Zebrafish TMEM47 is an effective blocker of IFN production during RNA and DNA virus infection.

IMPORTANCE: Mitochondrial antiviral signaling protein (MAVS) and stimulator of interferon (IFN) genes (STING) are key adaptor proteins required for innate immune responses to RNA and DNA virus infection. Here, we show that zebrafish transmembrane protein 47 (TMEM47) plays a critical role in regulating MAVS- and STING-triggered IFN production in a negative feedback manner. TMEM47 interacted with MAVS and STING for autophagic degradation, and ATG5 was essential for this process. These findings suggest the inhibitory function of TMEM47 on MAVS- and STING-mediated signaling responses during RNA and DNA virus infection.

Coronaviruses exploit a host cysteine-aspartic protease for replication.

Highly pathogenic coronaviruses, including severe acute respiratory syndrome coronavirus 2 (refs. 1,2) (SARS-CoV-2), Middle East respiratory syndrome coronavirus3 (MERS-CoV) and SARS-CoV-1 (ref. 4), vary in their transmissibility and pathogenicity. However, infection by all three viruses results in substantial apoptosis in cell culture5-7 and in patient tissues8-10, suggesting a potential link between apoptosis and pathogenesis of coronaviruses. Here we show that caspase-6, a cysteine-aspartic protease of the apoptosis cascade, serves as an important host factor for efficient coronavirus replication. We demonstrate that caspase-6 cleaves coronavirus nucleocapsid proteins, generating fragments that serve as interferon antagonists, thus facilitating virus replication. Inhibition of caspase-6 substantially attenuates lung pathology and body weight loss in golden Syrian hamsters infected with SARS-CoV-2 and improves the survival of mice expressing human DPP4 that are infected with mouse-adapted MERS-CoV. Our study reveals how coronaviruses exploit a component of the host apoptosis cascade to facilitate virus replication.

African Swine Fever Virus EP364R and C129R Target Cyclic GMP-AMP To Inhibit the cGAS-STING Signaling Pathway.

African swine fever virus (ASFV) is a highly pathogenic swine DNA virus with high mortality that causes African swine fever (ASF) in domestic pigs and wild boars. For efficient viral infection, ASFV has developed complex strategies to evade key components of antiviral innate immune responses. However, the immune escape mechanism of ASFV remains unclear. Upon ASFV infection, cyclic GMP-AMP (2',3'-cGAMP) synthase (cGAS), a cytosolic DNA sensor, recognizes ASFV DNA and synthesizes the second messenger 2',3'-cGAMP, which triggers interferon (IFN) production to interfere with viral replication. In this study, we demonstrated a novel immune evasion mechanism of ASFV EP364R and C129R, which blocks cellular cyclic 2',3'-cGAMP-mediated antiviral responses. ASFV EP364R and C129R with nuclease homology inhibit IFN-mediated responses by specifically interacting with 2',3'-cGAMP and exerting their phosphodiesterase (PDE) activity to cleave 2',3'-cGAMP. Particularly notable is that ASFV EP364R had a region of homology with the stimulator of interferon genes (STING) protein containing a 2',3'-cGAMP-binding motif and point mutations in the Y76S and N78A amino acids of EP364R that impaired interaction with 2',3'-cGAMP and restored subsequent antiviral responses. These results highlight a critical role for ASFV EP364R and C129R in the inhibition of IFN responses and could be used to develop ASFV live attenuated vaccines. IMPORTANCE African swine fever (ASF) is a highly contagious hemorrhagic disease in domestic pigs and wild boars caused by African swine fever virus (ASFV). ASF is a deadly epidemic disease in the global pig industry, but no drugs or vaccines are available. Understanding the pathogenesis of ASFV is essential to developing an effective live attenuated ASFV vaccine, and investigating the immune evasion mechanisms of ASFV is crucial to improve the understanding of its pathogenesis. In this study, for the first time, we identified the EP364R and C129R, uncharacterized proteins that inhibit type I interferon signaling. ASFV EP364R and C129R specifically interacted with 2',3'-cGAMP, the mammalian second messenger, and exerted phosphodiesterase activity to cleave 2',3'-cGAMP. In this study, we discovered a novel mechanism by which ASFV inhibits IFN-mediated antiviral responses, and our findings can guide the understanding of ASFV pathogenesis and the development of live attenuated ASFV vaccines.

Fifty Shades of Erns: Innate Immune Evasion by the Viral Endonucleases of All Pestivirus Species.

The genus Pestivirus, family Flaviviridae, includes four historically accepted species, i.e., bovine viral diarrhea virus (BVDV)-1 and -2, classical swine fever virus (CSFV), and border disease virus (BDV). A large number of new pestivirus species were identified in recent years. A common feature of most members is the presence of two unique proteins, Npro and Erns, that pestiviruses evolved to regulate the host's innate immune response. In addition to its function as a structural envelope glycoprotein, Erns is also released in the extracellular space, where it is endocytosed by neighboring cells. As an endoribonuclease, Erns is able to cleave viral ss- and dsRNAs, thus preventing the stimulation of the host's interferon (IFN) response. Here, we characterize the basic features of soluble Erns of a large variety of classified and unassigned pestiviruses that have not yet been described. Its ability to form homodimers, its RNase activity, and the ability to inhibit dsRNA-induced IFN synthesis were investigated. Overall, we found large differences between the various Erns proteins that cannot be predicted solely based on their primary amino acid sequences, and that might be the consequence of different virus-host co-evolution histories. This provides valuable information to delineate the structure-function relationship of pestiviral endoribonucleases.

The parapoxvirus Orf virus ORF116 gene encodes an antagonist of the interferon response.

Orf virus (ORFV) is the type species of the Parapoxvirus genus of the Poxviridae family. Genetic and functional studies have revealed ORFV has multiple immunomodulatory genes that manipulate innate immune responses, during the early stage of infection. ORF116 is a novel gene of ORFV with hitherto unknown function. Characterization of an ORF116 deletion mutant showed that it replicated in primary lamb testis cells with reduced levels compared to the wild-type and produced a smaller plaque phenotype. ORF116 was shown to be expressed prior to DNA replication. The potential function of ORF116 was investigated by gene-expression microarray analysis in HeLa cells infected with wild-type ORFV or the ORF116 deletion mutant. The analysis of differential cellular gene expression revealed a number of interferon-stimulated genes (ISGs) differentially expressed at either 4 or 6 h post infection. IFI44 showed the greatest differential expression (4.17-fold) between wild-type and knockout virus. Other ISGs that were upregulated in the knockout included RIG-I, IFIT2, MDA5, OAS1, OASL, DDX60, ISG20 and IFIT1 and in addition the inflammatory cytokine IL-8. These findings were validated by infecting HeLa cells with an ORF116 revertant recombinant virus and analysis of transcript expression by quantitative real time-PCR (qRT-PCR). These observations suggested a role for the ORFV gene ORF116 in modulating the IFN response and inflammatory cytokines. This study represents the first functional analysis of ORF116.

Pseudorabies Virus Inhibits Type I and Type III Interferon-Induced Signaling via Proteasomal Degradation of Janus Kinases.

Both type I and III interferons (IFNs) play a crucial role in host antiviral response by activating the JAK/STAT (Janus kinase/signal transducer and activator of transcription) signaling pathway to trigger the expression of antiviral IFN-stimulated genes (ISGs). We report that the porcine alphaherpesvirus pseudorabies virus (PRV) triggers proteasomal degradation of the key Janus kinases Jak1 and to a lesser extent Tyk2, thereby inhibiting both type I and III IFN-induced STAT1 phosphorylation and suppressing IFN-induced expression of ISGs. UV-inactivated PRV did not interfere with IFN signaling. In addition, deletion of the EP0 gene from the PRV genome or inhibition of viral genome replication did not affect PRV-induced inhibition of IFN signaling. To our knowledge, this is the first report describing Janus kinase degradation by alphaherpesviruses. These findings thus reveal a novel alphaherpesvirus evasion mechanism of type I and type III IFNs. IMPORTANCE Type I and III interferons (IFNs) trigger signaling via Janus kinases that phosphorylate and activate signal transducer and activator of transcription (STAT) transcription factors, leading to the expression of antiviral interferon-stimulated genes (ISGs) that result in an antiviral state of host cells. Viruses have evolved various mechanisms to evade this response. Our results indicate that an alphaherpesvirus, the porcine pseudorabies virus (PRV), inhibits both type I and III IFN signaling pathways by triggering proteasome-dependent degradation of the key Janus kinases Jak1 and Tyk2 and consequent inhibition of STAT1 phosphorylation and suppression of ISG expression. Moreover, we found that this inhibition is not caused by incoming virions and does not depend on expression of the viral EP0 protein or viral true late proteins. These data for the first time address alphaherpesvirus evasion of type III IFN-mediated signaling and reveal a previously uncharacterized alphaherpesvirus mechanism of IFN evasion via proteasomal degradation of Janus kinases.

A Comparative Analysis of Coronavirus Nucleocapsid (N) Proteins Reveals the SADS-CoV N Protein Antagonizes IFN-ß Production by Inducing Ubiquitination of RIG-I.

Coronaviruses (CoVs) are a known global threat, and most recently the ongoing COVID-19 pandemic has claimed more than 2 million human lives. Delays and interference with IFN responses are closely associated with the severity of disease caused by CoV infection. As the most abundant viral protein in infected cells just after the entry step, the CoV nucleocapsid (N) protein likely plays a key role in IFN interruption. We have conducted a comprehensive comparative analysis and report herein that the N proteins of representative human and animal CoVs from four different genera [swine acute diarrhea syndrome CoV (SADS-CoV), porcine epidemic diarrhea virus (PEDV), severe acute respiratory syndrome CoV (SARS-CoV), SARS-CoV-2, Middle East respiratory syndrome CoV (MERS-CoV), infectious bronchitis virus (IBV) and porcine deltacoronavirus (PDCoV)] suppress IFN responses by multiple strategies. In particular, we found that the N protein of SADS-CoV interacted with RIG-I independent of its RNA binding activity, mediating K27-, K48- and K63-linked ubiquitination of RIG-I and its subsequent proteasome-dependent degradation, thus inhibiting the host IFN response. These data provide insight into the interaction between CoVs and host, and offer new clues for the development of therapies against these important viruses.

SARS-CoV-2 Accessory Proteins in Viral Pathogenesis: Knowns and Unknowns.

There are still many unanswered questions concerning viral SARS-CoV-2 pathogenesis in COVID-19. Accessory proteins in SARS-CoV-2 consist of eleven viral proteins whose roles during infection are still not completely understood. Here, a review on the current knowledge of SARS-CoV-2 accessory proteins is summarized updating new research that could be critical in understanding SARS-CoV-2 interaction with the host. Some accessory proteins such as ORF3b, ORF6, ORF7a and ORF8 have been shown to be important IFN-I antagonists inducing an impairment in the host immune response. In addition, ORF3a is involved in apoptosis whereas others like ORF9b and ORF9c interact with cellular organelles leading to suppression of the antiviral response in infected cells. However, possible roles of ORF7b and ORF10 are still awaiting to be described. Also, ORF3d has been reassigned. Relevant information on the knowns and the unknowns in these proteins is analyzed, which could be crucial for further understanding of SARS-CoV-2 pathogenesis and to design strategies counteracting their actions evading immune responses in COVID-19.

Interfering With Inflammation: Heterogeneous Effects of Interferons in Graft-Versus-Host Disease of the Gastrointestinal Tract and Inflammatory Bowel Disease.

The intestine can be the target of several immunologically mediated diseases, including graft-versus-host disease (GVHD) and inflammatory bowel disease (IBD). GVHD is a life-threatening complication that occurs after allogeneic hematopoietic stem cell transplantation. Involvement of the gastrointestinal tract is associated with a particularly high mortality. GVHD development starts with the recognition of allo-antigens in the recipient by the donor immune system, which elicits immune-mediated damage of otherwise healthy tissues. IBD describes a group of immunologically mediated chronic inflammatory diseases of the intestine. Several aspects, including genetic predisposition and immune dysregulation, are responsible for the development of IBD, with Crohn's disease and ulcerative colitis being the two most common variants. GVHD and IBD share multiple key features of their onset and development, including intestinal tissue damage and loss of intestinal barrier function. A further common feature in the pathophysiology of both diseases is the involvement of cytokines such as type I and II interferons (IFNs), amongst others. IFNs are a family of protein mediators produced as a part of the inflammatory response, typically to pathogens or malignant cells. Diverse, and partially paradoxical, effects have been described for IFNs in GVHD and IBD. This review summarizes current knowledge on the role of type I, II and III IFNs, including basic concepts and controversies about their functions in the context of GVHD and IBD. In addition, therapeutic options, research developments and remaining open questions are addressed.

Rotavirus NSP1 Inhibits Type I and Type III Interferon Induction.

Type I interferons (IFNs) are produced by most cells in response to virus infection and stimulate a program of anti-viral gene expression in neighboring cells to suppress virus replication. Type III IFNs have similar properties, however their effects are limited to epithelial cells at mucosal surfaces due to restricted expression of the type III IFN receptor. Rotavirus (RV) replicates in intestinal epithelial cells that respond predominantly to type III IFNs, and it has been shown that type III rather than type I IFNs are important for controlling RV infections in vivo. The RV NSP1 protein antagonizes the host type I IFN response by targeting IRF-3, IRF-5, IRF-7, or ß-TrCP for proteasome-mediated degradation in a strain-specific manner. Here we provide the first demonstration that NSP1 proteins from several human and animal RV strains antagonize type III as well as type I IFN induction. We also show that NSP1 is a potent inhibitor of IRF-1, a previously undescribed property of NSP1 which is conserved among human and animal RVs. Interestingly, all NSP1 proteins were substantially more effective inhibitors of IRF-1 than either IRF-3 or IRF-7 which has significance for evasion of basal anti-viral immunity and type III IFN induction in the intestinal epithelium.

Interferon system deficiencies exacerbating severe pandemic virus infections.

Pandemics are caused by novel pathogens to which pre-existing antibody immunity is lacking. Under these circumstances, the body must rely on innate interferon-mediated defenses to limit pathogen replication and allow development of critical humoral protection. Here, we highlight studies on disease susceptibility during H1N1 influenza and COVID-19 (SARS-CoV-2) pandemics. An emerging concept is that genetic and non-genetic deficiencies in interferon system components lead to uncontrolled virus replication and severe illness in a subset of people. Intriguingly, new findings suggest that individuals with autoantibodies neutralizing the antiviral function of interferon are at increased risk of severe COVID-19. We discuss key questions surrounding how such autoantibodies develop and function, as well as the general implications of diagnosing interferon deficiencies for personalized therapies.

Type I and III IFN-mediated antiviral actions counteracted by SARS-CoV-2 proteins and host inherited factors.

SARS-CoV-2 is a recently identified coronavirus accountable for the current pandemic disease known as COVID-19. Different patterns of disease progression infer a diverse host immune response, with interferon (IFN) being pivotal. IFN-I and III are produced and released by virus-infected cells during the interplay with SARS-CoV-2, thus establishing an antiviral state in target cells. However, the efficacy of IFN and its role in the possible outcomes of the disease are not yet defined, as it is influenced both by factors inherent to the virus and to the host. The virus exhibits multiple strategies to counteract the innate immune response, including those shared by SARS-CoV and MERS-CoV and other novel ones. Inborn errors in the host may affect IFN-related effector proteins or decrease its levels in plasma upon neutralization by preexistent autoantibodies. This battle between the IFN response triggered upon SARS-CoV-2 infection, its magnitude and timing, and the efficacy of its antiviral tools in dispute against the viral evasion strategies together with the genetic factors of the host, generate a scenario whose fate contributes to defining the severity of COVID-19.

Global absence and targeting of protective immune states in severe COVID-19.

Although infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has pleiotropic and systemic effects in some individuals1-3, many others experience milder symptoms. Here, to gain a more comprehensive understanding of the distinction between severe and mild phenotypes in the pathology of coronavirus disease 2019 (COVID-19) and its origins, we performed a whole-blood-preserving single-cell analysis protocol to integrate contributions from all major immune cell types of the blood-including neutrophils, monocytes, platelets, lymphocytes and the contents of the serum. Patients with mild COVID-19 exhibit a coordinated pattern of expression of interferon-stimulated genes (ISGs)3 across every cell population, whereas these ISG-expressing cells are systemically absent in patients with severe disease. Paradoxically, individuals with severe COVID-19 produce very high titres of anti-SARS-CoV-2 antibodies and have a lower viral load compared to individuals with mild disease. Examination of the serum from patients with severe COVID-19 shows that these patients uniquely produce antibodies that functionally block the production of the ISG-expressing cells associated with mild disease, by activating conserved signalling circuits that dampen cellular responses to interferons. Overzealous antibody responses pit the immune system against itself in many patients with COVID-19, and perhaps also in individuals with other viral infections. Our findings reveal potential targets for immunotherapies in patients with severe COVID-19 to re-engage viral defence.

Development and utilization of an infectious clone for porcine deltacoronavirus strain USA/IL/2014/026.

We report the generation of a full-length infectious cDNA clone for porcine deltacoronavirus strain USA/IL/2014/026. Similar to the parental strain, the infectious clone virus (icPDCoV) replicated efficiently in cell culture and caused mild clinical symptoms in piglets. To investigate putative viral interferon (IFN) antagonists, we generated two mutant viruses: a nonstructural protein 15 mutant virus that encodes a catalytically-inactive endoribonuclease (icEnUmut), and an accessory gene NS6-deletion virus in which the NS6 gene was replaced with the mNeonGreen sequence (icDelNS6/nG). By infecting PK1 cells with these recombinant PDCoVs, we found that icDelNS6/nG elicited similar levels of type I IFN responses as icPDCoV, however icEnUmut stimulated robust type I IFN responses, demonstrating that the deltacoronavirus endoribonuclease, but not NS6, functions as an IFN antagonist in PK1 cells. Collectively, the construction of a full-length infectious clone and the identification of an IFN-antagonistic endoribonuclease will aid in the development of live-attenuated deltacoronavirus vaccines.

Loss of orf3b in the circulating SARS-CoV-2 strains.

The newly emerged betacoronavirus, SARS-CoV-2, causes the COVID-19 pandemic since December 2019 with more than 35 million laboratory confirmed human infections and over one million deaths within nine months. The genome of SARS-CoV-2 continues to evolve during the global transmission with the notable emergence of the spike D614G substitution that enhances infectivity. Some of these viral adaptations may alter not only the infectivity but also viral pathogenesis. Continuous phylogenomic analysis of circulating viral strains and functional investigation of new non-synonymous substitutions may help to understand the evolution of virus, its virulence and transmissibility. Here we describe a loss of an accessory protein orf3b (57 amino acids) in current circulating SARS-CoV-2 strains, contributing around 24% of more than 100,000 complete viral genomes analysed. The loss of 3b is caused by the presence of an early stop codon which is created by an orf3a Q57H substitution. There is an increasing trend in the loss of orf3b which has reached 32% in May 2020. Geographically, loss of 3b is more prevalent in certain countries including Colombia (46%), USA (48%), South Korea (51%), France (66%), Saudi Arabia (72%), Finland (76%) and Egypt (77%). Interestingly, the loss of 3b coincides with the emergence of spike D614G substitution. In addition, we found that truncated orf3b has lost the interferon antagonism compared to the full-length orf3b, suggesting a loss of function by the newly adapted virus. Further investigation of orf3b deletion and spike D614G substitution on virulence and infectivity respectively will provide important insights into SARS-CoV-2 evolution.

Porcine deltacoronavirus (PDCoV) infection antagonizes interferon-λ1 production.

Porcine deltacoronavirus (PDCoV) is a novel swine enteropathogenic coronavirus that causes watery diarrhea, vomiting and mortality in nursing piglets. Type III interferons (IFN-λs) are the major antiviral cytokines in intestinal epithelial cells, the target cells in vivo for PDCoV. In this study, we found that PDCoV infection remarkably inhibited Sendai virus-induced IFN-λ1 production by suppressing transcription factors IRF and NF-κB in IPI-2I cells, a line of porcine intestinal mucosal epithelial cells. We also confirmed that PDCoV infection impeded the activation of IFN-λ1 promoter stimulated by RIG-I, MDA5 and MAVS, but not by TBK1 and IRF1. Although the expression levels of IRF1 and MAVS were not changed, PDCoV infection resulted in reduction of the number of peroxisomes, the platform for MAVS to activate IRF1, and subsequent type III IFN production. Taken together, our study demonstrates that PDCoV suppresses type III IFN responses to circumvent the host's antiviral immunity.

SARS-CoV-2 nsp13, nsp14, nsp15 and orf6 function as potent interferon antagonists.

The Coronavirus disease 2019 (COVID-19), which is caused by the novel SARS-CoV-2 virus, is now causing a tremendous global health concern. Since its first appearance in December 2019, the outbreak has already caused over 5.8 million infections worldwide (till 29 May 2020), with more than 0.35 million deaths. Early virus-mediated immune suppression is believed to be one of the unique characteristics of SARS-CoV-2 infection and contributes at least partially to the viral pathogenesis. In this study, we identified the key viral interferon antagonists of SARS-CoV-2 and compared them with two well-characterized SARS-CoV interferon antagonists, PLpro and orf6. Here we demonstrated that the SARS-CoV-2 nsp13, nsp14, nsp15 and orf6, but not the unique orf8, could potently suppress primary interferon production and interferon signalling. Although SARS-CoV PLpro has been well-characterized for its potent interferon-antagonizing, deubiquitinase and protease activities, SARS-CoV-2 PLpro, despite sharing high amino acid sequence similarity with SARS-CoV, loses both interferon-antagonising and deubiquitinase activities. Among the 27 viral proteins, SARS-CoV-2 orf6 demonstrated the strongest suppression on both primary interferon production and interferon signalling. Orf6-deleted SARS-CoV-2 may be considered for the development of intranasal live-but-attenuated vaccine against COVID-19.

Grass carp reovirus VP56 represses interferon production by degrading phosphorylated IRF7.

Grass carp reovirus (GCRV) is an efficient pathogen causing high mortality in grass carp, meanwhile, fish interferon (IFN) is a powerful cytokine enabling host cells to establish an antiviral state; therefore, the strategies used by GCRV to escape the cellular IFN response need to be investigated. Here, we report that GCRV VP56 inhibits host IFN production by degrading the transcription factor IFN regulatory factor 7 (IRF7). First, overexpression of VP56 inhibited the IFN production induced by the polyinosinic-polycytidylic acid (poly I:C) and mitochondrial antiviral signaling protein (MAVS), while the capacity of IRF7 on IFN induction was unaffected. Second, VP56 interacted with RLRs but did not affect the stabilization of the proteins in the normal state, while the phosphorylated IRF7 activated by TBK1 was degraded by VP56 through K48-linked ubiquitination. Finally, overexpression of VP56 remarkably reduced the host cellular ifn transcription and facilitated viral proliferation. Taken together, our results demonstrate that GCRV VP56 suppresses the host IFN response by targeting phosphorylated IRF7 for ubiquitination and degradation.

Nonstructural Protein 11 of Porcine Reproductive and Respiratory Syndrome Virus Induces STAT2 Degradation To Inhibit Interferon Signaling.

Interferons (IFNs) play a crucial role in host antiviral response by activating the JAK/STAT (Janus kinase/signal transducer and activator of transcription) signaling pathway to induce the expression of myriad genes. STAT2 is a key player in the IFN-activated JAK/STAT signaling. Porcine reproductive and respiratory syndrome virus (PRRSV) is an important viral pathogen, causing huge losses to the swine industry. PRRSV infection elicits a meager protective immune response in pigs. The objective of this study was to investigate the effect of PRRSV on STAT2 signaling. Here, we demonstrated that PRRSV downregulated STAT2 to inhibit IFN-activated signaling. PRRSV strains of both PRRSV-1 and PRRSV-2 species reduced the STAT2 protein level, whereas the STAT2 transcript level had minimal change. PRRSV reduced the STAT2 level in a dose-dependent manner and shortened STAT2 half-life significantly from approximately 30 to 5 h. PRRSV-induced STAT2 degradation could be restored by treatment with the proteasome inhibitor MG132 and lactacystin. In addition, PRRSV nonstructural protein 11 (nsp11) was identified to interact with and reduce STAT2. The N-terminal domain (NTD) of nsp11 was responsible for STAT2 degradation and interacted with STAT2 NTD and the coiled-coil domain. Mutagenesis analysis showed that the amino acid residue K59 of nsp11 was indispensable for inducing STAT2 reduction. Mutant PRRSV with the K59A mutation generated by reverse genetics almost lost the ability to reduce STAT2. Together, these results demonstrate that PRRSV nsp11 antagonizes IFN signaling via mediating STAT2 degradation and provide further insights into the PRRSV interference of the innate immunity.IMPORTANCE PRRSV infection elicits a meager protective immune response in pigs. One of the possible reasons is that PRRSV antagonizes interferon induction and its downstream signaling. Interferons are key components in the innate immunity and play crucial roles against viral infection and in the activation of adaptive immune response via JAK/STAT signaling. STAT2 is indispensable in the JAK/STAT signaling since it is also involved in activation of antiviral activity in the absence of STAT1. Here, we discovered that PRRSV nsp11 downregulates STAT2. Interestingly, the N-terminal domain of nsp11 is responsible for inducing STAT2 degradation and directly interacts with STAT2 N-terminal domain. We also identified a crucial amino acid residue K59 in nsp11 since a mutation of it led to loss of the ability to downregulate STAT2. A mutant PRRSV with mutation of K59 had minimal effect on STAT2 reduction. Our data provide further insights into PRRSV interference with interferon signaling.

The Nucleoprotein and Phosphoprotein of Peste des Petits Ruminants Virus Inhibit Interferons Signaling by Blocking the JAK-STAT Pathway.

Peste des petits ruminants virus (PPRV) is associated with global peste des petits ruminants resulting in severe economic loss. Peste des petits ruminants virus dampens host interferon-based signaling pathways through multiple mechanisms. Previous studies deciphered the role of V and C in abrogating IFN-ß production. Moreover, V protein directly interacted with signal transducers and activators of transcription 1 (STAT1) and STAT2 resulting in the impairment of host IFN responses. In our present study, PPRV infection inhibited both IFN-ß- and IFN-γ-induced activation of IFN-stimulated response element (ISRE) and IFN-γ-activated site (GAS) element, respectively. Both N and P proteins, functioning as novel IFN response antagonists, markedly suppressed IFN-ß-induced ISRE and IFN-γ-induced GAS promoter activation to impair downstream upregulation of various interferon-stimulated genes (ISGs) and prevent STAT1 nuclear translocation. Specifically, P protein interacted with STAT1 and subsequently inhibited STAT1 phosphorylation, whereas N protein neither interacted with STAT1 nor inhibited STAT1 phosphorylation as well as dimerization, suggesting that the N and P protein antagonistic effects were different. Though they differed in their relationship to STAT1, both proteins blocked JAK-STAT signaling, severely negating the host antiviral immune response. Our study revealed a new mechanism employed by PPRV to evade host innate immune response, providing a platform to study the interaction of paramyxoviruses and host response.