Journal of General Virology最新文献

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nonstructural protein 15 (Nsp15) is a conserved uridine-specific endoribonuclease (EndoU) and is implicated in innate immune evasion, yet its precise molecular mechanism remains incompletely understood. Here, we demonstrate that Nsp15 limits antiviral innate immune responses in part by downregulating the cGAS-STING pathway. To investigate how Nsp15 antagonizes host innate immune responses, we engineered recombinant SARS-CoV-2 bearing WT or EndoU-inactive Nsp15 (H234A). Compared with the WT virus, the Nsp15-H234A mutant exhibited a 2-log decrease in peak viral titres in interferon (IFN)-competent A549-ACE2 cells, but not in their STAT1 knockout counterparts. This attenuation was partially reversed by STING knockout or STING inhibitors, highlighting STING's involvement in Nsp15-driven immune evasion. Transcriptomic analyses revealed the upregulation of IFNs and IFN-stimulated genes in cells infected with the Nsp15-H234A mutant virus. Notably, cGAS and STING transcripts and proteins were suppressed during WT but not mutant virus infections - even in STAT1-deficient cells - suggesting that Nsp15 contributes directly to their downregulation. In vitro, Nsp15 targeted cGAS and STING transcripts in an EndoU activity-dependent manner, thereby reducing cGAS-STING-driven IFN-β and NF-κB reporter activation. Infection of Syrian hamsters confirmed that the Nsp15-H234A mutant virus replicated to lower titres in respiratory tissues and elicited stronger expression of innate immune-related genes. Collectively, these findings define a key strategy by which SARS-CoV-2 Nsp15 subverts the cGAS-STING pathway to facilitate viral replication and immune evasion and underscore Nsp15 EndoU activity as a potential target for future coronavirus antiviral or vaccine design.

The guinea pig with guinea pig cytomegalovirus (GPCMV) is the only small-animal model for congenital cytomegalovirus, a leading cause of cognitive impairment and hearing loss in newborns. GPCMV encodes human cytomegalovirus (HCMV) homologues of viral entry glycoprotein complexes, which are neutralizing-antibody vaccine targets. As with HCMV, GPCMV has two pathways of cell entry (direct and endocytic). Specific viral gH/gL-based complexes are necessary for receptor interaction and cell entry: gH/gL/gO trimer (direct) and pentamer complex (PC) (endocytic). Both pathways also require gB as the fusogenic protein. Direct GPCMV cell entry requires platelet-derived growth factor receptor alpha (PDGFRA), but an endocytic PC receptor remains unknown. We hypothesized that cellular knockout of direct and endocytic receptors would completely block infection, which cannot be achieved by gB-based antibodies. Candidate receptors including neuropilin proteins (NRP1, NRP2) and CD147 present on all established guinea pig cell lines were selected based on importance as common virus receptors or in fetal development. Results demonstrated that NRP2 interacted with PC, unlike NRP1 or CD147, in immunoprecipitation assays and eliminated NRP1/NRP2 heterodimer receptor interaction. The viral trimer only interacted with PDGFRA. Double knockout of PDGFRA/NRP2 completely blocked GPCMV infection. In contrast, the CD147/PDGFRA double knockout had limited GPCMV inhibition, and the single knockout of CD147 had no impact. Knockout of the various receptors had no effect on control HSV-1 infection. Ectopic expression of guinea pig cell receptors restored GPCMV infection but not human NRP2/PDGFRA, indicating a basis for the species-specific barrier for GPCMV and HCMV infection. Overall, results increase the translational relevance of GPCMV for the development of CMV intervention strategies.

Processing bodies (P-bodies/PBs) are non-membranous foci involved in coordinating RNA fate by regulating translation and mRNA decay. In this study, we characterize the anti-viral factor Shiftless (SHFL) as a potent disruptor of PB dynamics. We show SHFL expression restricts PB accumulation even in the context of oxidative stress, suggesting that SHFL expression impedes PB formation. Mutational approaches revealed that SHFL RNA-binding activity is not required to restrict PB formation. However, we have identified a new region of SHFL, a bridge between two distant SHFL domains, as necessary for SHFL-mediated PB disruption. Furthermore, we show that SHFL's ability to disrupt PB formation also impacts its anti-viral activity during infection by the gammaherpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV). While WT SHFL efficiently restricts KSHV lytic reactivation, SHFL mutants defective in PB disruption no longer restrict KSHV reactivation. SHFL-mediated PB disruption also leads to increased expression of several anti-viral cytokines, further emphasizing the connection among SHFL, PB dynamics and the SHFL-dependent anti-viral state. Taken together, our observations suggest a role of SHFL in inhibiting PB formation to restrict KSHV lytic replication, reinforcing the importance of crosstalk between RNA fate and the innate immune response to viral infection.

Understanding the genomic diversity of the Mpox (formerly known as 'monkeypox') virus (MPXV) is important for monitoring viral evolution and dissemination. We encountered three clinical cases in our hospital during the outbreak period of Mpox in 2023 in Japan. The present study aims to report the largest genomic rearrangement observed to date in one of the clinical isolates and to demonstrate its viability in cell culture. Whole-genome sequencing and digital PCR were used to characterize the viral genomes. Viral isolation, microscopic observation and growth kinetics in Vero cells were performed to confirm viral replication. All three patients were men living with human immunodeficiency virus (HIV) and presented typical Mpox symptoms, such as rash, fever and pustules on the body surfaces, including near the genitals. Virus isolation was successful in all three cases. All viral strains belonged to clade IIb, lineage C.1. Notably, one strain exhibited a large-scale genomic rearrangement: a 5.5 kb deletion at the left variable region replaced by a 30.5 kb inverted sequence, the largest reported in clinical isolates. Despite this extensive genomic change, the strain maintained robust replication capacity and marked fusogenicity in vitro. We report, for the first time in clinical isolates, a massive genomic rearrangement in MPXV that does not impair viral replicability. This finding represents an example of genomic plasticity and provides a rare but noteworthy resource for future studies. Taken together, when performing genomic analyses of MPXV, aberrant genomic rearrangements should also be carefully considered alongside single-base substitutions.

Hepatitis C virus (HCV) is often associated with chronic liver diseases and significant alterations in host cellular signalling. However, the molecular mechanisms underlying HCV-related liver pathogenesis remain to be elucidated. The Hippo signalling pathway, a key regulator of cell proliferation and survival, plays a critical role in maintaining liver homeostasis. Here, we investigated the role of the Hippo pathway in HCV-related pathogenesis. We demonstrated that HCV infection induces degradation of LATS1, a key regulator of the Hippo pathway. Degradation of LATS1 protein was restored by a proteasomal inhibitor, but not a lysosome inhibitor, indicating that HCV promotes proteasomal degradation of LATS1 protein. HCV-induced degradation of LATS1 protein was suppressed in si-Itch-transfected Huh-7.5 cells. These results suggest that Itch ubiquitin ligase is involved in ubiquitin-dependent degradation of LATS1 protein. Cell fractionation assays and immunofluorescence staining revealed that HCV infection promoted nuclear translocation of YAP1 protein, suggesting that HCV infection suppresses the Hippo pathway. Furthermore, the transcription of YAP1 target genes, CYR61 and CTGF, that are involved in tissue remodelling and proliferation, was upregulated in HCV-infected Huh-7.5 cells and in HCV-infected patients. Taken together, we propose that HCV promotes the ubiquitin-dependent proteasomal degradation of LATS1 protein, leading to suppression of the Hippo pathway, thereby upregulating transcription of CYR61 and CTGF genes, which may contribute to HCV-related pathogenesis.

The family Nipumfusiviridae includes DNA viruses with hosts deduced to be marine ammonia-oxidizing archaea, specifically those in the archaeal family Nitrosopumilaceae. Virus genomes have been discovered through metagenomics of samples from inlets, coastal areas, intertidal zones, epipelagic and oceanic waters, and soil. Viruses have neither been isolated nor enriched through experiments. The family Nipumfusiviridae includes several genera. The virions of nipumfusiviruses are predicted to have spindle-shaped morphology based on the analysis of the deduced structural models of the major capsid protein. Limited information can be provided about translation and replication from the genome. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Nipumfusiviridae, which is available at ictv.global/report/nipumfusiviridae.

Cyvirus cyprinidallo2 (CyHV-2) is an alloherpesvirus and the causative agent of herpesviral haematopoietic necrosis in goldfish. Whole-genome sequence comparison of the developed live-attenuated vaccine P7-P8 with virulent CyHV-2 strains revealed seven single-nucleotide polymorphisms, five deletions and one inversion in the ORFs, which may be involved in attenuation. A start codon loss in ORF 113, a putative apoptosis-inhibition gene, was observed in the mutations. In vitro assays indicated that apoptosis-related genes were upregulated in cells inoculated with the vaccine or virulent virus compared to uninfected cells. However, the vaccine group showed increased phosphatidylserine externalization and DNA damage, suggesting the apoptosis-inducing properties of P7-P8. In the in vivo experiment, histopathology demonstrated that vaccinated goldfish exhibited immune responses, such as leucocyte aggregation and melanomacrophage centre formation, without marked degeneration. Gene expression analysis showed upregulation of proinflammatory and granzyme B genes in vaccinated fish. In addition, the vaccine strain triggered apoptosis of the infected cells during the early stage of infection, potentially promoting virus clearance and preventing excessive virus replication. The results show that the P7-P8 potentially induces apoptosis and immune responses, contributing to low virus propagation without tissue damage. This study provides insights into CyHV-2 pathogenesis, suggesting that apoptosis-related genes can be the targets for vaccine development against alloherpesviruses in aquaculture species.

Curly top disease (CTD) affects sugar beet, tomato and other crops, resulting in stunted plants with severely curled leaves and reduced yields. The disease occurs worldwide and is caused by geographically associated monopartite geminiviruses transmitted mostly by leafhoppers. However, the aetiology of CTD in South America remains unknown because the disease disappeared in the 1950s. Here, we describe how the chance finding of tomato plants with CTD-like symptoms in Brazil in 2016 and high-throughput sequencing helped identify a novel ~2.6 kb geminivirus DNA associated with curly top disease in Mato Grosso (GV-CTD-MT) that induced CTD symptoms in agroinoculated tomato and Nicotiana benthamiana Domin. plants and produced geminivirus-like virions in infected plants. Evidence GV-CTD-MT may be the genomic DNA of the historic Brazilian curly top virus (BraCTV) includes (i) occurring in the same geographic location (Brazil); (ii) inducing nearly identical CTD symptoms in tobacco (Nicotiana tabacum L.) and tomato plants to those described in the 1940s for the BraCTV; (iii) inducing CTD in a similar broad host range of plant species as previously reported for BraCTV based on leafhopper transmission experiments; (iv) the co-phylogenetic analysis predicting the vector of GV-CTD-MT is Agallia sp. leafhoppers; and, importantly, (v) transmission experiments showing Agallia albidula, the known vector of BraCTV, is a competent vector of GV-CTD-MT. Sequence and phylogenetic analyses revealed that this novel CTD-inducing geminivirus has a chimeric genome, a long evolutionary history, and is closely related to monopartite geminiviruses recently identified in South America. These viruses were placed in the new genus Topilevirus, the fifth genus whose members induce CTD. Thus, our results suggest that BraCTD, which disappeared over 70 years ago, and possibly historic CTDs of sugar beet in South America were caused by topileviruses transmitted by indigenous leafhoppers, thereby solving the conundrum left by first-generation researchers.

Feline morbillivirus (FeMV), first identified in 2012, is a member of the genus Morbillivirus. Like other morbilliviruses, FeMV utilizes signalling lymphocytic activation molecule (CD150 or SLAMF1) as a cellular receptor; however, it is genetically distinct and notable for its unique tissue tropism, particularly its ability to establish persistent infections in the feline kidney. In this study, we established a plaque assay system using Vero cells stably expressing feline SLAMF1 and quantitatively evaluated neutralizing antibodies against FeMV in the sera of domestic cats. The results showed that FeMV-infected feline sera contained exceptionally high titres of FeMV-specific neutralizing antibodies, often far exceeding those observed in animals infected with other morbilliviruses. Importantly, sera with high neutralizing activity against FeMV exhibited undetectable cross-neutralizing activity against other morbilliviruses. In contrast, significant cross-neutralization was observed among non-FeMV morbilliviruses. These findings indicate that FeMV surface glycoproteins are antigenically distinct, underscoring the immunological uniqueness of FeMV. Moreover, while feline SLAMF1 functions as a receptor for various morbilliviruses, FeMV utilized only feline SLAMF1. These data suggest that the receptor-binding domain of FeMV is uniquely adapted to the feline SLAMF1. This study offers valuable tools and insights for FeMV research, advancing our understanding of its antigenicity and host receptor usage.

Prion diseases manifest clinically in three different forms. Sporadic and infectious forms of prion disease are caused by the conversion of WT, cellular prion protein (PrP^C) into its pathogenic conformer (PrP^Sc). In contrast, genetic forms of prion diseases are caused by mutations in the PrP sequence that promote mutant PrP^Sc formation. When reconstituted with either polyanionic or lipid cofactors, purified PrP^C substrate can be converted in vitro into PrP^Sc products that display high levels of specific infectivity when inoculated in WT hosts. In contrast, various protein-only PrP^Sc molecules formed in the absence of cofactors display much lower levels of specific infectivity. Here, we report that protein-only PrP^Sc molecules with different sequences can induce the formation of proteinase K-resistant PrP^Sc molecules and spongiform degeneration in the brains of knock-in mice expressing PrP harbouring the pathogenic E200K mutation, but not in hosts expressing WT PrP. These results indicate that the E200K mutation enhances host susceptibility to various protein-only PrP^Sc fibrils, suggesting fundamental differences in the replication mechanisms of WT versus mutant prions.