, Necl-3, and SynCAM2) are host factors enabling cell-cell membrane fusion mediated by hyperfusogenic F proteins of neuropathogenic MeVs as well as MeV spread between neurons lacking the known receptors. CADM1 and CADM2 interact in cis with the H protein on the same cell membrane, triggering hyperfusogenic F protein-mediated membrane fusion. Multiple isoforms of CADM1 and CADM2 containing various lengths of their stalk regions are generated by alternative splicing. Here, we show that only short-stalk isoforms of CADM1 and CADM2 predominantly expressed in the brain induce hyperfusogenic F protein-mediated membrane fusion. While the known receptors interact in trans with the H protein through its head domain, these isoforms can interact in cis even with the H protein lacking the head domain and trigger membrane fusion, presumably through its stalk domain. Thus, our results unveil a new mechanism of viral fusion triggering by host factors. IMPORTANCE Measles, an acute febrile illness with skin rash, is still an important cause of childhood morbidity and mortality worldwide. Measles virus (MeV), the causative agent of measles, may also cause a progressive neurological disorder, subacute sclerosing panencephalitis (SSPE), several years after acute infection. The disease is fatal, and no effective therapy is available. Recently, we reported that cell adhesion molecule 1 (CADM1) and CADM2 are host factors enabling MeV cell-to-cell spread in neurons. These molecules interact in cis with the MeV attachment protein on the same cell membrane, triggering the fusion protein and causing membrane fusion. CADM1 and CADM2 are known to exist in multiple splice isoforms. In this study, we report that their short-stalk isoforms can induce membrane fusion by interacting in cis with the viral attachment protein independently of its receptor-binding head domain. This finding may have important implications for cis-acting fusion triggering by host factors.

The Mechanism and Consequences of SARS-CoV-2 Spike-Mediated Fusion and Syncytia Formation.

Syncytia are formed when individual cells fuse. SARS-CoV-2 induces syncytia when the viral spike (S) protein on the surface of an infected cell interacts with receptors on neighboring cells. Syncytia may potentially contribute to pathology by facilitating viral dissemination, cytopathicity, immune evasion, and inflammatory response. SARS-CoV-2 variants of concern possess several mutations within the S protein that enhance receptor interaction, fusogenicity and antibody binding. In this review, we discuss the molecular determinants of S mediated fusion and the antiviral innate immunity components that counteract syncytia formation. Several interferon-stimulated genes, including IFITMs and LY6E act as barriers to S protein-mediated fusion by altering the composition or biophysical properties of the target membrane. We also summarize the effect that the mutations associated with the variants of concern have on S protein fusogenicity. Altogether, this review contextualizes the current understanding of Spike fusogenicity and the role of syncytia during SARS-CoV-2 infection and pathology.

Human Cytomegalovirus Replication and Infection-Induced Syncytia Formation in Labial, Foreskin, and Fetal Lung Fibroblasts.

Only a handful of cell types, including fibroblasts, epithelial, and endothelial cells, can support human cytomegalovirus (CMV) replication in vitro, in striking contrast to the situation in vivo. While the susceptibility of epithelial and endothelial cells to CMV infection is strongly modulated by their anatomical site of origin, multiple CMV strains have been successfully isolated and propagated on fibroblasts derived from different organs. As oral mucosal cells are likely involved in CMV acquisition, we sought to evaluate the ability of infant labial fibroblasts to support CMV replication, compared to that of commonly used foreskin and fetal lung fibroblasts. No differences were found in the proportion of cells initiating infection, or in the amounts of viral progeny produced after exposure to the fibroblast-adapted CMV strain AD169 or to the endothelial cell-adapted strain TB40/E. Syncytia formation was, however, significantly enhanced in infected labial and lung fibroblasts compared to foreskin-derived cells, and did not occur after infection with AD169. Together, these data indicate that fibroblast populations derived from different tissues are uniformly permissive to CMV infection but retain phenotypic differences of potential importance for infection-induced cell-cell fusion, and ensuing viral spread and pathogenesis in different organs.

SARS-CoV-2 Alpha, Beta, and Delta variants display enhanced Spike-mediated syncytia formation.

Severe COVID-19 is characterized by lung abnormalities, including the presence of syncytial pneumocytes. Syncytia form when SARS-CoV-2 spike protein expressed on the surface of infected cells interacts with the ACE2 receptor on neighboring cells. The syncytia forming potential of spike variant proteins remain poorly characterized. Here, we first assessed Alpha (B.1.1.7) and Beta (B.1.351) spread and fusion in cell cultures, compared with the ancestral D614G strain. Alpha and Beta replicated similarly to D614G strain in Vero, Caco-2, Calu-3, and primary airway cells. However, Alpha and Beta formed larger and more numerous syncytia. Variant spike proteins displayed higher ACE2 affinity compared with D614G. Alpha, Beta, and D614G fusion was similarly inhibited by interferon-induced transmembrane proteins (IFITMs). Individual mutations present in Alpha and Beta spikes modified fusogenicity, binding to ACE2 or recognition by monoclonal antibodies. We further show that Delta spike also triggers faster fusion relative to D614G. Thus, SARS-CoV-2 emerging variants display enhanced syncytia formation.

Micronucleus production, activation of DNA damage response and cGAS-STING signaling in syncytia induced by SARS-CoV-2 infection.

SARS-CoV-2 infection could cause severe acute respiratory syndrome, largely attributed to dysregulated immune activation and extensive lung tissue damage. However, the underlying mechanisms are not fully understood. Here, we reported that viral infection could induce syncytia formation within cells expressing ACE2 and the SARS-CoV-2 spike protein, leading to the production of micronuclei with an average rate of about 4 per syncytium (> 93%). Remarkably, these micronuclei were manifested with a high level of activation of both DNA damage response and cGAS-STING signaling, as indicated by micronucleus translocation of γH2Ax and cGAS, and upregulation of their respective downstream target genes. Since activation of these signaling pathways were known to be associated with cellular catastrophe and aberrant immune activation, these findings help explain the pathological effects of SARS-CoV-2 infection at cellular and molecular levels, and provide novel potential targets for COVID-19 therapy.

Cell Fusion and Syncytium Formation in Betaherpesvirus Infection.

Cell-cell fusion is a fundamental and complex process that occurs during reproduction, organ and tissue growth, cancer metastasis, immune response, and infection. All enveloped viruses express one or more proteins that drive the fusion of the viral envelope with cellular membranes. The same proteins can mediate the fusion of the plasma membranes of adjacent cells, leading to the formation of multinucleated syncytia. While cell-cell fusion triggered by alpha- and gammaherpesviruses is well-studied, much less is known about the fusogenic potential of betaherpesviruses such as human cytomegalovirus (HCMV) and human herpesviruses 6 and 7 (HHV-6 and HHV-7). These are slow-growing viruses that are highly prevalent in the human population and associated with several diseases, particularly in individuals with an immature or impaired immune system such as fetuses and transplant recipients. While HHV-6 and HHV-7 are strictly lymphotropic, HCMV infects a very broad range of cell types including epithelial, endothelial, mesenchymal, and myeloid cells. Syncytia have been observed occasionally for all three betaherpesviruses, both during in vitro and in vivo infection. Since cell-cell fusion may allow efficient spread to neighboring cells without exposure to neutralizing antibodies and other host immune factors, viral-induced syncytia may be important for viral dissemination, long-term persistence, and pathogenicity. In this review, we provide an overview of the viral and cellular factors and mechanisms identified so far in the process of cell-cell fusion induced by betaherpesviruses and discuss the possible consequences for cellular dysfunction and pathogenesis.

M protein of subacute sclerosing panencephalitis virus, synergistically with the F protein, plays a crucial role in viral neuropathogenicity.

Subacute sclerosing panencephalitis (SSPE) is a rare fatal neurodegenerative disease caused by a measles virus (MV) variant, SSPE virus, that accumulates mutations during long-term persistent infection of the central nervous system (CNS). Clusters of mutations identified around the matrix (M) protein in many SSPE viruses suppress productive infectious particle release and accelerate cell-cell fusion, which are features of SSPE viruses. It was reported, however, that these defects of M protein function might not be correlated directly with promotion of neurovirulence, although they might enable establishment of persistent infection. Neuropathogenicity is closely related to the character of the viral fusion (F) protein, and amino acid substitution(s) in the F protein of some SSPE viruses confers F protein hyperfusogenicity, facilitating viral propagation in the CNS through cell-cell fusion and leading to neurovirulence. The F protein of an SSPE virus Kobe-1 strain, however, displayed only moderately enhanced fusion activity and required additional mutations in the M protein for neuropathogenicity in mice. We demonstrated here the mechanism for the M protein of the Kobe-1 strain supporting the fusion activity of the F protein and cooperatively inducing neurovirulence, even though each protein, independently, has no effect on virulence. The occurrence of SSPE has been estimated recently as one in several thousand in children who acquired measles under the age of 5 years, markedly higher than reported previously. The probability of a specific mutation (or mutations) occurring in the F protein conferring hyperfusogenicity and neuropathogenicity might not be sufficient to explain the high frequency of SSPE. The induction of neurovirulence by M protein synergistically with moderately fusogenic F protein could account for the high frequency of SSPE.