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BACKGROUND: We previously reported that impaired type I IFN activity, due to inborn errors of TLR3- and TLR7-dependent type I interferon (IFN) immunity or to autoantibodies against type I IFN, account for 15-20% of cases of life-threatening COVID-19 in unvaccinated patients. Therefore, the determinants of life-threatening COVID-19 remain to be identified in ~ 80% of cases. METHODS: We report here a genome-wide rare variant burden association analysis in 3269 unvaccinated patients with life-threatening COVID-19, and 1373 unvaccinated SARS-CoV-2-infected individuals without pneumonia. Among the 928 patients tested for autoantibodies against type I IFN, a quarter (234) were positive and were excluded. RESULTS: No gene reached genome-wide significance. Under a recessive model, the most significant gene with at-risk variants was TLR7, with an OR of 27.68 (95%CI 1.5-528.7, P = 1.1 × 10-4) for biochemically loss-of-function (bLOF) variants. We replicated the enrichment in rare predicted LOF (pLOF) variants at 13 influenza susceptibility loci involved in TLR3-dependent type I IFN immunity (OR = 3.70[95%CI 1.3-8.2], P = 2.1 × 10-4). This enrichment was further strengthened by (1) adding the recently reported TYK2 and TLR7 COVID-19 loci, particularly under a recessive model (OR = 19.65[95%CI 2.1-2635.4], P = 3.4 × 10-3), and (2) considering as pLOF branchpoint variants with potentially strong impacts on splicing among the 15 loci (OR = 4.40[9%CI 2.3-8.4], P = 7.7 × 10-8). Finally, the patients with pLOF/bLOF variants at these 15 loci were significantly younger (mean age [SD] = 43.3 [20.3] years) than the other patients (56.0 [17.3] years; P = 1.68 × 10-5). CONCLUSIONS: Rare variants of TLR3- and TLR7-dependent type I IFN immunity genes can underlie life-threatening COVID-19, particularly with recessive inheritance, in patients under 60 years old. ispartof: Genome Med vol:15 issue:1 pages:22- ispartof: location:England status: Published online

BackgroundWe previously reported that impaired type I IFN activity, due to inborn errors of TLR3- and TLR7-dependent type I interferon (IFN) immunity or to autoantibodies against type I IFN, account for 15-20% of cases of life-threatening COVID-19 in unvaccinated patients. Therefore, the determinants of life-threatening COVID-19 remain to be identified in ~ 80% of cases.MethodsWe report here a genome-wide rare variant burden association analysis in 3269 unvaccinated patients with life-threatening COVID-19, and 1373 unvaccinated SARS-CoV-2-infected individuals without pneumonia. Among the 928 patients tested for autoantibodies against type I IFN, a quarter (234) were positive and were excluded.ResultsNo gene reached genome-wide significance. Under a recessive model, the most significant gene with at-risk variants was TLR7, with an OR of 27.68 (95%CI 1.5-528.7, P = 1.1 × 10-4) for biochemically loss-of-function (bLOF) variants. We replicated the enrichment in rare predicted LOF (pLOF) variants at 13 influenza susceptibility loci involved in TLR3-dependent type I IFN immunity (OR = 3.70[95%CI 1.3-8.2], P = 2.1 × 10-4). This enrichment was further strengthened by (1) adding the recently reported TYK2 and TLR7 COVID-19 loci, particularly under a recessive model (OR = 19.65[95%CI 2.1-2635.4], P = 3.4 × 10-3), and (2) considering as pLOF branchpoint variants with potentially strong impacts on splicing among the 15 loci (OR = 4.40[9%CI 2.3-8.4], P = 7.7 × 10-8). Finally, the patients with pLOF/bLOF variants at these 15 loci were significantly younger (mean age [SD] = 43.3 [20.3] years) than the other patients (56.0 [17.3] years; P = 1.68 × 10-5).ConclusionsRare variants of TLR3- and TLR7-dependent type I IFN immunity genes can underlie life-threatening COVID-19, particularly with recessive inheritance, in patients under 60years old.

ObjectivePrior genomewide association studies have identified variation in major histocompatibility complex (MHC) class I alleles and C-C chemokine receptor type 5 gene (CCR5Δ32) as genetic predictors of viral control, especially in 'elite' controllers, individuals who remain virally suppressed in the absence of therapy.DesignCross-sectional genomewide association study.MethodsWe analyzed custom whole exome sequencing and direct human leukocyte antigen (HLA) typing from 202 antiretroviral therapy (ART)-suppressed HIV+ noncontrollers in relation to four measures of the peripheral CD4+ T-cell reservoir: HIV intact DNA, total (t)DNA, unspliced (us)RNA, and RNA/DNA. Linear mixed models were adjusted for potential covariates including age, sex, nadir CD4+ T-cell count, pre-ART HIV RNA, timing of ART initiation, and duration of ART suppression.ResultsPreviously reported 'protective' host genetic mutations related to viral setpoint (e.g. among elite controllers) were found to predict smaller HIV reservoir size. The HLA 'protective' B∗57:01 was associated with significantly lower HIV usRNA (q = 3.3 × 10-3), and among the largest subgroup, European ancestry individuals, the CCR5Δ32 deletion was associated with smaller HIV tDNA (P = 4.3 × 10-3) and usRNA (P = 8.7 × 10-3). In addition, genomewide analysis identified several single nucleotide polymorphisms in MX1 (an interferon stimulated gene) that were significantly associated with HIV tDNA (q = 0.02), and the direction of these associations paralleled MX1 gene eQTL expression.ConclusionsWe observed a significant association between previously reported 'protective' MHC class I alleles and CCR5Δ32 with the HIV reservoir size in noncontrollers. We also found a novel association between MX1 and HIV total DNA (in addition to other interferon signaling relevant genes, PPP1CB, DDX3X). These findings warrant further investigation in future validation studies.

Auto-antibodies (Abs) to type I interferons (IFNs) are found in up to 25% of patients with severe COVID-19, and are implicated in disease pathogenesis. It has remained unknown, however, whether type I IFN auto-Abs are unique to COVID-19, or are also found in other types of severe respiratory illnesses. To address this, we studied a prospective cohort of 284 adults with acute respiratory failure due to causes other than COVID-19. We measured type I IFN auto-Abs by radio ligand binding assay and screened for respiratory viruses using clinical PCR and metagenomic sequencing. Three patients (1.1%) tested positive for type I IFN auto-Abs, and each had a different underlying clinical presentation. Of the 35 patients found to have viral infections, only one patient tested positive for type I IFN auto-Abs. Together, our data suggest that type I IFN auto-Abs are uncommon in critically ill patients with acute respiratory failure due to causes other than COVID-19.

Funder: Charité - Universitätsmedizin Berlin, PURPOSE: Six to 19% of critically ill COVID-19 patients display circulating auto-antibodies against type I interferons (IFN-AABs). Here, we establish a clinically applicable strategy for early identification of IFN-AAB-positive patients for potential subsequent clinical interventions. METHODS: We analyzed sera of 430 COVID-19 patients from four hospitals for presence of IFN-AABs by ELISA. Binding specificity and neutralizing activity were evaluated via competition assay and virus-infection-based neutralization assay. We defined clinical parameters associated with IFN-AAB positivity. In a subgroup of critically ill patients, we analyzed effects of therapeutic plasma exchange (TPE) on the levels of IFN-AABs, SARS-CoV-2 antibodies and clinical outcome. RESULTS: The prevalence of neutralizing AABs to IFN-α and IFN-ω in COVID-19 patients from all cohorts was 4.2% (18/430), while being undetectable in an uninfected control cohort. Neutralizing IFN-AABs were detectable exclusively in critically affected (max. WHO score 6-8), predominantly male (83%) patients (7.6%, 18/237 for IFN-α-AABs and 4.6%, 11/237 for IFN-ω-AABs in 237 patients with critical COVID-19). IFN-AABs were present early post-symptom onset and at the peak of disease. Fever and oxygen requirement at hospital admission co-presented with neutralizing IFN-AAB positivity. IFN-AABs were associated with lower probability of survival (7.7% versus 80.9% in patients without IFN-AABs). TPE reduced levels of IFN-AABs in three of five patients and may increase survival of IFN-AAB-positive patients compared to those not undergoing TPE. CONCLUSION: IFN-AABs may serve as early biomarker for the development of severe COVID-19. We propose to implement routine screening of hospitalized COVID-19 patients for rapid identification of patients with IFN-AABs who most likely benefit from specific therapies.

Type I interferons (IFN), including IFNβ, play a protective role in multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). Type I IFNs are induced by the stimulation of innate signaling, including via cytoplasmic RIG-I-like receptors. In the present study, we investigated the potential effect of a chimeric protein containing the key domain of RIG-I signaling in the production of CNS endogenous IFNβ and asked whether this would exert a therapeutic effect against EAE. We intrathecally administered an adeno-associated virus vector (AAV) encoding a fusion protein comprising RIG-I 2CARD domains (C) and the first 200 amino acids of mitochondrial antiviral-signaling protein (MAVS) (M) (AAV-CM). In vivo imaging in IFNβ/luciferase reporter mice revealed that a single intrathecal injection of AAV-CM resulted in dose-dependent and sustained IFNβ expression within the CNS. IFNβ expression was significantly increased for 7 days. Immunofluorescent staining in IFNβ-YFP reporter mice revealed extraparenchymal CD45+ cells, choroid plexus, and astrocytes as sources of IFNβ. Moreover, intrathecal administration of AAV-CM at the onset of EAE induced the suppression of EAE, which was IFN-I-dependent. These findings suggest that accessing the signaling pathway downstream of RIG-I represents a promising therapeutic strategy for inflammatory CNS diseases, such as MS.

Kaposi’s sarcoma-associated herpesvirus (KSHV) is an oncogenic gammaherpesvirus that is the causative agent of primary effusion lymphoma and Kaposi’s sarcoma. In healthy carriers, KSHV remains latent, but a compromised immune system can lead to lytic viral replication that increases the probability of tumorigenesis. RIG-I-like receptors (RLRs) are members of the DExD/H box helicase family of RNA binding proteins that recognize KSHV to stimulate the immune system and prevent reactivation from latency. To determine if other DExD/H box helicases can affect KSHV lytic reactivation, we performed a knock-down screen that revealed DHX29-dependent activities appear to support viral replication but, in contrast, DDX24 and DDX49 have antiviral activity. When DDX24 or DDX49 are overexpressed in BCBL-1 cells, transcription of all lytic viral genes and genome replication were significantly reduced. RNA immunoprecipitation of tagged DDX24 and DDX49 followed by next-generation sequencing revealed that the helicases bind to mostly immediate-early and early KSHV mRNAs. Transfection of expression plasmids of candidate KSHV transcripts, identified from RNA pull-down, demonstrated that KSHV mRNAs stimulate type I interferon (alpha/beta) production and affect the expression of multiple interferon-stimulated genes. Our findings reveal that host DExD/H box helicases DDX24 and DDX49 recognize gammaherpesvirus transcripts and convey an antiviral effect in the context of lytic reactivation.

STAT2 is distinguished from other STAT family members by its exclusive involvement in type I and III interferon (IFN-I/III) signaling pathways, and its unique behavior as both positive and negative regulator of IFN-I signaling. The clinical relevance of these opposing STAT2 functions is exemplified by monogenic diseases of STAT2. Autosomal recessive STAT2 deficiency results in heightened susceptibility to severe and/or recurrent viral disease, whereas homozygous missense substitution of the STAT2-R148 residue is associated with severe type I interferonopathy due to loss of STAT2 negative regulation. Here we review the clinical presentation, pathogenesis, and management of these disorders of STAT2.

The type I interferon (IFN) response is one of the primary defense systems against various pathogens. Although rubella virus (RuV) infection is known to cause dysfunction of various organs and systems, including the central nervous system, little is known about how human neural cells evoke protective immunity against RuV infection, leading to controlling RuV replication. Using cultured human neural cells experimentally infected with RuV RA27/3 strain, we characterized the type I IFN immune response against the virus. RuV infected cultured human neural cell lines and induced IFN-β production, leading to the activation of signal transducer and activator of transcription 1 (STAT1) and the increased expression of IFN-stimulated genes (ISGs). Melanoma-differentiation-associated gene 5 (MDA5), one of the cytoplasmic retinoic acid-inducible gene I (RIG-I)-like receptors, is required for the RuV-triggered IFN-β mRNA induction in U373MG cells. We also showed that upregulation of RuV-triggered ISGs was attenuated by blocking IFN-α/β receptor subunit 2 (IFNAR2) using an IFNAR2-specific neutralizing antibody or by repressing mitochondrial antiviral signaling protein (MAVS) expression using MAVS-targeting short hairpin RNA (shRNA). Furthermore, treating RuV-infected cells with BX-795, a TANK-binding kinase 1 (TBK1)/I kappa B kinase ε (IKKε) inhibitor, robustly reduced STAT1 phosphorylation and expression of ISGs, enhancing viral gene expression and infectious virion production. Overall, our findings suggest that the RuV-triggered type I IFN-mediated antiviral response is essential in controlling RuV gene expression and viral replication in human neural cells.

Rift Valley fever (RVF) is a zoonotic and emerging disease, caused by the RVF virus (RVFV). In ruminants, it leads to “abortion storms” and enhanced mortality rates in young animals, whereas in humans it can cause symptoms like severe hemorrhagic fever or encephalitis. The role of the innate and adaptive immune response in disease initiation and progression is still poorly defined. The present study used the attenuated RVFV strain clone 13 to investigate viral spread, tissue tropism, and histopathological lesions after intranasal infection in C57BL/6 wild type (WT) and type I interferon (IFN-I) receptor I knockout (IFNAR−/−) mice. In WT mice, 104 PFU RVFV (high dose) resulted in a fatal encephalitis, but no hepatitis 7–11 days post infection (dpi), whereas 103 PFU RVFV (low dose) did not cause clinical disease or significant histopathological lesions in liver and the central nervous system (CNS). In contrast, IFNAR−/− mice infected with 103 PFU RVFV developed hepatocellular necrosis resulting in death at 2–5 dpi and lacked encephalitis. These results show that IFNAR signaling prevents systemic spread of the attenuated RVFV strain clone 13, but not the dissemination to the CNS and subsequent fatal disease. Consequently, neurotropic viruses may be able to evade antiviral IFN-I signaling pathways by using the transneuronal instead of the hematogenous route.

Zika virus (ZIKV) exhibits distinct selectivity for infection of various cells and tissues, but how host cellular factors modulate varying permissivity remains largely unknown. Previous studies showed that the neuroblastoma cell line SK-N-AS (expressing low levels of cellular protein CD24) was highly restricted for ZIKV infection, and that this restriction was relieved by ectopic expression of CD24. We tested the hypothesis that CD24 expression allowed ZIKV replication by suppression of the antiviral response. SK-N-AS cells expressing an empty vector (termed CD24-low cells) showed elevated basal levels of phosphorylated STAT1, IRF-1, IKKE, and NFκB. In response to exogenously added type I interferon (IFN-I), CD24-low cells had higher-level induction of antiviral genes and activity against two IFN-I-sensitive viruses (VSV and PIV5-P/V) compared to SK-N-AS cells with ectopic CD24 expression (termed CD24-high cells). Media-transfer experiments showed that the inherent antiviral state of CD24-low cells was not dependent on a secreted factor such as IFN-I. Transcriptomics analysis revealed that CD24 expression decreased expression of genes involved in intracellular antiviral pathways, including IFN-I, NFκB, and Ras. Our findings that CD24 expression in neuroblastoma cells represses intracellular antiviral pathways support the proposal that CD24 may represent a novel biomarker in cancer cells for susceptibility to oncolytic viruses.

Human platelet lysate (hPL) contains abundant growth factors for inducing human cell proliferation and may be a suitable alternative to fetal bovine serum (FBS) as a culture medium supplement. However, the application of hPL in virological research remains blank. Parechovirus type-A3 (PeV-A3) belongs to Picornaviridae, which causes meningoencephalitis in infants and young children. To understand the suitability of hPL-cultured cells for PeV-A3 infection, the infection of PeV-A3 in both FBS- and hPL-cultured glioblastoma (GBM) cells were compared. Results showed reduced PeV-A3 infection in hPL-cultured cells compared with FBS-maintained cells. Mechanistic analysis revealed hPL stimulating type I interferon (IFN) antiviral pathway, through which phospho-signal transducer and activator of transcription 1 (STAT1), STAT2, interferon regulatory factor 3 (IRF3) were activated and antiviral genes, such as IFN-α, IFN-β, and Myxovirus resistance protein 1 (MxA), were also detected. In addition, an enhanced PeV-A3 replication was detected in the hPL-cultured GBM cells treated with STAT-1 inhibitor (fludarabine) and STAT1 shRNA. These results in vitro suggested an unexpected effect of hPL-activated type I IFN pathway response to restrict virus replication and that hPL may be a potential antiviral bioreagent.

Lymphoid organ hypertrophy is a characteristic feature of acute infection and is considered to enable efficient induction of adaptive immune responses. Accordingly, oral infection with rotavirus induced a robust increase in cellularity in the mesenteric LNs, whose kinetics correlated with viral load and was caused by halted lymphocyte egress and increased recruitment of cells without altered cellular proliferation. Lymphocyte sequestration and mesenteric LN hypertrophy were independent of type 1 IFN receptor signaling or the continuous presence of TNF‐α. Our results support previous findings that adaptive immunity toward rotavirus is initiated primarily in the mesenteric LNs and show that type I IFN or TNF‐α are not required to coordinate the events involved in the LN response., Lymphoid organ hypertrophy facilitates lymphocyte priming in response to immune activation. We here report that mesenteric LN expansion in response to asymptomatic rotavirus infection in adult mice is independent of type I IFN and TNF‐α and caused both by enhanced recruitment and halted egress of lymphocytes.

Virus strains in the live attenuated influenza vaccine (LAIV) for swine in the United States that was on the market until 2020 encode a truncated nonstructural protein 1 of 126 amino acids (NS1del126). Their attenuation is believed to be due to an impaired ability to counteract the type I interferon (IFN)-mediated antiviral host response. However, this mechanism has been documented only in vitro for H3N2 strain A/swine/Texas/4199-2/98 NS1del126 (lvTX98), and several cases of clinical respiratory disease in the field were associated with the LAIV strains. We therefore further examined the pathobiology, including type I IFN induction, of lvTX98 in pigs and compared it with IFN induction in pig kidney-15 (PK-15) cells. lvTX98 induced up to 3-fold-higher type I IFN titers than wild-type TX98 (wtTX98) after inoculation of PK-15 cells at a high multiplicity of infection, while virus replication kinetics were similar. Mean nasal lvTX98 excretion by intranasally inoculated pigs was on average 50 times lower than that for wtTX98 but still reached titers of up to 4.3 log(10) 50% tissue culture infective doses/mL. After intratracheal inoculation, mean lvTX98 titers in the lower respiratory tract were significantly reduced at 18 to 48 h postinoculation (hpi) but similar to wtTX98 titers at 72 hpi. lvTX98 caused milder clinical signs than wtTX98 but induced comparable levels of microscopic and macroscopic lung lesions, peak neutrophil infiltration, and peak type I IFN. Thus, lvTX98 was partly attenuated in pigs, but this could not be associated with higher type I IFN levels. IMPORTANCE Swine influenza A viruses (swIAVs) with a truncated NS1del126 protein were strongly attenuated in previous laboratory-based safety studies and therefore approved for use as LAIVs for swine in the United States. In the field, however, the LAIV strains were detected in diagnostic samples and could regain a wild-type NS1 via reassortment with endemic swIAVs. This suggests a significant degree of LAIV replication and urges further investigation of the level and mechanism of attenuation of these LAIV strains in vivo. Here, we show that H3N2 LAIV strain lvTX98 is only partly attenuated in pigs and is excreted at significant titers after intranasal vaccination. Attenuation and restricted replication of lvTX98 in vivo seemed to be associated with the loss of NS1 functions other than type I IFN antagonism. Our findings can help to explain the occurrence of clinical respiratory disease and reassortment events associated with NS1del126-based LAIV strains in the field. Swine influenza A viruses (swIAVs) with a truncated NS1del126 protein were strongly attenuated in previous laboratory-based safety studies and therefore approved for use as LAIVs for swine in the United States. In the field, however, the LAIV strains were detected in diagnostic samples and could regain a wild-type NS1 via reassortment with endemic swIAVs.

Duck enteritis virus (DEV) can infect several types of waterfowl can cause high mortality and huge economic losses to the global waterfowl industry. Type I interferons (IFN) are important for host defense against virus infection through induction of antiviral effector molecules. TANK-binding kinase 1 (TBK1) is a key kinase required for the induction of type I IFNs; however, the role of TBK1 on DEV infection remains unclear. Here, we observed that the expression levels of TBK1 and IFN-β were upregulated during DEV infection in vivo and in vitro. Thus, the function of TBK1 on DEV infection was determined. The results showed that overexpression of TBK1 reduced DEV infection and knockdown of TBK1 resulted in the increased of DEV infection. Additionally, TBK1 overexpression upregulated the expression of IFN-β and a few interferon-stimulated genes (ISGs), which thus inhibited the synthesis of DEV glycoprotein B. On the other hand, the TBK1 inhibitor Amlexanox down-regulated the expression levels of IFN-β and IRF3. Interestingly, the expression levels of MAVS and GSK-3β were decreased in the cells treated with Amlexanox. Furthermore, overexpression of TBK1 activated the expression of upstream molecules MAVS and GSK-3β. Whereas, the expression of TBK1, IRF3 and IFN-β was inhibited by the GSK-3β inhibitor SB216763. Our findings suggest that DEV–stimulated TBK1 may be involved in defense against DEV infection.

We present a case of complete deficiency of the interferon alpha/beta receptor alpha chain (IFNAR1) in a child with fatal systemic hyperinflammation, apparently provoked by live-attenuated viral vaccination. Such pathologic hyperinflammation, fulfilling criteria for hemophagocytic lymphohistiocytosis, is an emerging phenotype accompanying inborn errors of type I interferon immunity.

Early type I interferon is essential for antagonizing against malaria infection, which remains a significant global infectious disease. After Plasmodium yoelii YM infection, the activation of MAVS‐, STING‐ and inflammasome‐IRF3‐mediated pathway could trigger the Socs1 expression to inhibit the TLR7‐MyD88‐IRF7‐induced type I interferon production. However, the dynamic regulatory mechanisms of type I interferon response to YM infection and delicate cross‐regulation of these signalling are far from clear. In current study, we established a mathematical model to systematically demonstrate that the MAVS‐, STING‐ and inflammasome‐mediated signalling pathways play distinct roles in regulating type I interferon response after YM infection; and the YM dose could significantly affect the difference of resistance to YM infection among MAVS, STING and inflammasome deficiency. Collectively, our study systematically elucidated the precise regulatory mechanisms of type I interferon signalling after YM infection and advanced the research on therapy of plasmodium infection by incorporating multiple signalling pathways at diverse time.

Objective In mouse models, deficiency of TTC39B (T39) decreases hepatic lipogenic gene expression and protects against diet-induced steatohepatitis. While assessing the therapeutic potential of antisense oligonucleotides (ASOs) targeting T39, we discovered an unexpected weight loss phenotype. The objective of this study was to determine the mechanism of the resistance to diet-induced obesity. Methods To assess therapeutic potential, we used antisense oligonucleotides (ASO) to knock down T39 expression in a Western or high-fat, high-cholesterol, high-sucrose-diet-fed Ldlr−/− or wild-type mice. Results T39 ASO treatment led to decreased hepatic lipogenic gene expression and decreased hepatic triglycerides. Unexpectedly, T39 ASO treatment protected against diet-induced obesity. The reduced weight gain was seen with two different ASOs that decreased T39 mRNA in adipose tissue macrophages (ATMs), but not with a liver-targeted GalNac-ASO. Mice treated with the T39 ASO displayed increased browning of gonadal white adipose tissue (gWAT) and evidence of increased lipolysis. However, T39 knockout mice displayed a similar weight loss response when treated with T39 ASO, indicating an off-target effect. RNA-seq analysis of gWAT showed a widespread increase in type I interferon (IFN)-responsive genes, and knockout of the IFN receptor abolished the weight loss phenotype induced by the T39 ASO. Some human T39 ASOs and ASOs with different modifications targeting LDLR also induced a type I IFN response in THP1 macrophages. Conclusion Our data suggest that extrahepatic targeting of T39 by ASOs in ATMs produced an off-target type 1 IFN response, leading to activation of lipolysis, browning of WAT, and weight loss. While our findings suggest that ASOs may induce off-target type 1 IFN response more commonly than previously thought, they also suggest that therapeutic induction of type 1 IFN selectively in ATMs could potentially represent a novel approach to the treatment of obesity., Highlights • T39 ASOs produced an off-target type I IFN response resulting in reduced obesity, increased lipolysis and browning of WAT. • The resistance to diet-induced obesity and browning of white adipose tissue was dependent on IFNAR1. • Multiple ASOs produced a type I IFN response in vitro in human THP1 cells, suggesting they may be more common than suspected. • Therapeutic induction of low-grade type 1 IFN responses by ASO targeting to ATMs may represent a novel treatment for obesity.

Nucleic acids derived from microorganisms are powerful triggers for innate immune responses. Proteins called RNA and DNA sensors detect foreign nucleic acids and, in mammalian cells, include RIG-I, cGAS, and AIM2. On binding to nucleic acids, these proteins initiate signaling cascades that activate host defense responses. An important aspect of this defense program is the production of cytokines such as type I interferons and IL-1β. Studies conducted over recent years have revealed that nucleic acid sensors also activate programmed cell death pathways as an innate immune response to infection. Indeed, RNA and DNA sensors induce apoptosis, pyroptosis, and necroptosis. Cell death via these pathways prevents replication of pathogens by eliminating the infected cell and additionally contributes to the release of cytokines and inflammatory mediators. Interestingly, recent evidence suggests that programmed cell death triggered by nucleic acid sensors plays an important role in a number of noninfectious pathologies. In addition to nonself DNA and RNA from microorganisms, nucleic acid sensors also recognize endogenous nucleic acids, for example when cells are damaged by genotoxic agents and in certain autoinflammatory diseases. This review article summarizes current knowledge on the links between nucleic acid sensing and cell death and explores important open questions for future studies in this area., Graphical abstract Image 1, Highlights • Nucleic acid sensors detect foreign or unusual DNA and RNA. • DNA/RNA sensing triggers innate immune responses, including programmed cell death. • Nucleic acid sensors induce apoptosis, necroptosis, and pyroptosis. • Programmed cell death eliminates infected cells and contributes to inflammation. • These responses are highly cell type and species specific.

Children generally develop a mild disease after SARS-CoV-2 infection whereas older adults are at risk of developing severe COVID-19. Recent transcriptomic analysis showed pre-activated innate immunity in children, resulting in a more effective anti-SARS-CoV-2 response upon infection. To further characterize age-related differences, we studied type I and III interferon (IFN) response in SARS-CoV-2 infected and non-infected individuals of different ages. Specifically, levels of expression of type I (IFN-α, -β, -ε and -ω), type III (IFN-λ1, -λ2 and -λ3) IFNs and of the IFN-stimulated genes, ISG15 and ISG56 were quantified in nasopharyngeal cells from diagnostic swabs. Basal transcription of type I/III IFN genes was highest among children and decreased with age. Among SARS-CoV-2-infected individuals, only IFN-ε and -ω levels were significantly higher in children and young adults whereas ISGs were overexpressed in infected adults. The occurrence of symptoms in children and the need for hospitalization in adults were associated to higher transcription of several IFN genes. Starting from a pre-activated transcription level, the expression of type I and III IFNs was not highly up-regulated in children upon SARS-CoV-2 infection; young adults activated IFNs’ transcription at intermediate levels whereas older adults were characterized by higher ISGs and lower IFN-ε and -ω relative expression levels. Overall, our findings contribute to recognize components of a protective IFN response as a function of age, in the context of SARS-CoV-2 infection.

BackgroundThe role of type I interferons (IFNs) in the early phase of COVID-19 remains unclear.ObjectivesTo evaluate the relationship between IFN-I levels in patients with COVID-19 and clinical presentation, SARS-CoV-2 viral load, and other major pro-inflammatory cytokines.MethodsThis prospective observational study recruited patients hospitalized with COVID-19. The levels of interferon-alpha (IFN-α), interferon-beta (IFN-β), interleukin-6 (IL-6), and C-X-C motif chemokine ligand (CXCL10) within 5 days after symptom onset were measured using an ELISA, in serum from blood collected within 5 days after the onset of symptoms. The SARS-CoV-2 viral load was determined via qPCR using nasal-swab specimens and serum.ResultsThe study enrolled 50 patients with COVID-19. IFN-α levels were significantly higher in patients who presented with pneumonia or developed hypoxemic respiratory failure (p < 0.001). Furthermore, IFN-α levels were associated with viral load in nasal-swab specimens and RNAemia (p < 0.05). In contrast, there was no significant association between IFN-β levels and the presence of pneumonia or RNAemia, despite showing a stronger association with nasal-swab viral load (p < 0.001). Correlation analysis showed that the serum levels of IFN-α significantly correlated with those of IFN-β, IL-6, and CXCL10, while the levels of IFN-β did not correlate with those of IL-6 or CXCL10.ConclusionsSerum IFN-I levels in the early phase of SARS-CoV-2 infection were higher in patients who developed hypoxemic respiratory failure. The association between IFN-α, IL-6, and CXCL10 may reflect the systemic immune response against SARS-CoV-2 invasion into pulmonary circulation, which might be an early predictor of respiratory failure due to COVID-19.

Type I interferons (IFNs) are cytokines with both antiviral properties and protective roles in innate immune responses to viral infection. They induce an antiviral cellular state and link innate and adaptive immune responses. Yet, viruses have evolved different strategies to inhibit such host responses. One of them is the existence of viral proteins which subvert type I IFN responses to allow quick and successful viral replication, thus, sustaining the infection within a host. We propose mathematical models to characterise the intra-cellular mechanisms involved in viral protein antagonism of type I IFN responses, and compare three different molecular inhibition strategies. We study the Ebola viral protein, VP35, with this mathematical approach. Approximate Bayesian computation sequential Monte Carlo, together with experimental data and the mathematical models proposed, are used to perform model calibration, as well as model selection of the different hypotheses considered. Finally, we assess if model parameters are identifiable and discuss how such identifiability can be improved with new experimental data.

Dengue Virus (DENV) infection can cause severe illness such as highly fatality dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Innate immune activation by Nod-like receptors (NLRs) is a critical part of host defense against viral infection. Here, we revealed a key mechanism of NLRP12-mediated regulation in DENV infection. Firstly, NLRP12 expression was inhibited in human macrophage following DENV or other flaviviruses (JEV, YFV, ZIKV) infection. Positive regulatory domain 1 (PRDM1) was induced by DENV or poly(I:C) and suppressed NLRP12 expression, which was dependent on TBK-1/IRF3 and NF-κB signaling pathways. Moreover, NLRP12 inhibited DENV and other flaviviruses (JEV, YFV, ZIKV) replication, which relied on the well-conserved nucleotide binding structures of its NACHT domain. Furthermore, NLRP12 could interact with heat shock protein 90 (HSP90) dependent on its Walker A and Walker B sites. In addition, NLRP12 enhanced the production of type I IFNs (IFN-α/β) and interferon-stimulated genes (ISGs), including IFITM3, TRAIL and Viperin. Inhibition of HSP90 with 17-DMAG impaired the upregulation of type I IFNs and ISGs induced by NLRP12. Taken together, we demonstrated a novel mechanism that NLRP12 exerted anti-viral properties in DENV and other flaviviruses (JEV, YFV, ZIKV) infection, which brings up a potential target for the treatment of DENV infection.

BackgroundCOVID-19, caused by SARS-CoV-2 virus, is a global pandemic with high mortality and morbidity. Limited diagnostic methods hampered the infection control. Since the direct detection of virus mainly by RT-PCR may cause false-negative outcome, host response-dependent testing may serve as a complementary approach for improving COVID-19 diagnosis.ObjectiveOur study discovered a highly-preserved transcriptional profile of Type I interferon (IFN-I)-dependent genes for COVID-19 complementary diagnosis.MethodsComputational language R-dependent machine learning was adopted for mining highly-conserved transcriptional profile (RNA-sequencing) across heterogeneous samples infected by SARS-CoV-2 and other respiratory infections. The transcriptomics/high-throughput sequencing data were retrieved from NCBI-GEO datasets (GSE32155, GSE147507, GSE150316, GSE162835, GSE163151, GSE171668, GSE182569). Mathematical approaches for homological analysis were as follows: adjusted rand index-related similarity analysis, geometric and multi-dimensional data interpretation, UpsetR, t-distributed Stochastic Neighbor Embedding (t-SNE), and Weighted Gene Co-expression Network Analysis (WGCNA). Besides, Interferome Database was used for predicting the transcriptional factors possessing IFN-I promoter-binding sites to the key IFN-I genes for COVID-19 diagnosis.ResultsIn this study, we identified a highly-preserved gene module between SARS-CoV-2 infected nasal swab and postmortem lung tissue regulating IFN-I signaling for COVID-19 complementary diagnosis, in which the following 14 IFN-I-stimulated genes are highly-conserved, including BST2, IFIT1, IFIT2, IFIT3, IFITM1, ISG15, MX1, MX2, OAS1, OAS2, OAS3, OASL, RSAD2, and STAT1. The stratified severity of COVID-19 may also be identified by the transcriptional level of these 14 IFN-I genes.ConclusionUsing transcriptional and computational analysis on RNA-seq data retrieved from NCBI-GEO, we identified a highly-preserved 14-gene transcriptional profile regulating IFN-I signaling in nasal swab and postmortem lung tissue infected by SARS-CoV-2. Such a conserved biosignature involved in IFN-I-related host response may be leveraged for COVID-19 diagnosis.

BackgroundHost factors that influence Congenital Zika Syndrome (CZS) outcome remain elusive. Interferons have been reported as the main antiviral factor in Zika and other flavivirus infections.MethodsWe accessed samples from Zika pregnancies, conducted a case-control study to verify whether interferon alfa receptor 1 (IFNAR1) and interferon lambda 2 and 4 (IFNL2/4) single nucleotide polymorphisms (SNPs) contribute to CZS newborn outcome and we characterized placenta gene expression profile at term.FindingsNewborns carrying CG/CC genotypes of rs2257167 inIFNAR1presented higher risk of developing CZS (OR=3.73; IC=1.36-10.21;Pcorrected=0.02646). No association betweenIFNLSNPs and CZS was observed. Placenta from CZS cases displayed lower levels ofIFNL2andISG15along with higherIFIT5.The rs2257167 CG/CC placentas also demonstrated high levels ofIFIT5and inflammation-related genes.InterpretationWe found CZS to be related with exacerbated type I IFN and insufficient type III IFN in placenta at term, forming an unbalanced response modulated by theIFNAR1rs2257167 genotype. These findings shed light on the host-pathogen interaction focusing on the genetically regulated type I / type III IFN axis that could lead to better management of Zika and other TORCH (Toxoplasma, Others, Rubella, Cytomegalovirus, Herpes) congenital infections.FundingThis work was supported by the Instituto Oswaldo Cruz (Rio de Janeiro, Brazil) and by the Instituto de Tecnologia em Imunobiológicos (Rio de Janeiro, Brazil).Research in contextEvidence before this studyLevels of type I and type III interferons are genetically controlled and decisively regulate outcome of spontaneous viral infections or response to antiviral treatment. Hepatitis C virus, Yellow Fever and Zika virus belong to the Flaviviridae family and elicit similar host immune responses. Congenital Zika Syndrome presents well-known risk factors, mainly the first trimester of pregnancy as well as social and nutritional factors, however, these do not entirely explain abnormal outcomes.Added value of this studyWe conducted a case-control study to evaluate SNPs in type I and III interferon genes using samples from newborns and mothers who had zika infection during pregnancy. We have shown that newborn interferon type I background contributes to the development of abnormal CSZ. This specific genetic makeup regulates placental immunological responses and prevents an exacerbated type I, and lack of type III, interferon response in syndromic cases.Implications of all the available evidenceOur study suggests an important factor regulating the host-pathogen interaction during Zika virus (ZIKV) infections in humans. During pregnancy, genetic variations play a role in balancing tissue-specific type I and III interferons during ZIKV congenital infection influencing fetal neurological damage. Custom pharmacological interventions could be used to modulate immunity and inflammation towards protective responses.Graphical abstract

MicroRNAs are involved in the pathogenesis of systemic lupus erythematosus (SLE) and dysregulated in the kidneys of lupus nephritis (LN) patients, but their pathogenic roles in LN are largely unknown. Janus Kinase 1 (JAK1) mediates the activation of the downstream signaling pathways of many inflammatory cytokines, including type I interferon (IFN-I) signaling pathway which is critical to the development of SLE and LN. Thus, JAK1 is a potent therapeutic target for these autoimmune diseases. However, there are few studies demonstrating the dysregulation of JAK1 in autoimmune diseases and the molecular mechanism behind it. In this concise report, we show an inhibitory effect of hsa-miR-127-3p, a microRNA that is downregulated in the renal tissues of LN patients, on IFN-I signaling. We found the overexpression of hsa-miR-127-3p could inhibit the induction of ISRE and GAS mediated gene expression, the phosphorylation of STAT1 and STAT2, and the upregulation of IFN stimulated genes (ISGs) stimulated by IFN-I. While, hsa-miR-127-3p antagonist enhanced the activation of IFN-I signaling in primary renal mesangial cells. Subsequently, we identified JAK1 as a bona fide target gene of hsa-miR-127-3p and showed hsa-miR-127-3p targeted JAK1 through binding to its 3’UTR and coding region. Consistently, we found the downregulation of hsa-miR-127-3p was associated with the upregulation of JAK1 and ISGs in kidney tissues of LN patients. Our data indicate renal downregulation of hsa-miR-127-3p contributes to the overactivation of IFN-I signaling pathway in the kidneys of LN patients through promoting the expression of JAK1, suggesting hsa-miR-127-3p mimics may be used to inhibit JAK1 and IFN-I signaling pathway in LN.

Porcine reproductive and respiratory syndrome (PRRS), a significant viral infectious disease that commonly occurs among farmed pigs, leads to considerable economic losses to the swine industry worldwide. Major vault protein (MVP) is a host factor that induces type Ⅰ interferon (IFN) production. In this study, we evaluated the effect of MVP on PRRSV infection in CRL2843CD163 cell lines and porcine alveolar macrophages (PAMs). Our results showed that MVP expression was downregulated by PRRSV infection. Adenoviral overexpression of MVP inhibited PRRSV replication, whereas the siRNA knockdown of MVP promoted PRRSV replication. In addition, MVP knockdown has an adverse effect on the inhibitive role of MVP overexpression on PRRSV replication. Moreover, MVP could induce the expression of type Ⅰ IFNs and IFN-stimulated gene 15 (ISG15) in PRRSV-infected PAMs. Based on these results, MVP may be a potential molecular target of drugs for the effective prevention and treatment of PRRSV infection.

Oncolytic viruses are promising agents for cancer therapy because they selectively infect and kill tumor cells, and because they trigger immune responses that can boost anticancer immunity. Key to the latter process is the production of type I interferons (IFN-Is) that can turn noninflamed "cold" tumors into "hot" ones. Besides this desired anticancer effect, IFN-Is are antiviral and successful oncolytic virotherapy thus relies on tightly controlled IFN-I levels. This requires a profound understanding of when and how tumor cells induce IFN-I in response to specific viruses. In this study, we uncovered two key factors that augment IFN-I production in transformed human myeloid cells infected with a tumor-selective reovirus. Viral replication and IFN-alpha/beta receptor (IFNAR) signaling progressively reinforced the levels of IFN-I expressed by infected cells. Mechanistically, both augmented the activation of interferon regulatory factor 3, a key transcription factor for IFN beta expression. Our findings imply that reovirus-permissive tumor cells themselves are a major source of IFN-I expression. As tumors can perturb the IFNAR pathway for their own survival, reovirus-exposed IFNAR-unresponsive tumors may need additional therapeutic intervention to promote the secretion of sufficient IFN-I into the tumor microenvironment. Our increased understanding of the parameters that affect reovirus-induced IFN-I levels could aid in the design of tailored virus-based cancer therapies.

Type I interferon and apoptosis elicit multifaceted effects on host defense and various diseases, such as viral infections and cancers. However, the gene/protein network regulating type I interferon and apoptosis has not been elucidated completely. In this study, we selected grass carp (Ctenopharyngodon idella) as an experimental model to investigate the modulation of RNASEK on the secretion of type I interferon and apoptosis. We first cloned two paralogs RNASEK-a and -b in grass carp, defined three exons in each gene, and found the length of both coding regions is 306 bp with 73.27% of protein homology. The protein sequences of the two paralogs are highly conserved across species. Two proteins were mainly localized in early and late endosomes and endoplasmic reticulum. Further, quantitative real-time PCR demonstrated that dsRNA poly I:C and grass carp reovirus upregulated RNASEK-a and -b in grass carp cells and tissues. Overexpression of RNASEK-a and -b individually induced type I interferon expression and the phosphorylation of IRF3/IRF7 shown by Western blot and immunofluorescent staining, increased Bax/Bcl-2 mRNA ratio, DNA fragmentations, TUNEL-positive cells, and the proportion of Annexin V-positive signals in flow cytometry, and activated eIF2α, opposite to that observed when RNASEK-a and -b were knocked down in multiple cell types. Taken together, we claim for the first time that fish paralog proteins RNASEK-a and -b enhance type I interferon secretion and promote apoptosis, which may be involved in the phosphorylation of IRF3/IRF7 and eIF2α, respectively. Our study reveals a previously unrecognized role of RNASEK as a new positive regulator of type I interferon and apoptosis.

Acute lung injury (ALI) results in acute respiratory disease that causes fatal respiratory diseases; however, little is known about the incidence of influenza infection in ALI. Using a ALI-mouse model, we investigated the pro-inflammatory cytokine response to ALI and influenza infection. Mice treated with bleomycin (BLM), which induces ALI, were more resistant to influenza virus infection and exhibited higher levels of type I interferon (IFN-I) transcription during the early infection period than that in PBS-treated control mice. BLM-treated mice also exhibited a lower viral burden, reduced pro-inflammatory cytokine production, and neutrophil levels. In contrast, BLM-treated IFN-I receptor 1 (IFNAR1)-knockout mice failed to show this attenuated phenotype, indicating that IFN-I is key to the antiviral response in ALI-induced mice. The STING/TBK1/IRF3 pathway was found to be involved in IFN-I production and the establishment of an antiviral environment in the lung. The depletion of plasmacytoid dendritic cells (pDCs) reduced the effect of BLM treatment against influenza virus infection, suggesting that pDCs are the major source of IFN-I and are crucial for defense against viral infection in BLM-induced lung injury. Overall, this study showed that BLM-mediated ALI in mice induced the release of double-stranded DNA, which in turn potentiated IFN-I-dependent pulmonary viral resistance by activating the STING/TBK1/IRF3 pathway in association with pDCs.

Viral infections pose a severe threat to humans by causing many infectious, even fatal, diseases, such as the current pandemic disease (COVID-19) since 2019, and understanding how the host innate immune system recognizes viruses has become more important. Endosomal and cytosolic sensors can detect viral nucleic acids to induce type I interferon and proinflammatory cytokines, subsequently inducing interferon-stimulated genes for restricting viral infection. Although viral RNA and DNA sensing generally rely on diverse receptors and adaptors, the crosstalk between DNA and RNA sensing is gradually appreciated. This minireview highlights the overlap between the RNA- and DNA-sensing mechanisms in antiviral innate immunity, which significantly amplifies the antiviral innate responses to restrict viral infection and might be a potential novel target for preventing and treating viral diseases.

Intestinal microbiomes are of vital importance in antagonizing systemic viral infection. However, very little literature has shown whether commensal bacteria play a crucial role in protecting against enteric virus systemic infection from the aspect of modulating host innate immunity. In the present study, we utilized an enteric virus, encephalomyocarditis virus (EMCV), to inoculate mice treated with phosphate-buffered saline (PBS) or given an antibiotic cocktail (Abx) orally or intraperitoneally to examine the impact of microbiota depletion on virulence and viral replication in vivo. Microbiota depletion exacerbated the mortality, neuropathogenesis, viremia, and viral burden in brains following EMCV infection. Furthermore, Abx-treated mice exhibited severely diminished mononuclear phagocyte activation and impaired type I interferon (IFN) production and expression of IFN-stimulated genes (ISG) in peripheral blood mononuclear cells (PBMC), spleens, and brains. With the help of fecal bacterial 16S rRNA sequencing of PBS- and Abx-treated mice, we identified a single commensal bacterium, Blautia coccoides, that can restore mononuclear phagocyte- and IFNAR (IFN-α/β receptor)-dependent type I IFN responses to restrict systemic enteric virus infection. These findings may provide insight into the development of novel therapeutics for preventing enteric virus infection or possibly alleviating clinical diseases by activating host systemic innate immune responses via respective probiotic treatment using B. coccoides. IMPORTANCE While cumulative data indicate that indigenous commensal bacteria can facilitate enteric virus infection, little is known regarding whether intestinal microbes have a protective role in antagonizing enteric systemic infection by modulating host innate immunity. Although accumulating literature has pointed out that the microbiota has a fundamental impact on host systemic antiviral innate immune responses mediated by type I interferon (IFN), only a few specific commensal bacteria species have been revealed to be capable of regulating IFN-I and ISG expression, not to mention the underlying mechanisms. Thus, it is important to understand the cross talk between microbiota and host anti-enteric virus innate immune responses and characterize the specific bacterial species that possess protective functions. Our study demonstrates how fundamental innate immune mediators such as mononuclear phagocytes and type I IFN are regulated by commensal bacteria to antagonize enteric virus systemic infection. In particular, we have identified a novel commensal bacterium, Blautia coccoides, that can restrict enteric virus replication and neuropathogenesis by activating IFN-I and ISG responses in mononuclear phagocytes via an IFNAR- and STAT1-mediated signaling pathway.

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. TheSuper susceptibility to tuberculosis 1 (Sst1)locus in mice confers resistance to diverse bacterial infections. Here we provide evidence thatSp140is a gene encoded within theSst1locus that represses type I IFN transcription during bacterial infections. We generatedSp140-/-mice and find they are susceptible to infection byLegionella pneumophilaandMycobacterium tuberculosis.Susceptibility ofSp140-/-mice to bacterial infection was rescued by crosses to mice lacking the type I IFN receptor(Ifnar-/-). Our results implicateSp140as an important negative regulator of type I IFNs that is essential for resistance to bacterial infections.Impact StatementRepression of type I interferons by SP140 is essential for resistance toLegionella pneumophilaandMycobacterium tuberculosis.

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 β-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.

Significance Clinical studies have shown that defects in type I interferon (IFN) production or antibodies to IFN appear to correlate with severe COVID-19 infection. Here, we demonstrate that proinflammatory cytokines but not type I IFN are produced by hypoxic human monocytes. We show that hypoxia suppresses production of type I IFN but not nuclear factor-κB–dependent proinflammatory cytokines. We demonstrate that hypoxia-inducible factor-1α is a direct transcriptional suppressor of interferon regulatory factors, the transcriptional activators of type I IFN. These findings may aid in understanding and control of impaired IFN production by severe acute respiratory syndrome coronavirus 2 infection., Patients with severe COVID-19 infection exhibit a low level of oxygen in affected tissue and blood. To understand the pathophysiology of COVID-19 infection, it is therefore necessary to understand cell function during hypoxia. We investigated aspects of human monocyte activation under hypoxic conditions. HMGB1 is an alarmin released by stressed cells. Under normoxic conditions, HMGB1 activates interferon regulatory factor (IRF)5 and nuclear factor-κB in monocytes, leading to expression of type I interferon (IFN) and inflammatory cytokines including tumor necrosis factor α, and interleukin 1β, respectively. When hypoxic monocytes are activated by HMGB1, they produce proinflammatory cytokines but fail to produce type I IFN. Hypoxia-inducible factor-1α, induced by hypoxia, functions as a direct transcriptional repressor of IRF5 and IRF3. As hypoxia is a stressor that induces secretion of HMGB1 by epithelial cells, hypoxia establishes a microenvironment that favors monocyte production of inflammatory cytokines but not IFN. These findings have implications for the pathogenesis of COVID-19.

RIG-I-like receptors (RLR), RIG-I and MDA5, are cytoplasmic viral RNA sensors that recognize viral double-stranded RNAs and trigger signals to induce antiviral responses, including type I interferon production. Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) caused the coronavirus disease 2019 pandemic. However, the RLR role in innate immune response to SARS-CoV-2 has not been fully elucidated. Here, we studied the roles of RLR in cytokine expression responding to SARS-CoV-2 and found that not only MDA5 but also RIG-I are involved in innate immune responses in some types of human cells. Transfection of total RNAs extracted from SARS-CoV-2-infected cells into epithelial cells induced IFN-β, IP-10, and Ccl5 mRNA expression. The cytokine expression was reduced by knockout of either RIG-I or MDA5, suggesting that both proteins are required for appropriate innate immune response to SARS-CoV-2. Two viral genomic RNA regions strongly induced type I IFN expression, and a 200-base fragment of viral RNA preferentially induced type I IFN in a RIG-I-dependent manner. In contrast, SARS-CoV-2 infectious particles hardly induced cytokine expression, suggesting viral escape from the host response. Viral 9b protein inhibited RIG-I and MAVS interaction, and viral 7a protein destabilized the TBK1 protein, leading to attenuated IRF-3 phosphorylation required for type I IFN expression. Our data elucidated the mechanism underlying RLR-mediated response to SARS-CoV-2 infection and viral escape from the host innate immune response.

Type I IFN (IFN-I) is thought to be rapidly internalized and degraded following binding to its receptor and initiation of signaling. However, many studies report the persistent effects mediated by IFN-I for days or even weeks, both ex vivo and in vivo. These long-lasting effects are attributed to downstream signaling molecules or induced effectors having a long half-life, particularly in specific cell types. Here, we describe a mechanism explaining the long-term effects of IFN-I. Following receptor binding, IFN-I is siloed into endosomal compartments. These intracellular “IFN silos” persist for days and can be visualized by fluorescence and electron microscopy. However, they are largely dormant functionally, due to IFN-I−induced negative regulators. By contrast, in individuals lacking these negative regulators, such as ISG15 or USP18, this siloed IFN-I can continue to signal from within the endosome. This mechanism may underlie the long-term effects of IFN-I therapy and may contribute to the pathophysiology of type I interferonopathies.

Prion diseases, including human Creutzfeldt-Jakob disease, are infectious, intractable central neurodegenerative diseases, which are also zoonoses that commonly infect not only higher organisms but also a wide variety of animals.Pathogenic prions induce abnormal prion protein (PrP), which is produced by structural conversion of normal PrP, a beta-sheet-rich structure with high aggregation propensity. Thus, it is believed that the host is immunotolerant against prions because there is no difference in the primary structure of normal and abnormal PrP, and prions do not induce a marked immune response. Recently, using mutated Toll-like receptor (TLR) 4-transgenic mice, a bioassay after prion inoculation has intriguingly found that the TLR4-signaling pathway may have a protective role against prion infection. Meanwhile, we reported that a transcription factor, interferon regulatory factor-3 (IRF-3), located downstream of TLR4 signaling, showed resistance to prions. IRF-3-inducing type I interferon (I-IFN) is a critical factor for the host defense against pathogen invasion. These findings indicate that the TLR-signaling pathway of the innate immune system might regulate prion invasion. However, the details have not been fully determined. In this symposium, we will introduce new findings including the relationship between I-IFN and prions., Journal of the Pharmaceutical Society of Japan, 139(7), pp.993-998; 2019

Background Type I interferons (IFN-I) have recently emerged as key regulators of tumor response to chemotherapy and immunotherapy. However, IFN-I function in cytotoxic T lymphocytes (CTLs) in the tumor microenvironment is largely unknown. Methods Tumor tissues and CTLs of human colorectal cancer patients were analyzed for interferon (alpha and beta) receptor 1 (IFNAR1) expression. IFNAR1 knock out (IFNAR-KO), mixed wild type (WT) and IFNAR1-KO bone marrow chimera mice, and mice with IFNAR1 deficiency only in T cells (IFNAR1-TKO) were used to determine IFN-I function in T cells in tumor suppression. IFN-I target genes in tumor-infiltrating and antigen-specific CTLs were identified and functionally analyzed. Results IFNAR1 expression level is significantly lower in human colorectal carcinoma tissue than in normal colon tissue. IFNAR1 protein is also significantly lower on CTLs from colorectal cancer patients than those from healthy donors. Although IFNAR1-KO mice exhibited increased susceptibility to methylcholanthrene-induced sarcoma, IFNAR1-sufficient tumors also grow significantly faster in IFNAR1-KO mice and in mice with IFNAR1 deficiency only in T cells (IFNAR1-TKO), suggesting that IFN-I functions in T cells to enhance host cancer immunosurveillance. Strikingly, tumor-infiltrating CTL levels are similar between tumor-bearing WT and IFNAR1-KO mice. Competitive reconstitution of mixed WT and IFNAR1-KO bone marrow chimera mice further determined that IFNAR1-deficient naïve CTLs exhibit no deficiency in response to vaccination to generate antigen-specific CTLs as compared to WT CTLs. Gene expression profiling determined that Gzmb expression is down-regulated in tumor-infiltrating CTLs of IFNAR1-KO mice as compared to WT mice, and in antigen-specific IFNAR1-KO CTLs as compared to WT CTLs in vivo. Mechanistically, we determined that IFN-I activates STAT3 that binds to the Gzmb promoter to activate Gzmb transcription in CTLs. Conclusion IFN-I induces STAT3 activation to activate Gzmb expression to enhance CTL effector function to suppress tumor development. Human colorectal carcinoma may use down-regulation of IFNAR1 on CTLs to suppress CTL effector function to evade host cancer immunosurveillance. Electronic supplementary material The online version of this article (10.1186/s40425-019-0635-8) contains supplementary material, which is available to authorized users.

MicroRNAs (miRNAs) are a type of small non-coding RNAs that participate in diverse cellular processes including microbial invasion and immune defense. In a previous study, we identified a large amount of Japanese flounder (Paralichthys olivaceus) miRNAs responsive to megalocytivirus infection. In the present study, we examined the function of one of these miRNAs, pol-miR-194a, in association with the infectivity of Edwardsiella tarda, an intracellular bacterial pathogen to many fish species including flounder. We found that pol-miR-194a was induced in expression to a significant extent in the spleen, liver, and gill of Japanese flounder infected by E. tarda. Transfection of flounder cells with pol-miR-194a mimic significantly enhanced the intracellular replication of E. tarda. pol-miR-194a was able to interact specifically with the 3′UTR of IRF7 in a negative manner, resulting in inhibition of IRF7 expression. Consistently, pol-miR-194a significantly blocked the promoter activity of type Ⅰ interferon. Taken together, these results indicate that pol-miR-194a plays an important role in the regulation of flounder immune response as well as microbial infection, and that pol-miR-194a probably serves as a target for E. tarda to manipulate and escape host immune defense.

The effects of casirivimab and imdevimab (C/I) on the innate immune response against SARS-CoV-2 infection remain unclear. We evaluated the effect of C/I on type I interferon (IFN-I) and cytokines in patients with SARS-CoV-2 infection. This prospective observational study recruited consecutive patients hospitalized with SARS-CoV-2 infection. Blood levels of IFN-I and cytokines before and after C/I administration were assessed using enzyme-linked immunoassay. The study enrolled 29 patients in the C/I group. In addition, 11 patients who received remdesivir and dexamethasone (R/D group) during the early phase (≤5 days after the onset of symptoms) were included as a comparator group. After treatment, IFN-α and IFN-β levels decreased significantly in both the C/I group and R/D group, whilst the post-treatment neutrophil-to-lymphoid ratio increased in the early C/I group but not the R/D group. In the C/I group, temporal temperature elevation and hypoxemia were observed after treatment in 58.6% and 41.4% of the cohort, respectively. However, most patients recovered by 5 days after treatment. This study could demonstrate the high therapeutic effect of C/I with an antibody-dependent enhancement-like response and decreased IFN-I production, which was likely due to the immediate induction of an antibody-dependent immune response against SARS-CoV-2.

Purpose Singleton-Merten syndrome manifests as dental dysplasia, glaucoma, psoriasis, aortic calcification, and skeletal abnormalities including tendon rupture and arthropathy. Pathogenic variants in IFIH1 have previously been associated with the classic Singleton-Merten syndrome, while variants in DDX58 has been described in association with a milder phenotype, which is suggested to have a better prognosis. We studied a family with severe, “classic” Singleton-Merten syndrome. Methods We undertook clinical phenotyping, next-generation sequencing, and functional studies of type I interferon production in patient whole blood and assessed the type I interferon promoter activity in HEK293 cells transfected with wild-type or mutant DDX58 stimulated with Poly I:C. Results We demonstrate a DDX58 autosomal dominant gain-of-function mutation, with constitutive upregulation of type I interferon. Conclusions DDX58 mutations may be associated with the classic features of Singleton-Merten syndrome including dental dysplasia, tendon rupture, and severe cardiac sequela.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is the causative pathogen of current COVID-19 pandemic, and insufficient production of type I interferon (IFN-I) is associated with the severe forms of the disease. Membrane (M) protein of SARS-CoV-2 has been reported to suppress host IFN-I production, but the underlying mechanism is not completely understood. In this study, SARS-CoV-2 M protein was confirmed to suppress the expression of IFNβ and interferon-stimulated genes induced by RIG-I, MDA5, IKKϵ, and TBK1, and to inhibit IRF3 phosphorylation and dimerization caused by TBK1. SARS-CoV-2 M could interact with MDA5, TRAF3, IKKϵ, and TBK1, and induce TBK1 degradation via K48-linked ubiquitination. The reduced TBK1 further impaired the formation of TRAF3–TANK–TBK1-IKKε complex that leads to inhibition of IFN-I production. Our study revealed a novel mechanism of SARS-CoV-2 M for negative regulation of IFN-I production, which would provide deeper insight into the innate immunosuppression and pathogenicity of SARS-CoV-2.

Tight regulation of inflammatory cytokine and interferon (IFN) production in innate immunity is pivotal for optimal control of pathogens and avoidance of immunopathology. The human Nod-like receptor (NLR) NLRP11 has been shown to regulate type I IFN and pro-inflammatory cytokine responses. Here, we identified the ATP-dependent RNA helicase DDX3X as a novel binding partner of NLRP11, using co-immunoprecipitation and LC-MS/MS. DDX3X is known to enhance type I IFN responses and NLRP3 inflammasome activation. We demonstrate that NLRP11 can abolish IKKϵ-mediated phosphorylation of DDX3X, resulting in lower type I IFN induction upon viral infection. These effects were dependent on the LRR domain of NLRP11 that we mapped as the interaction domain for DDX3X. In addition, NLRP11 also suppressed NLRP3-mediated caspase-1 activation in an LRR domain-dependent manner, suggesting that NLRP11 might sequester DDX3X and prevent it from promoting NLRP3-induced inflammasome activation. Taken together, our data revealed DDX3X as a central target of NLRP11, which can mediate the effects of NLRP11 on type I IFN induction as well as NLRP3 inflammasome activation. This expands our knowledge of the molecular mechanisms underlying NLRP11 function in innate immunity and suggests that both NLRP11 and DDX3X might be promising targets for modulation of innate immune responses.

Interleukin-18 receptor accessory protein (IL18RAP) is an indispensable subunit for the IL-18 receptor (IL-18R) complex’s ability to mediate high-affinity IL-18 binding and signalling transduction. Interest in IL-18 in systemic lupus erythematosus (SLE) has been mostly focused on its role as a type 1 T helper cell-driving cytokine. The functional significance of IL18RAP in mediating the IL-18-driven response in myeloid cells in SLE remains largely unexplored. This study aimed to investigate the expression and function significance of IL18RAP in neutrophils of SLE patients. By qRT-PCR and Western blot analyses, elevated expressions of IL18RAP mRNA and protein were observed in neutrophils from SLE patients—particularly those with a history of nephritis. IL18RAP expression correlated negatively with complement 3 level and positively with disease activity, with higher expression in patients exhibiting renal and immunological manifestations. The increased IL18RAP expression in SLE neutrophils could be attributed to elevated type I interferon level in sera. Functionally, neutrophils from SLE patients showed higher IL-18-mediated enhancement in reactive oxygen species (ROS) generation, which showed positive correlation with IL18RAP expression and could be neutralized by anti-IL18RAP blocking antibodies. Taken together, our findings suggest that IL-18 could contribute to SLE pathogenesis through mediation of neutrophil dysfunction via the upregulation of IL18RAP expression.

Salivary gland tumors are a heterogeneous group of neoplasms representing less than 10% of all head and neck tumors. Among salivary gland tumors, salivary duct carcinoma (SDC) is a rare, but highly aggressive malignant tumor resembling ductal breast carcinoma. Sublingual treatments are promising for SDC due to the induction of both local and systemic biological effects and to reduced systemic toxicity compared to other administration routes. In the present study, we first established that the sublingual administration of type I IFN (IFN-I) is safe and feasible, and exerts antitumor effects both as monotherapy and in combination with chemotherapy in transplantable tumor models, i.e., B16-OVA melanoma and EG.7-OVA lymphoma. Subsequently, we proved that sublingual IFN-I in combination with cyclophosphamide (CTX) induces a long-lasting reduction of tumor mass in NeuT transgenic mice that spontaneously develop SDC. Most importantly, tumor shrinkage in NeuT transgenic micewas accompanied by the emergence of tumor-specific cellular immune responses both in the blood and in the tumor tissue. Altogether, these results provide evidence that sublingual IFN holds promise in combination with chemotherapy for the treatment of cancer.

Interferons (IFNs) are induced by viruses and are the main regulators of the host antiviral response. They balance tissue tolerance and immune resistance against viral challenges. Like all cells in the human body, neutrophils possess the receptors for IFNs and contribute to antiviral host defense. To combat viruses, neutrophils utilize various mechanisms, such as viral sensing, neutrophil extracellular trap formation, and antigen presentation. These mechanisms have also been linked to tissue damage during viral infection and inflammation. In this review, we presented evidence that a complex cross-regulatory talk between IFNs and neutrophils initiates appropriate antiviral immune responses and regulates them to minimize tissue damage. We also explored recent exciting research elucidating the interactions between IFNs, neutrophils, and severe acute respiratory syndrome-coronavirus-2, as an example of neutrophil and IFN cross-regulatory talk. Dissecting the IFN-neutrophil paradigm is needed for well-balanced antiviral therapeutics and development of novel treatments against many major epidemic or pandemic viral infections, including the ongoing pandemic of the coronavirus disease that emerged in 2019.

One of the most powerful and multifaceted cytokines produced by immune cells are type I interferons (IFNs), the basal secretion of which contributes to the maintenance of immune homeostasis, while their activation-induced production is essential to effective immune responses. Although, each cell is capable of producing type I IFNs, plasmacytoid dendritic cells (pDCs) possess a unique ability to rapidly produce large amounts of them. Importantly, type I IFNs have a prominent role in the pathomechanism of various pDC-associated diseases. Deficiency in type I IFN production increases the risk of more severe viral infections and the development of certain allergic reactions, and supports tumor resistance; nevertheless, its overproduction promotes autoimmune reactions. Therefore, the tight regulation of type I IFN responses of pDCs is essential to maintain an adequate level of immune response without causing adverse effects. Here, our goal was to summarize those endogenous factors that can influence the type I IFN responses of pDCs, and thus might serve as possible therapeutic targets in pDC-associated diseases. Furthermore, we briefly discuss the current therapeutic approaches targeting the pDC-type I IFN axis in viral infections, cancer, autoimmunity, and allergy, together with their limitations defined by the Janus-faced nature of pDC-derived type I IFNs.

Neuromyelitis optica (NMO) is an inflammatory disease of the central nervous system (CNS) most frequently mediated by serum autoantibodies against the water channel aquaporin 4, expressed on CNS astrocytes, resulting in primary astrocytopathy. There is no cure for NMO, and treatment with Type I interferon (IFNI)-IFN beta is ineffective or even detrimental. We have previously shown that both NMO lesions and associated microglial activation were reduced in mice lacking the receptor for IFN beta. However, the role of microglia in NMO is not well understood. In this study, we clarify the pathomechanism for IFNI dependence of and the role of microglia in experimental NMO. Transcriptome analysis showed a strong IFNI footprint in affected CNS tissue as well as in microglial subpopulations. Treatment with IFN beta led to exacerbated pathology and further microglial activation as evidenced by expansion of a CD11c(+) subset of microglia. Importantly, depletion of microglia led to suppression of pathology and decrease of IFNI signature genes. Our data show a pro-pathologic role for IFNI-activated microglia in NMO and open new perspectives for microglia-targeted therapies.

It is believed that ∼7,000 messenger RNAs (mRNAs) are subject to N6-methyladenosine modification. The biological significance of this remains mysterious., Enteroviruses (EV) deploy two proteases that mediate viral polyprotein cleavage and host cell manipulation. Here, we report that EV 2A proteases cleave all three members of the YTHDF protein family, cytosolic N6-methyladenosine (m6A) “readers” that regulate target mRNA fate. YTHDF protein cleavage occurs very early during infection, before viral translation is detected or cytopathogenic effects are observed. Preemptive YTHDF protein depletion enhanced viral translation and replication but only in cells with restrained viral translation, signs of inefficient 2A protease activity, and protective innate host immune responses. This effect corresponded with repression of interferon (IFN)-stimulated gene (ISG) induction, while type I/III IFN production was not significantly altered. Moreover, YTHDF3 depletion impaired JAK/STAT signaling in cells treated with type I, but not type II, IFN. YTHDF3 depletion’s stimulatory effect on viral dynamics was dampened by JAK/STAT blockade and enhanced by type I IFN pretreatment of cells. We propose that EV 2A proteases cleave YTHDF proteins to antagonize ISG induction in infected cells.