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During virus-host co-evolution, viruses have developed multiple strategies to dampen IFN response and prevent its antiviral activity in host cells. To date, the interactions between host IFN response and the immune evasion strategies exploited by fish iridoviruses still remain largely uncertain. Here, a potential immune evasion protein candidate of Singapore grouper iridovirus (SGIV), VP82 (encoded by SGIV ORF82) was screened and its roles during viral replication were investigated in detail. Firstly, VP82 overexpression dramatically decreased IFN or ISRE promoter activity and the transcription levels of IFN stimulated genes (ISGs) stimulated by grouper cyclic GMP-AMP synthase (EccGAS)/stimulator of interferon genes (EcSTING), TANK-binding kinase 1 (EcTBK1), IFN regulatory factor 3 (EcIRF3)and EcIRF7. Secondly, Co-IP assays indicated that VP82 interacted with EcIRF3 and EcIRF7, but not EcSTING and EcTBK1, which was consistent with the co-localization between VP82 and EcIRF3 or EcIRF7. Furthermore, VP82 promoted the degradation of EcIRF3 and EcIRF7 in a dose-dependent manner via the autophagy pathway. Finally, VP82 overexpression accelerated SGIV replication, evidenced by the increased transcriptions of viral core genes and viral production. Moreover, the antiviral action of EcIRF3 or EcIRF7 was significantly depressed in VP82 overexpressed cells. Together, VP82 was speculated to exert crucial roles for SGIV replication by inhibiting the IFN response via the degradation of IRF3 and IRF7. Our findings provided new insights into understanding the immune evasion strategies utilized by fish iridovirus through IFN regulation., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The impact of a deficiency in interferon regulatory factor (IRF)3 and IRF7 was evaluated in an herpes simplex virus encephalitis (HSE) model. Compared to wild type (WT), the mortality rates of infected IRF3 -/- and IRF7 -/- mice were higher and associated with increased brain viral titers. At a critical time post-infection, IRF7 -/- mice exhibited a deficit in IFN-β production. At a later time point, levels of type I IFNs and cytokines were increased in brains of both deficient mice compared to WT. Our results suggest that IRF3, and especially IRF7, are important for an effective control of inflammatory responses during HSE.

Type I interferon (IFN-I) responses are critical for the control of RNA virus infections, however, many viruses, including Dengue (DENV) and Chikungunya (CHIKV) virus, do not directly activate plasmacytoid dendritic cells (pDCs), robust IFN-I producing cells. Herein, we demonstrated that DENV and CHIKV infected cells are sensed by pDCs, indirectly, resulting in selective IRF7 activation and IFN-I production, in the absence of other inflammatory cytokine responses. To elucidate pDC immunomodulatory functions, we developed a mouse model in which IRF7 signaling is restricted to pDC. Despite undetectable levels of IFN-I protein, pDC-restricted IRF7 signaling controlled both viruses and was sufficient to protect mice from lethal CHIKV infection. Early pDC IRF7-signaling resulted in amplification of downstream antiviral responses, including an accelerated natural killer (NK) cell-mediated type II IFN response. These studies revealed the dominant, yet indirect role of pDC IRF7-signaling in directing both type I and II IFN responses during arbovirus infections., Competing Interests: BW, SW, BZ, ST, SC, ED, HP, IB, PJ, DD, TW, MA, MD No competing interests declared, (© 2018, Webster et al.)

In mammals, IFN regulatory factor (IRF)1, IRF3, and IRF7 are three critical transcription factors that are pivotal for cooperative regulation of the type I IFN response. In this study, we explored the relative contribution of zebrafish (Danio rerio) IRF1 (DrIRF1), IRF3 (DrIRF3), and IRF7 (DrIRF7) (DrIRF1/3/7) to zebrafish IFNΦ1 (DrIFNΦ1) and IFNΦ3 (DrIFNΦ3) (DrIFNΦ1/3) activation. Following spring viremia of carp virus infection, DrIFNΦ1/3 and DrIRF1/3/7 transcripts are significantly induced in zebrafish tissues, which correlates with the replication of spring viremia of carp virus. DrIRF1/3/7 selectively bind to the IRF-binding element/IFN-stimulated regulatory element sites of DrIFNΦ1/3 promoters, with the exception that DrIRF3 has no preference for two IRF-binding element/IFN-stimulated regulatory element motifs within the DrIFNΦ3 promoter. Consistently, DrIRF3 alone activates DrIFNΦ1, but not DrIFNΦ3; DrIRF7 predominantly stimulates DrIFNΦ3; and DrIRF1 has similar potential to DrIFNΦ1 and DrIFNΦ3. Strikingly, DrIRF3 facilitates the binding of DrIRF1 and DrIRF7 to both zebrafish IFN promoters, and so does DrIRF7 for the binding of DrIRF1, particularly to the DrIFNΦ3 promoter. These binding properties correlate with differential responses of DrIFNΦ1 and DrIFNΦ3 to the combinatory stimulation of DrIRF1/3/7, depending on their relative amounts. Similar to the dual roles of human IRF3 in regulating IRF7-activated IFNα genes, DrIRF3 exerts dual effects on DrIRF1-mediated DrIFNΦ3 gene expression: an inhibitory effect at lower concentrations and a synergistic effect at higher concentrations. These data provide evidence that fish and mammals have evolved a similar IRF-dependent regulatory mechanism fine-tuning IFN gene activation., (Copyright © 2016 by The American Association of Immunologists, Inc.)

Human metapneumovirus (hMPV) is a leading cause of respiratory tract disease in children and is associated with acute bronchiolitis, pneumonia, and asthma exacerbations, yet the mechanisms by which the host immune response to hMPV is regulated are poorly understood. By using gene-deleted neonatal mice, we examined the contributions of the innate receptor signaling molecules interferon (IFN)-β promoter stimulator 1 (IPS-1), IFN regulatory factor (IRF) 3, and IRF7. Viral load in the lungs was markedly greater in IPS-1(-/-) > IRF3/7(-/-) > IRF3(-/-), but not IRF7(-/-), mice compared with wild-type mice. IFN-β and IFN-λ2/3 (IL-28A/B) production was attenuated in the bronchoalveolar lavage fluid in all factor-deficient mice compared with wild-type mice at 1 day after infection, although IFN-λ2/3 was greater in IRF3/7(-/-) mice at 5 days after infection. IRF7(-/-) and IRF3/7(-/-) mice presented with airway eosinophilia, whereas only IRF3/7(-/-) mice developed an exaggerated type 1 and 17 helper T-cell response, characterized by natural killer T-cell and neutrophilic inflammation. Despite having the highest viral load, IPS-1(-/-) mice did not develop a proinflammatory cytokine or granulocytic response to hMPV infection. Our findings demonstrate that IFN-β, but not IFN-λ2/3, produced via an IPS-1-IRF3 signaling pathway, is important for hMPV clearance. In the absence of a robust type I IFN-α/β response, targeting the IPS-1 signaling pathway may limit the overexuberant inflammatory response that occurs as a consequence of viral persistence., (Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Although the transcription factors IRF-3 and IRF-7 are considered master regulators of type I interferon (IFN) induction and IFN stimulated gene (ISG) expression, Irf3(-/-)×Irf7(-/-) double knockout (DKO) myeloid dendritic cells (mDC) produce relatively normal levels of IFN-β after viral infection. We generated Irf3(-/-)×Irf5(-/-)×Irf7(-/-) triple knockout (TKO) mice to test whether IRF-5 was the source of the residual induction of IFN-β and ISGs in mDCs. In pathogenesis studies with two unrelated positive-sense RNA viruses (West Nile virus (WNV) and murine norovirus), TKO mice succumbed at rates greater than DKO mice and equal to or approaching those of mice lacking the type I IFN receptor (Ifnar(-/-)). In ex vivo studies, after WNV infection or exposure to Toll-like receptor agonists, TKO mDCs failed to produce IFN-β or express ISGs. In contrast, this response was sustained in TKO macrophages following WNV infection. To define IRF-regulated gene signatures, we performed microarray analysis on WNV-infected mDC from wild type (WT), DKO, TKO, or Ifnar(-/-) mice, as well as from mice lacking the RIG-I like receptor adaptor protein MAVS. Whereas the gene induction pattern in DKO mDC was similar to WT cells, remarkably, almost no ISG induction was detected in TKO or Mavs(-/-) mDC. The relative equivalence of TKO and Mavs(-/-) responses suggested that MAVS dominantly regulates ISG induction in mDC. Moreover, we showed that MAVS-dependent induction of ISGs can occur through an IRF-5-dependent yet IRF-3 and IRF-7-independent pathway. Our results establish IRF-3, -5, and -7 as the key transcription factors responsible for mediating the type I IFN and ISG response in mDC during WNV infection and suggest a novel signaling link between MAVS and IRF-5.

Interferon (IFN) regulatory factors (IRFs) are a family of transcription factors involved in regulating type I IFN genes and other genes participating in the early antiviral host response. To better understand the mechanisms involved in virus-induced central nervous system (CNS) inflammation, we studied the influence of IRF1, -3, -7, and -9 on the transcriptional activity of key genes encoding antiviral host factors in the CNS of mice infected with lymphocytic choriomeningitis virus (LCMV). A key finding is that neither IRF3 nor IRF7 is absolutely required for induction of a type I IFN response in the LCMV-infected CNS, whereas concurrent elimination of both factors markedly reduces the virus-induced host response. This is unlike the situation in the periphery, where deficiency of IRF7 almost eliminates the LCMV-induced production of the type I IFNs. This difference is seemingly related to the local environment, as peripheral production of type I IFNs is severely reduced in intracerebrally (i.c.) infected IRF7-deficient mice, which undergo a combined infection of the CNS and peripheral organs, such as spleen and lymph nodes. Interestingly, despite the redundancy of IRF7 in initiating the type I IFN response in the CNS, the response is not abolished in IFN-β-deficient mice, as might have been expected. Collectively, these data demonstrate that the early type I IFN response to LCMV infection in the CNS is controlled by a concerted action of IRF3 and -7. Consequently this work provides strong evidence for differential regulation of the type I IFN response in the CNS versus the periphery during viral infection.

The interferon regulatory factor (IRF)3 and IRF7 are considered to play essential roles in innate immune system's antiviral responses. In this report, the full-length cDNA and genomic structure and immune response characterizations of IRF3 and IRF7 were investigated in large yellow croaker, Larimichthys crocea. The full-length cDNA of L. crocea (Lc)IRF3 was of 2204 bp, including a 5'-terminal untranslated region (UTR) of 41 bp, a 3'-terminal UTR of 774 bp and an open reading frame (ORF) of 1389 bp encoding a polypeptide of 462 amino acids residues. The full-length cDNA of LcIRF7 was of 1979 bp, including a 5'-terminal UTR of 47 bp, a 3'-terminal UTR of 636 bp and an ORF of 1296 bp encoding a polypeptide of 431 amino acids. The putative amino acid sequence of both LcIRF3 and LcIRF7 contained a typical IRF domain at the N-terminal and an IRF3 domain at the C-terminal. Furthermore, we obtained 4517 nucleotides (nt) LcIRF3 genome sequence based on the full-length cDNA, which contained 11 exons and 10 introns. The full-length genome sequence of LcIRF7 was of 3991 nucleotides, including 9 exons and 8 introns. Quantitative real-time reverse transcription PCR analysis revealed a broad expression of LcIRF3 and LcIRF7 with the most predominant expression of LcIRF3 and LcIRF7 in the liver and in the gill, respectively. The expression levels of LcIRF3 and LcIRF7 after challenged with LPS, poly I:C and Vibrio parahaemolyticus were tested in blood, spleen and liver. The results showed that the highest relative expression of LcIRF3 was in the liver at 24 h after poly I:C injection with 90 times greater than that of the non-injection group (p < 0.05). Moreover, LcIRF3 transcription increased significantly at most time point in blood and spleen tissue after poly I:C stimulation compared with that of the control group. After LPS injection, the peak value of LcIRF7 was in the liver with 207 times (at 3 h) as much as that in the control group (p < 0.05). In addition, LcIRF7 expression was significantly induced by poly I:C injection in spleen. Both LcIRF3 and LcIRF7 transcripts did not show significant change after V. parahaemolyticus stimulation. These results indicated that IRF3 and IRF7 might play an important role in large yellow croaker's defense against viral and bacterial infection., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The cytoplasmic helicase protein RIG-I (retinoic acid-inducible gene I) and downstream signaling molecules, MAVS (mitochondrial antiviral signaling protein), TRAF3 (TNF-receptor-associated factor 3) and TBK1 (TANK-binding kinase 1), have significant roles in the recognition of cytoplasmic 5'-triphosphate ssRNA and short dsRNA, and phosphorylation of IRF-3 (interferon regulatory factor 3) and IRF-7 which is responsible for the induction of type I interferons (IFN). In the present study, the full-length cDNAs of RIG-I, MAVS, TRAF3 and TBK1 were cloned and identified in common carp (Cyprinus carpio L.). The deduced protein of carp RIG-I is of 946 aa (amino acids), consisting of two CARDs (caspase-recruitment domain), a DEXDc (DExD/H box-containing domain), a HELICc (helicase superfamily c-terminal domain) and a RD (regulatory domain). Carp MAVS is of 585 aa, containing a CARD, a proline-rich region and a TM (transmembrane domain). Carp TRAF3 encodes a protein of 573 aa, including a RING (really interesting new gene), two TRAF-type zinc fingers, a coiled coil and a MATH-TRAF3 (meprin and TRAF homology) domain. Carp TBK1 is of 727 aa and contains a S&#95;TKc domain (Serine/Threonine protein kinases, catalytic domain). Carp RIG-I, MAVS, TRAF3 and TBK1 mRNAs are ubiquitously expressed in all tissues examined. In response to SVCV infection, carp RIG-I and MAVS mRNAs were up-regulated at different levels in spleen, head kidney and intestine tissues at different time points. Similarly, both carp IRF-3 and IRF-7 mRNAs were significantly up-regulated in the detected tissues. Especially in intestine, the IRF-3 and IRF-7 mRNAs of carp increased and reached 25.3-fold (at 3 dpi) and 224.7-fold (at 5 dpi). Noteworthily, a significant growth of carp TRAF3 and TBK1 mRNA was also mainly found in intestine (7.0-fold and 11.3-fold at 5 dpi, respectively). These data implied that the expression profiles of IRF-3/-7 mRNAs in carp correlate with the up-regulation of RIG-I/MAVS/TRAF3/TBK, and carp RIG-I and MAVS may be involved in antiviral responses through the RIG-I viral recognition signaling pathway in a TRAF3/TBK1-dependent manner., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Innate immune responses contribute to synovial inflammation in rheumatoid arthritis. The present study was designed to investigate the contribution of IFN regulatory factor (IRF)3 and IRF7 to type I IFN-regulated gene expression in synoviocytes. Fibroblast-like synoviocytes were stimulated with polyinosinic-polycytidylic acid (poly [I-C]) after transfection with IRF3 or IRF7 small interfering RNA to knockdown transcription factor expression. Western blots, luciferase assay after transfection with reporter constructs, quantitative PCR, and AP-1 DNA binding ELISA were performed to evaluate the role of IRF3 and IRF7 in poly (I-C)-induced signaling and synoviocyte gene expression. IRF3 regulates IFN-stimulated response element (ISRE) promoter activity as well as IFN-beta, IRF5, IRF7, RANTES, IFN-inducible protein-10, MCP-1, and MIP1alpha gene expression in response to poly (I-C). IRF7 knockdown modestly decreased a subset of genes and ISRE activity, although the results were not statistically significant. Surprisingly, IRF3 knockdown almost completely blocked expression of additional genes in which the ISRE is not traditionally considered a dominant promoter site in fibroblast-like synoviocytes, including matrix metalloproteinase (MMP)3, MMP9, IL-6, and IL-8. Transcription factor activation studies demonstrated a role for IRF3 in regulation of c-Jun phosphorylation and AP-1 binding. IRF3 rather than IRF7 regulates poly (I-C)-induced type I IFN responses in human synoviocytes by increasing ISRE promoter activity. IRF3 also partially regulates expression of other cytokines and MMP through activation of c-Jun and the AP-1 promoter site. Targeting synoviocyte IRF3 represents a potential approach to suppress diverse mediators while limiting suppression of IRF7-mediated immune responses.

Amplicons are helper-dependent herpes simplex virus type 1 (HSV-1)-based vectors that can deliver very large, foreign DNA sequences and, as such, are good candidates for both gene delivery and vaccine development. However, many studies have shown that innate immune responses induced by virus vectors can play a significant role in the control of transgenic expression and in the induction of inflammatory responses. Furthermore, amplicons are very interesting tools to study innate cellular responses elicited by entry of HSV-1 particles in the absence of any virus gene expression. For these reasons, in this study we characterized the innate antiviral response established in human fibroblasts of limited passage (HFFF-2) infected by amplicons. Our results indicate that infection with amplicons triggered an interferon (IFN)-regulatory factors 3 and 7 (IRF3/7)-dependent antiviral response, rendered the cells resistant to vesicular stomatitis virus infection and induced significant changes in the pattern of cellular gene expression, including the upregulation of Toll-like receptor 3 (TLR3), IRF7 and IFN-stimulated genes (ISGs). In contrast, we observed only a mild and contained type I IFN response in infected cells. Amplicon infection induced nuclear translocation and subsequent degradation of IRF3, without hyperphosphorylation of the protein. Inhibition of endosome-resident TLR signalling by blocking lysosome maturation or the knockdown of TLR3 and 4 did not abolish the cellular response to amplicons, whereas knockdown of IRF3 and 7 inhibited ISG and IFN-beta expression severely. Therefore, our results confirm the existence of TLR-independent, IRF3/7-dependent activation pathways triggered by HSV-1 particles in human fibroblasts.

IFN regulatory factor 7 (IRF7) has been described as the master regulator of type I IFN responses and has been shown to be critical for innate antiviral immunity in vivo. In addition to type I IFN, NK cell responses are involved in the control of viral replication during acute viral infection. To investigate the role of IRF7 in the context of a viral infection that induces a strong NK cell response, the murine cytomegalovirus (MCMV) infection model was used. WT, IRF7-deficient and IRF3/IRF7-double deficient mice were infected with MCMV. The systemic IFN-alpha response to MCMV was entirely dependent on IRF7, but independent of IRF3. However, peak IFN-beta production during MCMV infection was not affected by the lack of IRF7 or both IRF7 and IRF3. Despite the complete lack of IFN-alpha production IRF7- and IRF3/IRF7-deficient mice were surprisingly efficient in controlling MCMV replication and were only modestly more susceptible to MCMV infection than WT mice. NK cell cytotoxicity was unimpaired and NK cell IFN-gamma production was enhanced in IRF7-deficient mice correlating with increased levels of bioactive IL-12. Owing to these compensatory mechanisms IRF7-dependent antiviral immune responses were not essential for resistance against acute MCMV infection in vivo.

Interferon regulatory factors (IRF) 3 and 7 in mammals are known to be crucial in regulating the type I interferon (IFN) response to viral infection as part of transcriptional complexes binding to IRF-binding elements (IRF-Es) and interferon stimulatory response elements (ISREs) within IFN and interferon-stimulated genes (ISGs). Here we report the sequencing and characterization of full-length cDNA homologues of rainbow trout (rt)IRF7 and, for the first time in fish, IRF3. RtIRF3 consists of 2127 bp with a 159 bp 5'-UTR-containing two upstream AUGs and a 573 bp 3'-UTR. RtIRF7 was found to be 2055 bp, with a 102 bp 5'-UTR and a 705 bp 3'-UTR. The open reading frames (ORFs) translate into 464 amino acid and 415 amino acid proteins, respectively, each possessing a putative DNA-binding domain (DBD) containing a tryptophan cluster, which is characteristic of all IRF family members. The presence of putative IRF association domain (IAD)s, serine-rich C terminal domains (poorly conserved in trout IRF3), and phylogenetic analysis places the two genes in the IRF3 subfamily. Both genes were found to be upregulated by poly I:C, type I recombinant rainbow trout (r) IFN (second isoform, type I rIFN), type II rIFN (rIFNgamma), LPS, and rIL-1beta in the trout macrophage cell line, RTS-11. Poly I:C and type I rIFN also induced IRF3 and IRF7 expression in a trout fibroblast cell line (RTG-2). Transient transfection of RTG-2 cells with each IRF fused to GFP revealed a predominant cytoplasmic distribution found most intensely around the nucleus and, to a lesser extent, within cell nuclei. Transient transfection of rtIRF3 in the Mx-1-luciferase reporter cell line, RTG-P1, revealed a modest increase in luciferase activity relative to the vehicle control, which was lost in cells over-expressing a DBD-truncated form of rtIRF3. Both full-length and DBD-truncated forms of rtIRF7 increased reporter activity relative to the control, although to a non-significant extent. Electromobility shift assays (EMSAs) did not reveal a specific interaction between each IRF and the ISRE element found in the Mx-1 promoter, although the Mx-1 ISRE bound specifically to endogenous transcriptional complexes. These data support the premise that rtIRF3 and rtIRF7 are important molecules in the regulation of antiviral responses in fish, with the impact of rIFNgamma on rtIRF3/7 expression implying a role for these IRFs in immune processes other than type I IFN-driven antiviral responses.

Rapid induction of type I interferon (IFN) expression is a central event in the establishment of the innate immune response against viral infection and requires the activation of multiple transcriptional proteins following engagement and signaling through Toll-like receptor-dependent and -independent pathways. The transcription factor interferon regulatory factor-3 (IRF-3) contributes to a first line of defense against viral infection by inducing the production of IFN-beta that in turn amplifies the IFN response and the development of antiviral activity. In murine knock-out models, the absence of IRF-3 and the closely related IRF-7 ablates IFN production and increases viral pathogenesis, thus supporting a pivotal role for IRF-3/IRF-7 in the development of the host antiviral response.

Type I IFN (IFN-alpha/beta) have important biological functions ranging from immune cell development and activation, to tumor cell killing and most importantly inhibition of virus replication. Following viral infection or activation of Toll-like receptors (TLRs) via distinct ligands, IFN-alpha/beta are produced. Two members of the interferon regulatory factor (IRF) family - IRF-3 and IRF-7 - are the major modulators of IFN gene expression. Activation of IRF-3 and IRF-7 by TBK1/IKKvarepsilon mediated phosphorylation promotes IFN gene expression and potentiates the production of IFN responsive genes important to the development of an effective antiviral immune response. IFN treatment can augment anti-tumor properties and they are potentially key players in cancer therapy. For example, adoptive transfer of IFN-gamma-activated macrophages can mediate tumor cell killing via direct cell-cell contact, as well as release of soluble cytotoxic pro-inflammatory molecules. A recent study investigated whether IRF-3 and IRF-7 could mediate the acquisition of new anti-tumor effector functions in macrophages. Adenovirus mediated transduction of the active form of IRF-7 into primary macrophages resulted in the production of type I IFN, upregulation of target genes including TRAIL and increased tumoricidal activity of macrophages; in contrast, the active form of IRF-3 led to induction of cell death. These studies indicate that IRF-7 transduced macrophages may be an attractive candidate for in vivo adoptive therapy of cancer.