Journal of Virology最新文献

The Vpx protein encoded by HIV-2/simian immunodeficiency virus (SIV) can antagonize the restriction of the host intrinsic restriction factor, SAMHD1, in nondividing cells by promoting its polyubiquitination and subsequent degradation, thereby facilitating viral replication and immune evasion. However, the role of deubiquitinating enzymes (DUBs) in the dynamics of virus and host remains poorly understood. Here, we demonstrate that DUB USP37 significantly reverses the Vpx-mediated degradation of SAMHD1 in various HIV-2/SIV subtypes by interacting with SAMHD1 and removing its ubiquitin chains. Notably, USP37 deubiquitinates SAMHD1 by directly recognizing SAMHD1 rather than by targeting the E3 ubiquitin ligase. The deubiquitinase activity of USP37 and its ubiquitin interacting motifs are essential for the deubiquitination of SAMHD1, whereas the phosphorylation state of USP37 does not influence its activity. Additionally, USP37 enhances the suppression of the retrotransposition of LINE-1 elements by SAMHD1 via stabilizing SAMHD1. Our findings provide important evidence that enhancing the deubiquitinating activity of some DUBs results in the stability of the host restriction factor and might be a viable strategy against HIV/SIV infections.IMPORTANCESAMHD1 is a multifunctional protein, including restricting virus replication, maintaining genomic integrity through DNA repair, modulating the immune response by influencing the production of type I interferons and other cytokines, and affecting cancer cell proliferation and sensitivity to chemotherapy. However, HIV-2/simian immunodeficiency virus (SIV)-encoded Vpx and the host E3 ligase TRIM21 can induce the degradation of SAMHD1 via the ubiquitin-proteasome pathway. Therefore, it is necessary to find the strategy to stabilize SAMHD1. Our study demonstrates that the deubiquitinase USP37 reverses Vpx- and TRIM21-mediated degradation of SAMHD1, thereby inhibiting SIV replication and LINE-1 activity by stabilizing SAMHD1. Thus, we report a novel role of USP37, which represents a potentially useful target for the development of new drugs.

The contributions of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific T cells to vaccine efficacy and durability are unclear. We investigated relationships between mRNA vaccine-induced spike-specific interferon- gamma (IFN-γ) and interleukin-2 (IL-2) T-cell responses and neutralizing antibody development in long-term care home staff doubly vaccinated with BNT162b2 or mRNA-1273. The impacts of pre-existing cross-reactive T-cell immunity on cellular and humoral responses to vaccination were additionally assessed. Mathematical modeling of the kinetics of spike-specific IFN-γ and IL-2 T-cell responses over 6 months post-second dose was bifurcated into recipients who exhibited gradual increases with doubling times of 155 and 167 days or decreases with half-lives of 165 and 132 days, respectively. Differences in kinetics did not correlate with clinical phenotypes. Serological anti-spike IgG, anti-receptor binding domain (RBD) IgG, anti-spike IgA, and anti-RBD IgA antibody levels otherwise decayed in all participants with half-lives of 63, 57, 79, and 46 days, respectively, alongside waning neutralizing capacity (t_1/2 = 408 days). Spike-specific T-cell responses induced at 2-6 weeks positively correlated with live viral neutralization at 6 months post-second dose, especially in hybrid immune individuals. Participants with pre-existing cross-reactive T-cell immunity to SARS-CoV-2 exhibited greater spike-specific T-cell responses, reduced anti-RBD IgA antibody levels, and a trending increase in neutralization at 2-6 weeks post-second dose. Non-spike-specific T-cells predominantly targeted SARS-CoV-2 non-structural protein at 6 months post-second dose in cross-reactive participants. mRNA vaccination was lastly shown to induce off-target T-cell responses against unrelated antigens. In summary, vaccine-induced spike-specific T-cell immunity appeared to influence serological neutralizing capacity, with only a modest effect induced by pre-existing cross-reactivity.

Importance: Our findings provide valuable insights into the potential contributions of mRNA vaccine-induced spike-specific T-cell responses to the durability of neutralizing antibody levels in both uninfected and hybrid immune recipients. Our study additionally sheds light on the precise impacts of pre-existing cross-reactive T-cell immunity to severe acute respiratory syndrome coronavirus 2 on the magnitude and kinetics of cellular and humoral responses to vaccination. Accordingly, our data will help optimize the development of next-generation T cell-based coronavirus vaccines and vaccine regimens to maximize efficacy and durability.

Canine parvovirus type 2 (CPV-2) is a member of the Parvoviridae family, characterized by its small, non-enveloped virions containing a linear single-stranded DNA genome of approximately 5 kb. Parvoviruses entirely reliant on the host cell's division machinery for replication. In this study, we demonstrate that CPV-2 infection triggers the host translation shutoff, a process in which the nonstructural protein 1 (NS1) plays a pivotal role. Our findings indicate that the CPV-2 NS1-induced host translation shutoff is not associated with transcription, protein degradation pathways, or eIFα phosphorylation, but rather involves the reduction of phosphorylation of the mammalian target of rapamycin (mTOR). In conclusion, this research reveals that CPV-2 NS1 induces a host translation shutoff by reducing mTOR phosphorylation, a mechanism that could potentially inform the development of more efficacious control and therapeutic strategies for CPV-2 and other parvoviral infections.

Importance: Autonomous parvoviruses, which possess compact genomes, are obligate intracellular parasites that necessitate host cell division for their replication cycle. Consequently, the modulation of host translation and usurpation of cellular machinery are hypothesized to facilitate immune evasion, enhance viral transmission, and perpetuate long-term infection. Despite the biological significance, the precise mechanisms by which autonomous parvoviruses regulate host translation remain understudied. Our study elucidates that CPV-2 infection induces a shutoff of host translation through the attenuation of mTOR phosphorylation. This mechanism may enable the virus to subvert the host immune response and engender pathogenic effects.

Diaphorina citri transmits Candidatus Liberibacter asiaticus (CLas) between citrus plants which causes the expression of huanglongbing disease in citrus. D. citri flavi-like virus (DcFLV) co-occurs intracellularly with CLas in D. citri populations in the field. However, the impact(s) of DcFLV presence on the insect vector and its interaction with the CLas phytopathogen remain unclear. We compared CLas acquisition and transmission efficiencies as well as transcriptomic expression between viruliferous and non-viruliferous psyllids at multiple life stages. Viruliferous nymphs acquired higher titers of CLas than non-viruliferous nymphs, whereas viruliferous adults acquired less CLas than those without virus. The presence of DcFLV increased the transmission of CLas by both nymphs and adults. Furthermore, RNA-seq and functional gene expression analyses revealed that endoplasmic reticulum stress-, autophagy-, and defense-related genes were significantly upregulated in viruliferous adult psyllids, whereas most of these genes were downregulated in viruliferous nymphs. Our work demonstrates that DcFLV differentially modulates various cellular and physiological functions in D. citri in a life stage-dependent manner and promotes the acquisition of CLas at the nymphal stage and transmission of the pathogen at the adult stage of the vector. Collectively, our results suggest that D. citri vectors with DcFLV exhibit greater pathogen transmission efficiency than those without virus.

Importance: Huanglongbing (HLB), caused by fastidious bacteria from three Candidatus Liberibacter species, is the most damaging disease impacting the citrus industry worldwide. Spread by the Asian citrus psyllid (Diaphorina citri) in Asia and the Americas, HLB causes substantial financial losses, and has reduced citrus production in Florida by more than 90%. Although there are ongoing efforts to limit spread of the disease, effective HLB management remains elusive. Suppressing vector populations and decreasing CLas transmission are the two strategies that need to be urgently improved. Recently, a D. citri flavi-like virus (DcFLV) was characterized within its D. citri host, and it co-occurs intracellularly with CLas in psyllid populations. Here, we show that viruliferous nymphs exhibit higher CLas acquisition than non-viruliferous nymphs. Furthermore, both viruliferous adults and nymphs exhibit increased CLas transmission efficiency. We suggest the possibility of manipulating DcFLV in D. citri populations to reduce CLas transmission for HLB disease management.

Equine infectious anemia virus (EIAV) and HIV-1 are both members of the Lentivirus genus and are similar in virological characters. EIAV is of great concern in the equine industry. Lentiviruses establish a complex interaction with the host cell to counteract the antiviral responses. There are various pattern recognition receptors in the host, for instance, the cytosolic RNA helicases interact with viral RNA to activate the mitochondrial antiviral signaling protein (MAVS) and subsequent interferon (IFN) response. However, viruses also exploit multiple strategies to resist host immunity by targeting MAVS, but the mechanism by which lentiviruses are able to target MAVS has remained unclear. In this study, we found that EIAV infection induced MAVS degradation, and that EIAV Gag protein recruited the E3 ubiquitin ligase Smurf1 to polyubiquitinate and degrade MAVS. The CARD domain of MAVS and the WW domain of Smurf1 are responsible for the interaction with Gag. EIAV Gag is a precursor polyprotein of the membrane-interacting matrix p15, the capsid p26, and the RNA-binding nucleocapsid proteins p11 and p9. Therefore, we analyzed which protein domain of Gag could interact with MAVS and Smurf1. We found that p15 and p26, but not p11 or p9, target MAVS for degradation. Moreover, we identified the key amino acid residues that support the interactions between p15 or p26 and MAVS or Smurf1. The present study describes a novel role of the EIAV structural protein Gag in targeting MAVS to counteract innate immunity, and reveals the mechanism by which the equine lentivirus can antagonize against MAVS.IMPORTANCEHost anti-RNA virus innate immunity relies mainly on the recognition by retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated protein 5 (MDA5), and subsequently initiates downstream signaling through interaction with mitochondrial antiviral signaling protein (MAVS). However, viruses have developed various strategies to counteract MAVS-mediated signaling, although the method of antagonism of MAVS by lentiviruses is still unknown. In this article, we demonstrate that the precursor (Pr55gag) polyprotein of EIAV and its protein domains p15 and p26 target MAVS for ubiquitin-mediated degradation through E3 ubiquitin ligase Smurf1. MAVS degradation leads to the inhibition of the downstream IFN-β pathway. This is the first time that lentiviral structural protein has been found to have antagonistic effects on MAVS pathway. Overall, our study reveals a novel mechanism by which equine lentiviruses can evade host innate immunity, and provides insight into potential therapeutic strategies for the control of lentivirus infection.

Herpesviruses carry an assortment of proteins in the interstitial space between the capsid and membrane envelope, collectively referred to as the tegument. Upon virion fusion with a cell, envelope integrity is disrupted, and many tegument constituents disperse into the cytosol to carry out individual effector functions, while others direct transport of the capsid to the nucleus. To gain insight into the tegument dynamics that occur with disruption of envelope integrity, we used a combination of single-particle fluorescence and biochemical approaches that leveraged the previously established use of n-ethylmaleimide to inhibit virion dynamics. We document that the large tegument protein (pUL36), which is stably bound to the capsid surface at its C-terminus, is also conditionally bound to the capsid via its N-terminal deubiquitinase (DUB) domain. The DUB is released, while remaining tethered to the capsid by the pUL36 C-terminus, by a mechanism dependent on reactive cysteines. Mutation of these cysteines locks the DUB in a capsid bound state and suppresses enzymatic activity.

Importance: Neuroinvasive alphaherpesviruses, such as herpes simplex virus and pseudorabies virus, cause a broad range of diseases in humans and other animals. Novel strategies to interfere with the virion structural rearrangements required for infectivity could prove valuable to treat infections, yet critical aspects of the virion architecture and its metastability remain poorly defined. In this study, we document that the pUL36 tegument protein exhibits conditional capsid binding in its N-terminal deubiquitinase domain that regulates enzymatic activity during infection.

H5 highly pathogenic avian influenza (HPAI) viruses of the A/Goose/Guangdong/1/96 (Gs/Gd) lineage continue to evolve and cause outbreaks in domestic poultry and wild birds, with sporadic spillover infections in mammals. The global spread of clade 2.3.4.4b viruses via migratory birds since 2020 has facilitated the introduction of novel reassortants to China, where avian influenza of various subtypes have been epizootic or enzootic among domestic birds. To determine the impact of clade 2.3.4.4b re-introduction on local HPAI dynamics, we analyzed the genetic diversity of H5N6 and H5N8 detected from monthly poultry market surveillance in Guangdong, China, between 2020 and 2022. Our findings reveal that H5N6 viruses clustered in clades 2.3.4.4b and 2.3.4.4h, while H5N8 viruses were exclusively clustered in clade 2.3.4.4b. After 2020, the re-introduced clade 2.3.4.4b viruses replaced the clade 2.3.4.4h viruses detected in 2020. The N6 genes were divided into two clusters, distinguished by an 11 amino acid deletion in the stalk region, while the N8 genes clustered with clade 2.3.4.4 H5N8 viruses circulating among wild birds. Genomic analysis identified 10 transient genotypes. H5N6, which was more prevalently detected, was also clustered into more genotypes than H5N8. Specifically, H5N6 isolates contained genes derived from HPAI H5Nx viruses and low pathogenic avian influenza in China, while the H5N8 isolates contained genes derived from HPAI A(H5N8) 2.3.4.4b and A(H5N1) 2.3.2.1c. No positive selection on amino acid residues associated with mammalian adaptation was found. Our results suggest expanded genetic diversity of H5Nx viruses in China since 2021 with increasing challenges for pandemic preparedness.IMPORTANCESince 2016/2017, clade 2.3.4.4b H5Nx viruses have spread via migratory birds to all continents except Oceania. Here, we evaluated the impact of the re-introduction of clade of 2.3.4.4b on highly pathogenic avian influenza (HPAI) virus genetic diversity in China. Twenty-two H5N6 and H5N8 HPAI isolated from monthly surveillance in two poultry markets in Guangdong between 2020 and 2022 were characterized. Our findings showed that clade 2.3.4.4h, detected in 2020, was replaced by clade 2.3.4.4b in 2021-2022. H5N6 (n = 18) were clustered into more genotypes than H5N8 (n = 4), suggesting that H5N6 may possess better replication fitness in poultry. Conversely, the H5N8 genotypes are largely derived from the clade 2.3.4.4b wild bird isolates. As clade 2.3.4.4b continues to spread via migratory birds, it is anticipated that the genetic diversity of H5N6 viruses circulating in China may continue to expand in the coming years. Continuous efforts in surveillance, genetic analysis, and risk assessment are therefore crucial for pandemic preparedness.

Bacteriophages are viruses infecting bacteria. The vast majority of them bear a tail, allowing host recognition, cell wall perforation, and DNA injection into the host cytoplasm. Using electron cryo-microscopy (cryo-EM) and single particle analysis, we determined the organization of the tail proximal extremity of siphophage T5 that possesses a long flexible tail and solved the structure of its tail terminator protein p142 (TrP₁₄₂). It allowed us to confirm the common evolutionary origin between T5 TrP_p142 and other known or putative TrPs from siphophages, myophages, and bacterial tail-like machines, despite very poor sequence conservation. By also determining the structure of the T5 tail proximal extremity after interaction with T5 bacterial receptor FhuA, we showed that no conformational changes occur in TrP_p142 and confirmed that the infection signal transduction is not carried by the tube itself. We also investigated the location of T5 Neck1 or tail completion protein p143 (TCP_p143) and showed, thanks to a combination of cryo-EM and structure prediction using Alphafold2, that it is not located at the capsid-to-tail interface as suggested by its position in the genome, but instead, very unexpectedly, on the side of T5 tail tip, and that it appears to be monomeric. Based on structure comparison with other putative TCPs predicted structures, this feature could not be shared by other TCPs and questions the affiliation of p143 to this family of protein.IMPORTANCEBacteriophages, viruses infecting bacteria, are the most abundant living entities on Earth. They are present in all ecosystems where bacteria develop and are instrumental in the regulation, diversity, evolution, and pathogeny of microbial populations. Moreover, with the increasing number of pathogenic strains resistant to antibiotics, virulent phages are considered a serious alternative or complement to classical treatments. 96% of all phages present a tail that allows host recognition and safe channeling of the DNA to the host cytoplasm. We present the atomic model of the proximal extremity of the siphophage T5 tail, confirming structural similarities with other phages. This structure, combined with results previously published and further explored, also allowed a review and a discussion on the role and localization of a mysterious tail protein, the tail completion protein, which is known to be present in the phage tails, but that was never identified in a phage structure.

Following reactivation of a latent alphaherpesvirus infection, viral particles are assembled in neuronal cell bodies, trafficked anterogradely within axons to nerve termini, and spread to adjacent epithelial cells. The virally encoded membrane proteins US9p and the glycoprotein heterodimer gE/gI of pseudorabies virus (PRV) and herpes simplex virus type 1 (HSV-1) play critical roles in anterograde spread, likely as a tripartite gE/gI-US9p complex. Two kinesin motors, kinesin-1 and kinesin-3, are implicated in the egress of these viruses, but how gE/gI-US9p coordinates their activities is poorly understood. Here, we report that PRV, in addition to associating with the kinesin-3 motor KIF1A, recruits the neuronal kinesin-1 isoforms KIF5A and KIF5C, but not the broadly expressed isoform KIF5B, during egress from differentiated CAD neurons. Similarly, in the axons of dorsal root ganglia (DRG)-derived sensory neurons, PRV colocalized with KIF5C but not KIF5B. In differentiated CAD cells, the association of KIF1A with egressing PRV was dependent upon US9p, whereas the recruitment of KIF5 isoforms required gE/gI. Consistent with these findings, the number of PRV particles trafficking within CAD neurites and the axons of DRG neurons increased when kinesin-1 motor activity was upregulated by hyperacetylating microtubules using trichostatin A (TSA) or tubacin, and this enhanced trafficking depended upon the presence of gE/gI. We propose that, following its recruitment by US9p, KIF1A delivers PRV particles to a location where KIF5 motors are subsequently added by a gE/gI-dependent mechanism. KIF5A/C isoforms then serve to traffic viral particles along axons, resulting in characteristic recrudescent infection.

Importance: Alphaherpesviruses include important human and veterinary pathogens that share a unique propensity to establish life-long latent infections in the peripheral nervous system. Upon reactivation, these viruses navigate back to body surfaces and transmit to new hosts. In this study, we demonstrate that the virus gE/gI-US9p membrane complex routes virus particles down this complex neuronal egress pathway by coordinating their association with multiple kinesin microtubule motors.