Interferon-stimulated Response Element Research Articles

Marek’s disease virus inhibits the JAK-STAT signaling pathway to evade the innate immune response1

The Janus kinase (JAK)–signal transducer and activator of transcription (STAT) signaling pathway plays a crucial role in innate immunity by inducing antiviral proteins in response to interferon signals. Marek’s disease virus (MDV), a member of the alpha-herpes virus family, exerts potent tumorigenic and immunosuppressive effects. Recent studies have primarily focused on the tumorigenic mechanisms of MDV, and the mechanism of immune evasion has not been fully understood. In this study, we showed that MDV reduced the production of interferon-stimulated gene (ISGs) by inhibiting the phosphorylation and nuclear translocation of STAT1. Using a dual-luciferase reporter system, we screened for viral proteins that significantly suppress interferon-stimulated response element (ISRE) promoter activity. Meq overexpression markedly reduced ISRE promoter activity and ISG expression, whereas infection with Meq-deficient MDV induced higher ISG production in vitro and in vivo than infection with wild-type MDV. Meq also inhibited the phosphorylation and nuclear translocation of STAT1. Further experiments showed that Meq interacted with JAK1 and tyrosine kinase 2 (TYK2) and thereby inhibited JAK1–STAT1 interactions. Meq degraded TYK2 via a caspase-mediated pathway. The Meq-deficient MDV mutant replicated less efficiently than the wild-type MDV, both in vitro and in vivo. Collectively, these findings demonstrate that Meq played an immunosuppressive role in MDV by attenuating the JAK–STAT signaling pathway, which facilitated escape from innate immune-surveillance mechanisms.

Nuclear PCGF3 inhibits the antiviral immune response by suppressing the interferon-stimulated gene

Type I interferon (IFN-I) plays a crucial role in the antiviral immune response and inflammatory autoimmune diseases by inducing the expression of IFN-stimulated genes (ISGs). Hence, the regulation of ISG expression is fundamental for maintaining immune homeostasis. In this study, we found that PCGF3 negatively regulates the antiviral response by suppressing the expression of ISGs. The deficiency of PCGF3 in innate immune cells results in an augmented expression of ISGs in response to IFN-I stimulation. Mechanistically, PCGF3 is recruited to interferon-stimulated response elements (ISREs) region in an IFN-dependent way, precluding STAT1 from binding to the ISG promoter and diminishing ISRE activity. Additionally, we observed a negative correlation between decreased PCGF3 expression and elevated ISG expression in peripheral blood mononuclear cells (PBMCs) of patients with dermatomyositis (DM). Our findings clarified the epigenetic regulatory role of PCGF3 in inhibiting the excessive expression of ISGs induced by IFN-I under pathological circumstances.

Zinc-finger CCHC-type containing protein 8 promotes RNA virus replication by suppressing the type-I interferon responses.

Host cells have evolved an intricate regulatory network to fine tune the type-I interferon responses. However, the full picture of this regulatory network remains to be depicted. In this study, we found that knock out of zinc-finger CCHC-type containing protein 8 (ZCCHC8) impairs the replication of influenza A virus (IAV), Sendai virus (Sev), Japanese encephalitis virus (JEV), and vesicular stomatitis virus (VSV). Further investigation unveiled that ZCCHC8 suppresses the type-I interferon responses by targeting the interferon regulatory factor 3 (IRF3) signaling pathway. Mechanistically, ZCCHC8 associates with phosphorylated IRF3 and disrupts the interaction of IRF3 with the co-activator CREB-binding protein (CBP). Additionally, the direct binding of ZCCHC8 with the IFN-stimulated response element (ISRE) impairs the ISRE-binding of IRF3. Our study contributes to the comprehensive understanding for the negative regulatory network of the type-I interferon responses and provides valuable insights for the control of multiple viruses from a host-centric perspective.IMPORTANCEThe innate immune responses serve as the initial line of defense against invading pathogens and harmful substances. Negative regulation of the innate immune responses plays an essential role in avoiding auto-immune diseases and over-activated immune responses. Hence, the comprehensive understanding of the negative regulation network for innate immune responses could provide novel therapeutic insights for the control of viral infections and immune dysfunction. In this study, we report that ZCCHC8 negatively regulates the type-I interferon responses. We illustrate that ZCCHC8 impedes the IRF3-CBP association by interacting with phosphorylated IRF3 and competes with IRF3 for binding to ISRE. Our study demonstrates the role of ZCCHC8 in the replication of multiple RNA viruses and contributes to a deeper understanding of the negative regulation system for the type-I interferon responses.

Luciferase Reporter Assay for Determining the Signaling Activity of Interferons.

The classic dual luciferase reporter assay has been widely used to rapidly and accurately determine the transcriptional activity of a given promoter induced by certain signal pathways in the cells. In particular, the sensitive characteristics of luciferase highlight its significance in many experiments, such as weak promoter analysis, transfection studies using small amounts of DNA, and detection in cell lines with low transfection efficiency. This chapter presents detailed information and experimental procedures for measuring interferon (IFN)-induced Interferon-Stimulated Response Element (ISRE) promoter activity using the dual luciferase reporter assay.

Development of a highly sensitive platform for protein–protein interaction detection and regulation of T cell function

We developed a highly sensitive assay for detecting protein-protein interaction using chimeric receptors comprising two molecules of interest in the extracellular domain and interferon alpha and beta receptor subunit 1 or 2 (IFNAR1/2) in the intracellular domain. This intracellular IFNAR1/2 reconstitution system (IFNARRS) proved markedly more sensitive than the NanoBiT system, currently considered one of the best detection systems for protein interaction. Employing chimeric receptors with extracellular domains from the IFNγ or IL-2 receptor and the intracellular domains of IFNAR1/2, the IFNARRS system effectively identifies low IFNγ or IL-2 levels. Cells stably expressing these chimeric receptors responded to IFNγ secreted by activated T cells following various stimuli, including a specific peptide-antigen. The activation signals were further enhanced by the expression of relevant genes, such as costimulators, via IFN-stimulated response elements in the promoters. Besides IFNγ or IL-2, the IFNARRS system demonstrated the capability to detect other cytokines by using the corresponding extracellular domains from these target cytokine receptors.

Evolutionarily conserved Otub1 suppresses antiviral immune response by promoting Irf3 proteasomal degradation in miiuy croaker, Miichthys miiuy

Increasing evidence has been shown that OTUB1, a member of OTU deubiquitinases, is of importance in regulating the immune system. However, its molecular identification and functional characterization in teleosts are still rarely known. In this work, we cloned the otub1 of miiuy croaker (Miichthys miiuy), analyzed its sequence, structure, and evolution at genetic and protein levels, and determined its function in the antiviral immune response. The complete open reading frame (ORF) of miiuy croaker otub1 is 843 bp in length, encoding 280 amino acids. Miiuy croaker Otub1 has an OTU domain at the carboxyl terminus, which is a common functional domain that exists in OTU deubiquitinases. Molecular characteristics and evolution analysis results indicated that miiuy croaker Otub1, especially its functional domain, is highly conserved during evolution. The luciferase reporter assays showed that miiuy croaker Otub1 could significantly inhibit the poly(I:C) and Irf3-induced IFN1 and IFN-stimulated response element (ISRE) activation. Further experiments showed that miiuy croaker Otub1 decreases Irf3 protein abundance by promoting its proteasomal degradation. These data suggest that the evolutionarily conserved Otub1 acts as a suppressor in controlling antiviral immune response by promoting Irf3 proteasomal degradation in miiuy croaker.

Grouper OTUB1 and OTUB2 promote red-spotted grouper nervous necrosis virus (RGNNV) replication by inhibiting the host innate immune response

Red-spotted grouper nervous necrosis virus (RGNNV) is a major viral pathogen of grouper and is able to antagonize interferon responses through multiple strategies, particularly evading host immune responses by inhibiting interferon responses. Ovarian tumor (OTU) family proteins are an important class of DUBs and the underlying mechanisms used to inhibit interferon pathway activation are unknown. In the present study, primers were designed based on the transcriptome data, and the ovarian tumor (OTU) domain-containing ubiquitin aldehyde-binding protein 1 (OTUB1) and OTUB2 genes of Epinephelus coioides (EcOTUB1 and EcOTUB2) were cloned and characterized. The homology alignment showed that both EcOTUB1 and EcOTUB2 were most closely related to E. lanceolatus with 98 % identity. Both EcOTUB1 and EcOTUB2 were distributed to varying degrees in grouper tissues, and the transcript levels were significantly up-regulated following RGNNV stimulation. Both EcOTUB1 and EcOTUB2 promoted replication of RGNNV in vitro, and inhibited the promoter activities of interferon stimulated response element (ISRE), nuclear transcription factors kappaB (NF-κB) and IFN3, and the expression levels of interferon related genes and proinflammatory factors. Co-immunoprecipitation experiments showed that both EcOTUB1 and EcOTUB2 could interact with TRAF3 and TRAF6, indicating that EcOTUB1 and EcOTUB2 may play important roles in interferon signaling pathway. The results will provide a theoretical reference for the development of novel disease prevention and control techniques.

Key role of interferon regulatory factor 1 (IRF-1) in regulating liver disease: progress and outlook.

Interferon regulatory factor 1 (IRF-1) is a member of the IRF family. It is the first transcription factor to be identified that could bind to the interferon-stimulated response element (ISRE) on the target gene and displays crucial roles in the interferon-induced signals and pathways. IRF-1, as an important medium, has all of the advantages of full cell cycle regulation, cell death signaling transduction, and reinforcing immune surveillance, which are well documented. Current studies indicate that IRF-1 is of vital importance to the occurrence and evolution of multifarious liver diseases, including but not limited to inhibiting the replication of the hepatitis virus (A/B/C/E), alleviating the progression of liver fibrosis, and aggravating hepatic ischemia-reperfusion injury (HIRI). The tumor suppression of IRF-1 is related to the clinical characteristics of liver cancer patients, which makes it a potential indicator for predicting the prognosis and recurrence of liver cancer; additionally, the latest studies have revealed other effects of IRF-1 such as protection against alcoholic/non-alcoholic fatty liver disease (AFLD/NAFLD), cholangiocarcinoma suppression, and uncommon traits in other liver diseases that had previously received little attention. Intriguingly, several compounds and drugs have featured a protective function in specific liver disease models in which there is significant involvement of the IRF-1 signal. In this paper, we hope to propose a prospective research basis upon which to help decipher translational medicine applications of IRF-1 in liver disease treatment.

Systematic Review: JAK-STAT Regulation and Its Impact on Inflammation Response in ARDS from COVID-19

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has had a global impact and resulted in millions of deaths worldwide. The course of the Janus kinase signaling transducers and activators (JAK-STAT) pathway is an important molecular pathway that is involved in the cellular response to various cytokines and growth factors promoting an inflammatory response. The overactivation of the JAK-STAT signaling pathway in coronavirus disease 2019 (COVID-19) and its effect on acute respiratory distress syndrome (ARDS)-induced inflammatory processes was observed in various clinical articles that focused on JAK-STAT regulation regarding angiotensin converting enzyme 2 (ACE2) expression and cytokine storm release. Down-regulation of the JAK-STAT signaling pathway through inhibitors decreases the inflammatory response by decreasing cytokine storm release. However, the increased regulation of JAK-STAT in severe COVID-19 patients caused cytokines such as interferon alpha (IFN-α) to promote the phosphorylation of STATs. This response indicated an imbalance with JAK-STAT regulation and its inability to induce the transcription of interferon stimulated response elements. Furthermore, an increase in ACE2 regulation was noted to also increase JAK-STAT signaling, yet the down-regulation of JAK-STAT signaling can result in the overexpression of ACE2 by binding to SARS-CoV-2 and increasing STAT1 expression. Data suggest that inflammatory cytokines enhance the activation of ACE2 in endothelial cells via JAK-STAT pathway. Increasing the regulation of the JAK-STAT signaling pathway enhances the release of cytokines such as tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ), further expressing ACE2. The expression of ACE2 regulates STAT1 and STAT2 expression, leading to the up-regulation of the inflammasomal complexes in hyper-inflammatory responses from the JAK-STAT pathway. Through the review of various clinical reports, the effect of the JAK-STAT signaling pathway on ARDS-induced inflammatory response was observed and correlated with the expression of ACE2 and cytokine storm release in severe COVID-19 cases.

Rainbow trout DUBA inhibits type I interferon signaling by deubiquitinating TRAF3

Deubiquitinating enzyme A (DUBA), a member of the ovarian tumor (OTU) subfamily of deubiquitinases (DUBs), is recognized for its negative regulatory role in type I interferon (IFN) expression downstream of Toll-like receptor 3 (TLR3). However, its involvement in the TLR3 signaling pathway in fish remains largely unexplored. In this study, we investigated the regulatory role of DUBA (OmDUBA) in the TLR3 response in rainbow trout (Oncorhynchus mykiss). OmDUBA features a conserved OTU domain, and its expression increased in RTH-149 cells following stimulation with the TLR3 agonist poly(I:C). Gain- and loss-of-function experiments demonstrated that OmDUBA attenuated the activation of TANK-binding kinase 1 (TBK1), resulting in a subsequent reduction in type I IFN expression and IFN-stimulated response element (ISRE) activation in poly(I:C)-stimulated cells. OmDUBA interacted with TRAF3, a crucial mediator in TLR3-mediated type I IFN production. Under poly(I:C) stimulation, there was an augmentation in the K63-linked polyubiquitination of TRAF3, a process significantly inhibited upon OmDUBA overexpression. These findings suggest that OmDUBA may function similarly to its mammalian counterparts in downregulating the poly(I:C)-induced type I IFN response in rainbow trout by removing the K63-linked ubiquitin chain on TRAF3. Our study provides novel insights into the role of fish DUBA in antiviral immunity.

Foot-and-Mouth Disease Virus Capsid Protein VP1 Antagonizes Type I Interferon Signaling via Degradation of Histone Deacetylase 5.

Foot-and-mouth disease (FMD) is a highly contagious and economically important disease of cloven-hoofed animals that hampers trade and production. To ensure effective infection, the foot-and-mouth disease virus (FMDV) evades host antiviral pathways in different ways. Although the effect of histone deacetylase 5 (HDAC5) on the innate immune response has previously been documented, the precise molecular mechanism underlying HDAC5-mediated FMDV infection is not yet clearly understood. In this study, we found that silencing or knockout of HDAC5 promoted FMDV replication, whereas HDAC5 overexpression significantly inhibited FMDV propagation. IFN-β and IFN-stimulated response element (ISRE) activity was strongly activated through the overexpression of HDAC5. The silencing and knockout of HDAC5 led to an increase in viral replication, which was evident by decreased IFN-β, ISG15, and ISG56 production, as well as a noticeable reduction in IRF3 phosphorylation. Moreover, the results showed that the FMDV capsid protein VP1 targets HDAC5 and facilitates its degradation via the proteasomal pathway. In conclusion, this study highlights that HDAC5 acts as a positive modulator of IFN-β production during viral infection, while FMDV capsid protein VP1 antagonizes the HDAC5-mediated antiviral immune response by degrading HDAC5 to facilitate viral replication.

UMP-CMP kinase 2 inhibits ZIKV replication through activation of type I IFN signaling pathway.

Cytidine/uridine monophosphate kinase 2 (UMP-CMP kinase 2, CMPK2) has been reported as an antiviral interferon-stimulated gene (ISG). We previously observed that the expression of CMPK2 was significantly upregulated after Zika Virus (ZIKV) infection in A549 cells. However, the association and the underlying mechanisms between CMPK2 induction and ZIKV replication remain to be determined. We investigated the induction of CMPK2 during ZIKV infection and the effect of CMPK2 on ZIKV replication in A549, U251, Vero, IFNAR-deficient U5A and its parental 2fTGH cells, Huh7 and its RIG-I-deficient derivatives Huh7.5.1 cells. The activation status of Jak-STAT signaling pathway was determined by detecting the phosphorylation level of STAT1, the activity of interferon stimulated response element (ISRE) and the expression of several interferon stimulated genes (ISGs). We found that ZIKV infection induced CMPK2 expression through an IFNAR and RIG-I dependent manner. Overexpression of CMPK2 inhibited while CMPK2 knockdown promoted ZIKV replication in A549 and U251 cells. Mechanically, we found that CMPK2 overexpression increased IFNβ expression and activated Jak/STAT signaling pathway as shown by the increased level of p-STAT1, enhanced activity of ISRE, and the upregulated expression of downstream ISGs. These findings suggest that ZIKV infection induced CMPK2 expression, which inhibited ZIKV replication and serves as a positive feedback regulator for IFN-Jak/STAT pathway.

Deacetylase Sirtuin 1 mitigates type I IFN- and type II IFN-induced signaling and antiviral immunity.

Type I and type II IFNs are important immune modulators in both innate and adaptive immunity. They transmit signaling by activating JAK-STAT pathways. Sirtuin 1 (SIRT1), a class III NAD+-dependent deacetylase, has multiple functions in a variety of physiological processes. Here, we characterized the novel functions of SIRT1 in the regulation of type I and type II IFN-induced signaling. Overexpression of SIRT1 inhibited type I and type II IFN-induced interferon-stimulated response element activation. In contrast, knockout of SIRT1 promoted type I and type II IFN-induced expression of ISGs and inhibited viral replication. Treatment with SIRT1 inhibitor EX527 had similar positive effects. SIRT1 physically associated with STAT1 or STAT3, and this interaction was enhanced by IFN stimulation or viral infection. By deacetylating STAT1 at K673 and STAT3 at K679/K685/K707/K709, SIRT1 downregulated the phosphorylation of STAT1 (Y701) and STAT3 (Y705). Sirt1+/- primary peritoneal macrophages and Sirt1+/- mice exhibited enhanced IFN-induced signaling and antiviral activity. Thus, SIRT1 is a novel negative regulator of type I and type II IFN-induced signaling through its deacetylase activity.IMPORTANCESIRT1 has been reported in the precise regulation of antiviral (RNA and DNA) immunity. However, its functions in type I and type II IFN-induced signaling are still unclear. In this study, we deciphered the important functions of SIRT1 in both type I and type II IFN-induced JAK-STAT signaling and explored the potential acting mechanisms. It is helpful for understanding the regulatory roles of SIRT1 at different levels of IFN signaling. It also consolidates the notion that SIRT1 is an important target for intervention in viral infection, inflammatory diseases, or even interferon-related therapies.

The mechanism of Langchuangding in treatment of systemic lupus erythematosus via modulating TLR7-IRF7-IFNα pathway

ObjectSystemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by aberrant activity of the immune system. Plasmacytoid dendritic cells (pDCs) which the main producer of activated type I interferon, are related to SLE disease activity. To investigate the mechanism of Langchuangding (LCD) improving SLE based on TLR7-IRF7-IFNα pathway. MethodsSLE patients were randomly divided into Chinese medicine combined with western medicine (CWM) group and western medicine (WM) group, to observe the effect of LCD. The percent of pDCs in peripheral blood of SLE patients were detected by flow cytometry, and the influence of LCD on gene expression in SLE patients were detected by gene microarray. Mouse bone marrow cells were differentiated into dendritic like cells (DLC), then divided into Blank, immune complex (IC), LCD and dexamethasone (DXM) group. Employed RT-qPCR to detect MyD88, and IRF7 mRNA, and western blotting to determinate TLR7, MyD88, and p-IRF7 proteins. The IFNα in SLE patients were detected by enzyme-linked immunosorbent assay (ELISA). Employ dual luciferase to observe the interferon stimulated response element (ISRE) gene. ResultspDCs in WM group was higher than that of CWM group. The plasma IFNα in CWM group was significantly lower than that in WM group. The gene microarray showed that the gene expression of IFNα related signaling pathway in peripheral blood mononuclear cell (PBMC) and genes related to activation and proliferation of immune cells were down-regulated after LCD treatment. The DLCs MyD88, and IRF7 mRNA were down-regulated, TLR7, MyD88, and p-IRF7 proteins were significantly reduced, and the supernatant IFNα was significantly decreased in LCD group. LCD were mildly inhibited activation of ISRE in 293T cells. ConclusionsIn certain degree, LCD is beneficial to SLE patients. LCD therapy SLE may be through TLR7 signaling pathway, and IRF7 may be a promising therapeutic target for the treatment of SLE.

Identification and Characterization of the Stimulator of Interferon Genes (STING) in Chinese Giant Salamander Andrias davidianus

The stimulator of interferon genes (STING) is a central and multifaceted mediator in innate immunity, and plays vital roles in defending against pathogen infection. In this study, we identified and functionally characterized the STING homolog from the Chinese giant salamander Andrias davidianus (AdSTING) for the first time. The open reading frame of AdSTING encodes a 362 amino acid protein with a predicted molecular mass of 41.6 kDa, which shares 31.1–46.7% of its sequence identity with STING homologs in other vertebrates. Structural analysis revealed that AdSTING possesses four predicted transmembrane domains (TMs) at the N-terminal, and a C-terminal domain (CTD) featuring a dimerization domain (DD), a c-di-GMP-binding domain (CBD), and a short C-terminal tail (CTT). Tissue distribution analysis showed that AdSTING mRNA was ubiquitously expressed in all examined tissues, with abundant expression in muscles, intestine, and thymus. During Andrias davidianus ranavirus (ADRV) infection, significant up-regulation of AdSTING expression was observed in the thymus, spleen, and kidney. Upon different stimuli in vitro, the expression of AdSTING was significantly induced by ADRV infection or polyinosin-polycytidylic acid (poly I:C) stimulation, but no obvious changes were observed during lipopolysaccharide (LPS) stimulation. Subcellular localization analysis revealed that AdSTING mainly localized in the cytoplasm in the Chinese giant salamander thymus cell line (GSTC) and co-localized with the endoplasmic reticulum (ER). Luciferase reporter assays confirmed the ability of AdSTING to activate the interferon-stimulated response element (ISRE) and interferon (IFN) promoter. Furthermore, overexpression of AdSTING effectively decreased ADRV infection, as evidenced by the reduction of virus titers and viral gene expression. Collectively, our findings underscore the pivotal role of AdSTING in the antiviral innate immunity of the Chinese giant salamander, offering insights into the functional evolution of STING in amphibians.

A molluscan IRF interacts with IKKα/β family protein and modulates NF-κB and MAPK activity

Interferon regulatory factor (IRF) family proteins are key transcription factors involved in vital physiological processes such as immune defense. However, the function of IRF in invertebrates, especially in marine shellfish is not clear. In this study, a new IRF gene (CfIRF2) was identified in the Zhikong scallop, Chlamys farreri, and its immune function was analyzed. CfIRF2 has an open reading frame of 1107 bp encoding 368 amino acids. The N-terminus of CfIRF2 consists of a typical IRF domain, with conserved amino acid sequences. Phylogenetic analysis suggested close evolutionary relationship with shellfish IRF1 subfamily proteins. Expression pattern analysis showed that CfIRF2 mRNA was expressed in all tissues, with the highest expression in the hepatopancreas and gills. CfIRF2 gene expression was substantially enhanced by a pathogenic virus (such as acute viral necrosis virus) and poly(I:C) challenge. Co-immunoprecipitation assay identified CfIRF2 interaction with the IKKα/β family protein CfIKK1 of C. farreri, demonstrating a unique signal transduction mechanism in marine mollusks. Moreover, CfIRF2 interacted with itself to form homologous dimers. Overexpression of CfIRF2 in HEK293T cells activated reporter genes containing interferon stimulated response elements and NF-κB genes in a dose-dependent manner and promoted the phosphorylation of protein kinases (JNK, Erk1/2, and P38). Our results provide insights into the functions of IRF in mollusks innate immunity and also provide valuable information for enriching comparative immunological theory for the prevention of diseases in scallop farming.

Antiviral role of grouper FoxO1 against RGNNV and SGIV infection

As a key regulator of the innate immune system, FoxO1 has a variety of activities in biological organisms. In the present study, grouper FoxO1 (EcFoxO1) was cloned and the antiviral activity in red grouper neuron necrosis virus (RGNNV) and Singapore grouper iridescent virus (SGIV) was examined. The open reading frame (ORF) of EcFoxO1 contains 2,034 base pairs that encode a protein of 677 amino acids with a predicted molecular weight of 73.21 kDa. EcFoxO1 was shown to be broadly distributed in healthy grouper tissues, and was up-regulated in vitro in response to stimulation by RGNNV and SGIV. EcFoxO1 has a whole-cell distribution in grouper spleen (GS) cells. EcFoxO1 decreased the replication of RGNNV and SGIV, and activated interferon (IFN) 3, IFN-stimulated response element (ISRE), and nuclear factor-κB (NF-κB) promoter activities. EcFoxO1 could interact with EcIRF3. Together, the results demonstrated that EcFoxO1 might be an important regulator of grouper innate immune response against RGNNV and SGIV infection.

Seneca Valley virus 3Cpro antagonizes type I interferon response by targeting STAT1-STAT2-IRF9 and KPNA1 signals.

Type I interferon (IFN) signaling plays a principal role in host innate immune responses against invading viruses. Viruses have evolved diverse mechanisms that target the Janus kinase-signal transducer and activator of transcription (STAT) signaling pathway to modulate IFN response negatively. Seneca Valley virus (SVV), an emerging porcine picornavirus, has received great interest recently because it poses a great threat to the global pork industry. However, the molecular mechanism by which SVV evades host innate immunity remains incompletely clear. Our results revealed that SVV proteinase (3Cpro) antagonizes IFN signaling by degrading STAT1, STAT2, and IRF9, and cleaving STAT2 to escape host immunity. SVV 3Cpro also degrades karyopherin 1 to block IFN-stimulated gene factor 3 nuclear translocation. Our results reveal a novel molecular mechanism by which SVV 3Cpro antagonizes the type I IFN response pathway by targeting STAT1-STAT2-IRF9 and karyopherin α1 signals, which has important implications for our understanding of SVV-evaded host innate immune responses.

African Swine Fever Virus pF778R Attenuates Type I Interferon Response by Impeding STAT1 Nuclear Translocation

African swine fever virus (ASFV) is an extensive and intricate double-stranded DNA virus with approximately 100% lethality in domestic swine. There is no effective vaccine to combat this virus, and this has led to substantial economic losses in the swine industry. ASFV encodes various proteins that impede interferon-based immune defenses in the host by employing diverse mechanisms. However, the roles of most of these proteins remain unknown. Therefore, understanding the immune evasion mechanisms employed by ASFV may facilitate the development of effective measures against the virus. In this study, we discovered a negative regulation of the type I interferon (IFN) response by the ASFV ribonuclease reductase large subunit pF778R. This novel type Ⅰ IFN response antagonist significantly inhibits IFN-α-induced interferon-stimulated response element promoter activation, precludes the upregulation of various interferon-stimulated genes, and prevents STAT1 nuclear translocation. Mechanistically, pF778R did not affect the protein levels of crucial molecules in the JAK/STAT signaling pathway or engage in direct interactions. However, pF778R expression impedes type I IFN responses mediated by the JAK/STAT signaling pathway. Further investigations revealed that pF778R did not interfere with STAT1 phosphorylation or dimerization, but it inhibited IFN signaling by weakening the nuclear accumulation of activated STAT1. The critical role of the ASFV protein pF778R in evading IFN-I-mediated innate immunity highlights a unique mode of ASFV evasion and provides insights into the pathogenic mechanism of the virus.

ATG4B antagonizes antiviral immunity by GABARAP-directed autophagic degradation of TBK1

ABSTRACT TBK1 (TANK binding kinase 1) is an essential kinase of antiviral immunity, yet the regulatory mechanisms responsible for its stringent control via autophagy are not fully understood. Here, we identify the macroautophagy/autophagy-related cysteine protease ATG4B as a negative regulator of human antiviral immune responses by targeting TBK1 for autophagic degradation at the advanced stage of viral infection. Mechanistically, ATG4B serves as an adaptor for recruiting TBK1 to GABARAP (GABA type A receptor-associated protein), which subsequently leads to the TBK1-GABARAP interaction through the LC3-interacting region (LIR) motif of TBK1 ULD domain. Moreover, pharmacological ATG4B inhibitor, a small molecule named S130, contributes to host defense against viral infection and blocks ATG4B-dependent autophagic degradation of TBK1. Accordingly, S130 increases antiviral response and inhibits the VSV infection both in vitro and in vivo. Altogether, our study reveals the regulatory role of ATG4B in modulating TBK1-centered type I interferon (IFN) signaling, and indicates that ATG4B suppression can provide a potential therapy target for viral infection. Abbreviations Baf A1: bafilomycin A1; GABARAP: GABA type A receptor-associated protein; GFP: green fluorescent protein; IFN: interferon; IKBKE/IKKi: inhibitor of nuclear factor kappa B kinase subunit epsilon; IRF3: interferon regulatory factor 3; ISG: interferon-stimulated gene; ISRE: IFN-stimulated response element; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAVS: mitochondrial antiviral signaling protein; MOI: multiplicity of infection; PAMPs: pathogen-associated molecule patterns; RIGI/DDX58: RNA sensor RIG-I; SeV: Sendai virus; siRNA: small interfering RNA; TBK1: TANK binding kinase 1; WT: wild-type; VSV: vesicular stomatitis virus.