I Interferon Transcription Research Articles

FADD promotes type I interferon production to suppress porcine reproductive and respiratory syndrome virus infection.

Porcine reproductive and respiratory syndrome (PRRS) is an epidemic animal infectious disease worldwide, causing huge economic losses to the global swine industry. Fas-associated death domain (FADD) was previously reported to be an adaptor protein that functions in transferring the apoptotic signals regulated by the death receptors. In the current study, we unravel its unidentified role in promoting type I interferon (IFN) production during PRRS virus (PRRSV) infection. We identified that FADD inhibited PRRSV infection via promotion of type I IFN transcription. Overexpression of FADD suppressed the replication of PRRSV, while knockout of FADD increased viral titer and nucleocapsid protein expression. Mechanistically, FADD promoted mitochondrial antiviral signaling protein (MAVS)-mediated production of IFN-β and some IFN-stimulated genes (ISGs). Furthermore, FADD exerted anti-PRRSV effects in a MAVS-dependent manner and increased the type I IFN signaling during PRRSV infection. This study highlights the importance of FADD in PRRSV replication, which may have implications for the future control of PRRS.

Kikuchi-Fujimoto disease is mediated by an aberrant type I interferon response

Kikuchi-Fujimoto disease (KFD) is a reactive lymphadenitis of unclear etiology. To understand the pathogenesis of KFD, we performed targeted RNA sequencing of a well-characterized cohort of 15 KFD specimens with 9 non-KFD lymphadenitis controls. Two thousand and three autoimmunity-related genes were evaluated from archived formalin-fixed paraffin-embedded lymph node tissue and analyzed by a bioinformatics approach. Differential expression analysis of KFD cases compared to controls revealed 44 significantly upregulated genes in KFD. Sixty-eight percent of these genes were associated with the type I interferon (IFN) response pathway. Key component of the pathway including nucleic acid sensors, IFN regulatory factors, IFN-induced antiviral proteins, IFN transcription factors, IFN-stimulated genes, and IFN-induced cytokines were significantly upregulated. Unbiased gene expression pathway analysis revealed enrichment of IFN signaling and antiviral pathways in KFD. Protein–protein interaction analysis and a molecular complex detection algorithm identified a densely interacting 15-gene module of type I IFN pathway genes. Apoptosis regulator IFI6 was identified as a key seed gene. Transcription factor target analysis identified enrichment of IFN-response elements and IFN-response factors. T-cell-associated genes were upregulated while myeloid and B-cell-associated genes were downregulated in KFD. CD123+ plasmacytoid dendritic cells (PDCs) and activated T cells were noted in KFD. In conclusion, KFD is mediated by an aberrant type I interferon response that is likely driven by PDCs and T cells.

Role of the transcriptional regulator SP140 in resistance to bacterial infections via repression of type I interferons.

Type I interferons (IFNs) are essential for anti-viral immunity, but often impair protective immune responses during bacterial infections. An important question is how type I IFNs are strongly induced during viral infections, and yet are appropriately restrained during bacterial infections. The Super susceptibility to tuberculosis 1 (Sst1) locus in mice confers resistance to diverse bacterial infections. Here we provide evidence that Sp140 is a gene encoded within the Sst1 locus that represses type I IFN transcription during bacterial infections. We generated Sp140-/- mice and found that they are susceptible to infection by Legionella pneumophila and Mycobacterium tuberculosis. Susceptibility of Sp140-/- mice to bacterial infection was rescued by crosses to mice lacking the type I IFN receptor (Ifnar-/-). Our results implicate Sp140 as an important negative regulator of type I IFNs that is essential for resistance to bacterial infections.

Divergent effects of acute versus chronic glucocorticoids in COVID-19.

Divergent effects of acute versus chronic glucocorticoids in COVID-19.

The Cytomegalovirus Tegument Protein UL35 Antagonizes Pattern Recognition Receptor-Mediated Type I IFN Transcription.

The rapid activation of pattern recognition receptor (PRR)-mediated type I interferon (IFN) signaling is crucial for the host response to infection. In turn, human cytomegalovirus (HCMV) must evade this potent response to establish life-long infection. Here, we reveal that the HCMV tegument protein UL35 antagonizes the activation of type I IFN transcription downstream of the DNA and RNA sensors cGAS and RIG-I, respectively. We show that ectopic expression of UL35 diminishes the type I IFN response, while infection with a recombinant HCMV lacking UL35 induces an elevated type I IFN response compared to wildtype HCMV. With a series of luciferase reporter assays and the analysis of signaling kinetics upon HCMV infection, we observed that UL35 downmodulates PRR signaling at the level of the key signaling factor TANK-binding kinase 1 (TBK1). Finally, we demonstrate that UL35 and TBK1 co-immunoprecipitate when co-expressed in HEK293T cells. In addition, we show that a previously reported cellular binding partner of UL35, O-GlcNAc transferase (OGT), post-translationally GlcNAcylates UL35, but that this modification is not required for the antagonizing effect of UL35 on PRR signaling. In summary, we have identified UL35 as the first HCMV protein to antagonize the type I IFN response at the level of TBK1, thereby enriching our understanding of how this important herpesvirus escapes host immune responses.

Porcine sialoadhesin suppresses type I interferon production to support porcine reproductive and respiratory syndrome virus infection

Porcine reproductive and respiratory syndrome virus (PRRSV) is a significant threat to the global swine industry. Porcine sialoadhesin (poSn) has been previously shown to mediate PRRSV attachment and internalization. In the current study, we report its unidentified role in antagonism of type I interferon (IFN) production during PRRSV infection. We determined that poSn facilitated PRRSV infection via inhibition of type I IFN transcription. Mechanistically, poSn interacted with a 12 kDa DNAX-activation protein (DAP12), which was dependent on residues 51–57 within DAP12 transmembrane domain (TMD). PRRSV exploited the poSn-DAP12 pathway to attenuate activation of nuclear factor-kappa B (NF-κB). More importantly, the poSn-DAP12 pathway was involved in inhibiting poly (I:C)-triggered IFN production. All these results reveal a novel role of poSn in suppressing host antiviral responses, which deepens our understanding of PRRSV pathogenesis.

Duck karyopherin α4 (duKPNA4) is involved in type I interferon expression and the antiviral response against Japanese encephalitis virus

Karyopherin α4 (KPNA4) is an adaptor molecule that mediates type I interferon (IFN) production by facilitating the nuclear translocation of IFN transcription factors. Here, we cloned the duck KPNA4 (duKPNA4) gene and analyzed its involvement in type I IFN expression as well as antiviral response against Japanese encephalitis virus (JEV). The full-length duKPNA4 gene encoded a 520-amino acid protein that shared 97.3–98.7% sequence similarity with its orthologues in chickens, humans and mice. The duKPNA4 was extensively expressed in various duck tissues at the mRNA level. Analysis of the subcellular localization of duKPNA4 by immunofluorescence assays indicated that the duKPNA4 was primarily distributed in both the cytoplasm and nucleus in primary duck embryonic fibroblasts (DEFs). However, it translocated from the cytoplasm to the nucleus in response to poly(I:C) stimulation or JEV infection. The duKPNA4 interacted with duck IFN regulatory factor 7 and facilitated its nuclear translocation, thereby up-regulating the expression of IFN-α and IFN-β in DEFs in the presence of poly(I:C) stimulation. Exogenous expression of duKPNA4 significantly elevated the expression of IFN-α and IFN-β induced by JEV infection and inhibited JEV replication in DEFs. These data demonstrate the importance of duKPNA4 in type I IFN signaling as well as the antiviral response against JEV replication.

TRAF3 enhances STING-mediated antiviral signaling during the innate immune activation of black carp

Tumor necrosis factor receptor-associated factor 3 (TRAF3) is a main regulator of antiviral and anti-inflammatory pathways in mammals, which is considered to induce type I interferon (IFN) activation and negatively regulate the activation of the canonical and non-canonical NF-κB pathways. To elucidate its function in teleost fish, TRAF3 homologue of black carp (Mylopharyngodon piceus) has been cloned and characterized in this study. The open reading frame (ORF) of black carp TRAF3 (bcTRAF3) consists of 1722 nucleotides and bcTRAF3 contains 574 amino acids. bcTRAF3 protein migrated around 65 KDa in immunoblot analysis of both EPC and HEK293T cells. bcTRAF3 was identified as a cytosolic protein and suggested to form aggregates or be associated with vesicles scattering in the cytoplasm. It was interesting that both NF-κB and IFN transcription was activated by bcTRAF3 in reporter assay. When co-expressed with black carp STING (bcSTING), bcTRAF3 was redistributed in the cytoplasm and its subcellular location overlapped with that of bcSTING no matter what the cells was infected with GCRV or not, which suggested the association between these two molecules. bcSTING-mediated IFN production was up-regulated by bcTRAF3 in a dose dependent manner in reporter assay. Accordingly, EPC cells transfected with both bcSTING and bcTRAF3 showed enhanced antiviral activity comparing EPC cells expressing bcSTING alone. Taken together, the data generated in this paper supported the conclusion that bcTRAF3 was recruited into host innate immune activation and positively regulated bcSTING-mediated antiviral signaling.

The murine cytomegalovirus M35 protein antagonizes type I IFN induction downstream of pattern recognition receptors by targeting NF-κB mediated transcription.

The type I interferon (IFN) response is imperative for the establishment of the early antiviral immune response. Here we report the identification of the first type I IFN antagonist encoded by murine cytomegalovirus (MCMV) that shuts down signaling following pattern recognition receptor (PRR) sensing. Screening of an MCMV open reading frame (ORF) library identified M35 as a novel and strong negative modulator of IFNβ promoter induction following activation of both RNA and DNA cytoplasmic PRR. Additionally, M35 inhibits the proinflammatory cytokine response downstream of Toll-like receptors (TLR). Using a series of luciferase-based reporters with specific transcription factor binding sites, we determined that M35 targets NF-κB-, but not IRF-mediated, transcription. Expression of M35 upon retroviral transduction of immortalized bone marrow-derived macrophages (iBMDM) led to reduced IFNβ transcription and secretion upon activation of stimulator of IFN genes (STING)-dependent signaling. On the other hand, M35 does not antagonize interferon-stimulated gene (ISG) 56 promoter induction or ISG transcription upon exogenous stimulation of the type I IFN receptor (IFNAR). M35 is present in the viral particle and, upon MCMV infection of fibroblasts, is immediately shuttled to the nucleus where it exerts its immunomodulatory effects. Deletion of M35 from the MCMV genome and hence from the viral particle resulted in elevated type I IFN transcription and secretion in vitro and in vivo. In the absence of M35, lower viral titers are observed during acute infection of the host, and productive infection in the salivary glands was not detected. In conclusion, the M35 protein is released by MCMV immediately upon infection in order to deftly inhibit the antiviral type I IFN response by targeting NF-κB-mediated transcription. The identification of this novel viral protein reinforces the importance of timely countermeasures in the complex relationship between virus and host.

Depletion of elongation initiation factor 4E binding proteins by CRISPR/Cas9 enhances the antiviral response in porcine cells

Type I interferons (IFNs) are key mediators of the innate antiviral response in mammalian cells. Elongation initiation factor 4E binding proteins (4E-BPs) are translational controllers of interferon regulatory factor 7 (IRF-7), the “master regulator” of IFN transcription. Previous studies have suggested that mouse cells depleted of 4E-BPs are more sensitive to IFNβ treatment and had lower viral loads as compared to wild type (WT) cells. However, such approach has not been tested as an antiviral strategy in livestock species. In this study, we tested the antiviral activity of porcine cells depleted of 4E-BP1 by a Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR-associated protein-9 nuclease (Cas9) genome engineering system. We found that 4E-BP1 knockout (KO) porcine cells had increased expression of IFNα and β, IFN stimulated genes, and significant reduction in vesicular stomatitis virus titer as compare to WT cells. No phenotypical changes associated with CRISPR/Cas9 manipulation were observed in 4E-BP1 KO cells. This work highlights the use of the CRISPR/Cas9 system to enhance the antiviral response in porcine cells.

Finding a fairy in the forest: ELF4, a novel and critical element of type I interferon responses

Type I interferon (IFN) production is a key event during innate immune responses. This early and prompt mechanism is mediated by multiple factors that involve various pattern recognition receptors (PRRs), adaptor proteins, kinases and transcription factors. Stimulator of IFN genes (STING) is an important transmembrane adaptor protein that plays a role in the activation of downstream transcription factors, such as IFN regulatory factor 3 (IRF3) and signal transducer and activator of transcription 6, via TANK-binding kinase 1 (TBK1), which has been considered the main innate immune defense weaponry against viruses and intracellular pathogens. In a current issue of Nature Immunology, You et al. showed that, after viral infection, the ETS-related transcription factor ELF4 (previously known as MEF) interacts with STING and is consequently activated by TBK1. This process leads to the nuclear translocation of ELF4 and its binding to IFN promoters (Figure 1). The group also demonstrated that ELF4 acts as an IFN transcription factor, as it has the potential to increase the binding affinity of IRF3 and IRF7 through cooperative binding to newly identified enhancer elements (EICE) in the IFN promoters.1 These new findings not only extend our knowledge about the details of the fine-tuning of the innate IFN responses, but also represent a new milestone by describing the role of EICE elements in this process.

Virulence Factor NSs of Rift Valley Fever Virus Recruits the F-Box Protein FBXO3 To Degrade Subunit p62 of General Transcription Factor TFIIH

The nonstructural protein NSs is the main virulence factor of Rift Valley fever virus (RVFV; family Bunyaviridae, genus Phlebovirus), a serious pathogen of livestock and humans in Africa. RVFV NSs blocks transcriptional upregulation of antiviral type I interferons (IFN) and destroys the general transcription factor TFIIH subunit p62 via the ubiquitin/proteasome pathway. Here, we identified a subunit of E3 ubiquitin ligases, F-box protein FBXO3, as a host cell interactor of NSs. Small interfering RNA (siRNA)-mediated depletion of FBXO3 rescued p62 protein levels in RVFV-infected cells and elevated IFN transcription by 1 order of magnitude. NSs interacts with the full-length FBXO3 protein as well as with a truncated isoform that lacks the C-terminal acidic and poly(R)-rich domains. These isoforms are present in both the nucleus and the cytoplasm. NSs exclusively removes the nuclear pool of full-length FBXO3, likely due to consumption during the degradation process. F-box proteins form the variable substrate recognition subunit of the so-called SCF ubiquitin ligases, which also contain the constant components Skp1, cullin 1 (or cullin 7), and Rbx1. siRNA knockdown of Skp1 also protected p62 from degradation, suggesting involvement in NSs action. However, knockdown of cullin 1, cullin 7, or Rbx1 could not rescue p62 degradation by NSs. Our data show that the enzymatic removal of p62 via the host cell factor FBXO3 is a major mechanism of IFN suppression by RVFV. Rift Valley fever virus is a serious emerging pathogen of animals and humans. Its main virulence factor, NSs, enables unhindered virus replication by suppressing the antiviral innate immune system. We identified the E3 ubiquitin ligase FBXO3 as a novel host cell interactor of NSs. NSs recruits FBXO3 to destroy the general host cell transcription factor TFIIH-p62, resulting in suppression of the transcriptional upregulation of innate immunity.

PIAS1 negatively modulates virus triggered type I IFN signaling by blocking the DNA binding activity of IRF3

During viral infection, production of proinflammatory cytokines including type I interferons (IFNs) is under stringent control to avoid detrimental overreaction. The protein inhibitor of activated STAT (PIAS) family proteins have been recognized as anti-inflammatory molecules by restraining type I IFN induced amplifying signaling. Here we identified PIAS1 as an important negative regulator of virus-triggered type I IFN signaling. Overexpression of PIAS1 repressed virus-or RIG-I like receptor stimulated type I IFN transcription, whereas knockdown of PIAS1 expression augmented virus-induced production of type I IFNs. PIAS1 with a mutation in the SAP domain retained the inhibitory function in virus-induced IFN transcription, but abolished the inhibition in IFN-stimulated signaling. SUMO E3 ligase activity dead mutant PIAS1/C350S still had the comparable inhibitory function with WT PIAS1. Further study indicated that PIAS1 interacted with IRF3 and inhibited the DNA binding activity of IRF3. The C-terminal region of PIAS1 around a cluster of acidic amino acids is critical for the interaction with IRF3 and the inhibitory functions of PIAS1. Therefore, these results unveil PIAS1 functions both at the virus-induced early signaling stage and IFN stimulated amplifying stage with distinct mechanisms. PIAS1 is important in maintaining proper amounts of type I IFNs and restrains its magnitude when the antiviral response intensifies.

RING Finger Protein 11 Targets TBK1/IKKi Kinases to Inhibit Antiviral Signaling

A key feature of the innate antiviral immune response is a rapid nonspecific response to virus infection largely mediated by the induction and extracellular secretion of type I interferons (IFNs) that restrict virus replication. Cytoplasmic sensors such as RIG-I recognize viral RNA and trigger antiviral signaling pathways that upregulate IFN transcription. However, it remains largely unknown how antiviral signaling is negatively regulated to maintain homeostasis after the elimination of virus. In this report, we have identified the RING domain-containing protein RING finger 11 (RNF11) as a novel negative regulator of innate antiviral signaling. Overexpression of RNF11 downregulated IFN-β expression and enhanced viral replication whereas siRNA-mediated knockdown of RNF11 suppressed viral replication. RNF11 interacted with the noncanonical IKK kinases TBK1/IKKi and attenuated their Lys63-linked polyubiquitination by blocking interactions with the E3 ligase TRAF3. The inhibitory function of RNF11 was dependent on the ubiquitin-binding adaptor molecule TAX1BP1 which was required for RNF11 to target TBK1/IKKi. Collectively, these results indicate that RNF11 functions together with TAX1BP1 to restrict antiviral signaling and IFN-β production.

NEMO Binds Ubiquitinated TANK-Binding Kinase 1 (TBK1) to Regulate Innate Immune Responses to RNA Viruses

RIG-I-like receptors (RLR) are intracellular sensors utilized by nearly all cell types for recognition of viral RNA, initiation of antiviral defense, and induction of type I interferons (IFN). TBK1 is a critical kinase implicated in RLR-dependent IFN transcription. Posttranslational modification of TBK1 by K63-linked ubiquitin is required for RLR driven signaling. However, the TBK1 ubiquitin acceptor sites and the function of ubiquitinated TBK1 in the signaling cascade are unknown. We now show that TBK1 is ubiquitinated on residues K69, K154, and K372 in response to infection with RNA virus. The K69 and K154 residues are critical for innate antiviral responses and IFN production. Ubiquitinated TBK1 recruits the downstream adaptor NEMO through ubiquitin binding domains. The assembly of the NEMO/TBK1 complex on the mitochondrial protein MAVS leads to activation of TBK1 kinase activity and phosphorylation of the transcription factor, interferon response factor 3. The combined results refine current views of RLR signaling, define the role of TBK1 polyubiquitination, and detail the mechanisms involved in signalosome assembly.

NEMO recruitment of polyubiquitinated TBK1 to MAVS builds signalosome to regulate innate anti-viral immunity (108.3)

Abstract RIG-I-like receptors (RLRs) are intracellular sensors utilized by nearly all cell types for recognition of viral RNA, initiation of antiviral defense, and induction of type I interferons (IFN). TBK1 is a critical kinase implicated in RLR-dependent IFN transcription. The E3 ligases Mind bomb 1 (MIB1) and Mind bomb 2 (MIB2) conjugate TBK1 with K63-linked polyubiquitin on residues K69 and K154. TBK1 polyubiquitination is critical for recruitment of the downstream adaptor, NEMO. After viral infection NEMO and IRF3 are independently recruited to MAVS. NEMO bridges TBK1 to MAVS through ubiquitin binding domains. The assembly of the NEMO/TBK1 complex on MAVS leads to activation of TBK1 kinase activity and phosphorylation of the transcription factor, IRF3. In contrast, activation of NF-kB is not impaired by MIB deficiency. The combined results refine current views of RLR signaling, define the role of TBK1 polyubiquitination and detail the assembly of the MAVS signalosome.

Abstract 5396: Interferon-α receptor dependent and independent inhibition of type I interferon production in plasmacytoid dendritic cells from mice and melanoma patients

Abstract IFN-α is pleiotropic cytokine belonging to the type I Interferon (IFNs) family and can induce multiple biological effects, e.g. induction of apoptosis and inhibition of cell growth. These cytokines exhibit a long record of clinical use in patients with some types of cancer and viral diseases. Notably, certain autoimmune disorders have been postulated to be mediated by endogenous IFN-α and are often observed in some IFN-α-treated cancer patients. How IFN-α promotes the generation of antitumor T-cell immunity and its regulation mechanisms are still unclear. To address these questions, we focus on the main producer of IFNs, Plasmacytoid dendritic cells (pDCs), which reside in bone marrow and lymphoid organs and typically secrete type I IFNs on Toll-like receptor (TLR) triggering. We detected cytokines production by human pDCs from leukapheresis cells of melanoma patients who were treated with IFNs for 25 weeks. After stimulation with CpG-A, these pDCs after treatment (IFN-α-pDCs) produce 70% less IFN-α compare to before treatment, but compatible amount of other inflammatory cytokines e.g. IL-6. We observed the similar phenomena in murine models either after VSV infection or by systemic IFN-α delivery. We showed that these IFN-α-pDCs possess more active and mature morphology as DCs and they are particularly more effective in inducing T- and NK-cell immunity by increasing cell proliferation and differentiation. Of note, IFN-α-pDCs skewed the naïve CD4+ T cells to a Th17 antigen-presenting subset by producing more IL-17 but less IFN-γ compare to wide type pDCs. The ensemble of these results suggests that IFN-α-pDCs could be successfully used in strategies of cancer immunotherapy, especially in melanoma patients. We also demonstrate that pDCs possess two fundamental, but separate, physiologic mechanisms for negative feedback control of IFN-α production after IFNs treatment in melanoma patients. Both signaling through the canonical TLR-MyD88 pathway and feedback inhibition from IFN-α itself through the IFN-α/βR render pDCs unable to synthesize type I interferon upon TLR triggering. Mechanistically, we found that IFN-α-pDCs show increased IRF7 mRNA and protein upon TLR stimulation, but despite efficient nuclear IRF-7 translocation, type I IFN was not produced. Interestingly, Ifn-α6 promoter activity was blocked in the IFN-α-pDCs, suggesting a block of type I IFN transcription despite nuclear IRF-7 translocation. Taken together, triggering of TLR and of IFN-α/βR act independently to tightly limit systemic IFN-α production after IFNs treatment. All this reveals the complexity of the IFN-α-pDC interactions under normal and pathological conditions and stimulates further studies for identifying optimal modalities in either using these cytokines or controlling their production in melanoma patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5396. doi:1538-7445.AM2012-5396

Dcp2 Decapping Protein Modulates mRNA Stability of the Critical Interferon Regulatory Factor (IRF) IRF-7

The mammalian Dcp2 mRNA-decapping protein functions primarily on a subset of mRNAs in a transcript-specific manner. Here we show that Dcp2 is an important modulator of genes involved in the type I interferon (IFN) response, which is the initial line of antiviral innate immune response elicited by a viral challenge. Mouse embryonic fibroblast cells with reduced Dcp2 levels (Dcp2(β/β)) contained significantly elevated levels of mRNAs encoding proteins involved in the type I IFN response. In particular, analysis of a key type I IFN transcription factor, IFN regulatory factor 7 (IRF-7), revealed an increase in both IRF-7 mRNA and protein in Dcp2(β/β) cells. Importantly, the increase in IRF-7 mRNA within the background of reduced Dcp2 levels was attributed to a stabilization of the IRF-7 mRNA, suggesting that Dcp2 normally modulates IRF-7 mRNA stability. Moreover, Dcp2 expression was also induced upon viral infection, consistent with a role in attenuating the antiviral response by promoting IRF-7 mRNA degradation. The induction of Dcp2 levels following a viral challenge and the specificity of Dcp2 in targeting the decay of IRF-7 mRNA suggest that Dcp2 may negatively contribute to the innate immune response in a negative feedback mechanism to restore normal homeostasis following viral infection.

Primary Sjogren’s Syndrome and the Type I Interferon System

Patients with primary Sjögren's syndrome (pSS) have an activated type I interferon (IFN) system that contribute to the etiopathogenesis and clinical manifestations of the disease. The type I IFN system consists of the stimuli for type I IFN production, the receptors, cells and transcription factors involved in the synthesis of type I IFNs, the type I IFN-receptor and the effects on target cells. Increased type I IFN activity has been demonstrated in sera from patients with pSS and IFN-α, the main type I IFN, has been detected in the minor salivary glands. Gene expression profiling of peripheral blood mononuclear cells (PBMCs) and minor salivary glands from pSS patients display an up-regulation of type I IFN-induced genes, an "IFN signature". The professional IFN-α producing plasmacytoid dendritic cell (pDC) shows a reduced frequency in the peripheral blood, but has been detected in the salivary glands, possibly due to tissue recruitment. Polymorphisms in the interferon regulatory factor 5 (IRF5) and signal transducer and activator of transcription 4 (STAT4) genes in the type I IFN system, are associated with increased risk for pSS. A postulated disease model is that an initial viral infection induces type I IFN production in the salivary glands with subsequent activation of the adaptive immune system resulting in the production of autoantibodies against the RNA-binding proteins SSA/SSB/RNP. Interferogenic immune complexes are formed, which trigger the pDCs to an ongoing type I IFN production, which sustain the disease process. Potential therapeutic targets can be identified within the type I IFN system.

PIASy Inhibits Virus-induced and Interferon-stimulated Transcription through Distinct Mechanisms

The protein inhibitor of activated STAT (PIAS) family proteins regulates innate immune responses by controlling transcription induced by Toll-like receptor, RIG-I-like receptor signaling, and JAK/STAT pathways. Here, we show that PIASy negatively regulates type I interferon (IFN) transcription. Virus infection led to enhanced type I IFN induction in PIASy null cells, and conversely PIASy overexpression reduced IFN transcription. A mutation in the LXXLL motif of the SAP domain abolished inhibition of IFN-stimulated gene expression but did not affect virus or Toll-like receptor/RIG-I-like receptor-stimulated IFN transcription, indicating that PIASy employs distinct mechanisms to inhibit virus-induced and IFN-stimulated transcription. SUMO E3 activity was not required for PIASy inhibition of IFN transcription; however, PIASy relied on the SUMO modification mechanism to inhibit IFN transcription, because the activity of the SUMO-interacting motif was required for inhibition, and knockdown of SUMO E2 enzyme UBC9 decreased inhibitory activity of PIASy. Our results demonstrate that PIASy negatively regulates both IFN transcription and IFN-stimulated gene expression through multiple mechanisms utilizing the function of different domains.