Negative Regulator Of Type Research Articles

Autophagy mediated degradation of MITA/TBK1/IRF3 by a hnRNP family member attenuates interferon production in fish

HnRNP A/B belongs to the heterogeneous nuclear ribonucleoprotein (hnRNP) family and plays an important role in regulating viral protein translation and genome replication. Here, we found that overexpression of hnRNP A/B promoted spring viremia of carp virus (SVCV) and cyprinid herpesvirus 3 (CyHV3) replication. Further, hnRNP A/B was shown to act as a negative regulator of type I interferon (IFN) response. Mechanistically, hnRNP A/B interacted with MITA, TBK1 and IRF3 to initiate their degradation. In addition, hnRNP A/B bound to the kinase domain of TBK1, the C terminal domain of MITA and IAD domain of IRF3, and the RRM1 domain of hnRNP A/B bound to TBK1, RRM2 domain bound to IRF3 and MITA. Our study provides novel insights into the functions of hnRNP A/B in regulating host antiviral response.

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

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

Phosphorylation of aryl hydrocarbon receptor interacting protein by TBK1 negatively regulates IRF7 and the type I interferon response

The innate antiviral response to RNA viruses is initiated by sensing of viral RNAs by RIG-I-like receptors and elicits type I IFN production, which stimulates the expression of IFN-stimulated genes that orchestrate the antiviral response to prevent systemic infection. Negative regulation of type I IFN and its master regulator, transcription factor IRF7, is essential to maintain immune homeostasis. We previously demonstrated that AIP (aryl hydrocarbon receptor interacting protein) functions as a negative regulator of the innate antiviral immune response by binding to and sequestering IRF7 in the cytoplasm, thereby preventing IRF7 transcriptional activation and type I IFN production. However, it remains unknown how AIP inhibition of IRF7 is regulated. We show here that the kinase TBK1 phosphorylates AIP, and Thr-40 serves as the primary target for TBK1 phosphorylation. AIP Thr-40 plays critical roles in regulating AIP stability and mediating its interaction with IRF7. The AIP phospho-mimetic T40E exhibited increased proteasomal degradation and enhanced interaction with IRF7 compared to wild-type AIP. AIP T40E also blocked IRF7 nuclear translocation, which resulted in reduced type I IFN production and increased viral replication. In sharp contrast, AIP phospho-null mutant T40A had impaired IRF7 binding, and stable expression of AIP T40A in AIP-deficient mouse embryonic fibroblasts elicited a heightened type I IFN response and diminished RNA virus replication. Taken together, these results demonstrate that TBK1-mediated phosphorylation of AIP at Thr-40 functions as a molecular switch that enables AIP to interact with and inhibit IRF7, thus preventing overactivation of type I IFN genes by IRF7.

RNA-dependent RNA polymerase of SARS-CoV-2 regulate host mRNA translation efficiency by hijacking eEF1A factors

The RNA-dependent RNA polymerase (NSP12) of COVID-19 plays a significant role in the viral infection process, which promotes viral RNA replication by cooperating with NSP7 and NSP8, but little is known about its regulation on the function of host cells. We firstly found that overexpression of NSP12 had little effect on host mRNAs transcription. Using iCLIP technology, we found that NSP12 can bind a series of host RNAs with the conserved binding motif G(C/A/G)(U/G/A)UAG, especially ribosomal RNA. We found that NSP12 could directly bind to eEF1A factor via the NIRAN domain of NSP12 and N-terminal domain of eEF1A. NSP12 colocalized with eEF1A to inhibit type I interferon expression upon virus infection. In order to prove that NSP12 regulates the translation level of host cells, we found that NSP12 significantly affected the translation efficiency of many host mRNAs (such as ISG15, NF-κB2, ILK and SERPINI2) via ribosome profiling experiment, and the genes with significant upregulation in translation efficiency were mainly enriched in positive regulation of ubiquitin-dependent proteasomal process and NIK/NF-κB signaling pathway (such as NF-κB2, ILK), and negative regulation of type I interferon production, protein level of these genes were further confirmed in HEK293T and Calu3 cells upon NSP12 overexpression. These results indicate that NSP12 of SARS-CoV-2 can hijack the eEF1A factor to regulate translation efficiency of host mRNAs, which provides a new idea for us to evaluate the impact of SARS-CoV2 virus on the host and study the potential drug targets.

DDX5 inhibits type I IFN production by promoting degradation of TBK1 and disrupting formation of TBK1 - TRAF3 complex.

DExD/H-box helicase (DDX) 5 belongs to the DExD/H-box helicase family. DDX family members play differential roles in the regulation of innate antiviral immune response. However, whether DDX5 is involved in antiviral immunity remains unclear. In this study, we found that DDX5 serves as a negative regulator of type I interferon (IFN) response. Overexpression of DDX5 inhibited IFN production induced by Spring viremia of carp virus (SVCV) and poly(I:C) and enhanced virus replication by targeting key elements of the RLR signaling pathway (MAVS, MITA, TBK1, IRF3 and IRF7). Mechanistically, DDX5 directly interacted with TBK1 to promote its autophagy-mediated degradation. Moreover, DDX5 was shown to block the interaction between TRAF3 and TBK1, hence preventing nuclear translocation of IRF3. Together, these data shed light on the roles of DDX5 in regulating IFN response.

Capsaicin functions as a selective degrader of STAT3 to enhance host resistance to viral infection.

Although STAT3 has been reported as a negative regulator of type I interferon (IFN) signaling, the effects of pharmacologically inhibiting STAT3 on innate antiviral immunity are not well known. Capsaicin, approved for the treatment of postherpetic neuralgia and diabetic peripheral nerve pain, is an agonist of transient receptor potential vanilloid subtype 1 (TRPV1), with additional recognized potencies in anticancer, anti-inflammatory, and metabolic diseases. We investigated the effects of capsaicin on viral replication and innate antiviral immune response and discovered that capsaicin dose-dependently inhibited the replication of VSV, EMCV, and H1N1. In VSV-infected mice, pretreatment with capsaicin improved the survival rate and suppressed inflammatory responses accompanied by attenuated VSV replication in the liver, lung, and spleen. The inhibition of viral replication by capsaicin was independent of TRPV1 and occurred mainly at postviral entry steps. We further revealed that capsaicin directly bound to STAT3 protein and selectively promoted its lysosomal degradation. As a result, the negative regulation of STAT3 on the type I IFN response was attenuated, and host resistance to viral infection was enhanced. Our results suggest that capsaicin is a promising small-molecule drug candidate, and offer a feasible pharmacological strategy for strengthening host resistance to viral infection.

Negative regulation of type I interferon signaling by integrin-linked kinase permits dengue virus replication.

Dengue virus (DENV) infection can induce life-threatening dengue hemorrhagic fever/dengue shock syndrome in infected patients. DENV is a threat to global health due to its growing numbers and incidence of infection in the last 50 years. During infection, DENV expresses ten structural and nonstructural proteins modulating cell responses to benefit viral replication. However, the lack of knowledge regarding the cellular proteins and their functions in enhancing DENV pathogenesis impedes the development of antiviral drugs and therapies against fatal DENV infection. Here, we identified that integrin-linked kinase (ILK) is a novel enhancing factor for DENV infection by suppressing type I interferon (IFN) responses. Mechanistically, ILK binds DENV NS1 and NS3, activates Akt and Erk, and induces NF-κB-driven suppressor of cytokine signaling 3 (SOCS3) expression. Elevated SOCS3 in DENV-infected cells inhibits phosphorylation of STAT1/2 and expression of interferon-stimulated genes (ISGs). Inhibiting ILK, Akt, or Erk activation abrogates SOCS3 expression. In DENV-infected mice, the treatment of an ILK inhibitor significantly reduces viral loads in the brains, disease severity, and mortality rate. Collectively, our results show that ILK is a potential therapeutic target against DENV infection.

Type I Interferonopathy due to a Homozygous Loss-of-Inhibitory Function Mutation in STAT2

PurposeSTAT2 is both an effector and negative regulator of type I interferon (IFN-I) signalling. We describe the characterization of a novel homozygous missense STAT2 substitution in a patient with a type I interferonopathy.MethodsWhole-genome sequencing (WGS) was used to identify the genetic basis of disease in a patient with features of enhanced IFN-I signalling. After stable lentiviral reconstitution of STAT2-null human fibrosarcoma U6A cells with STAT2 wild type or p.(A219V), we performed quantitative polymerase chain reaction, western blotting, immunofluorescence, and co-immunoprecipitation to functionally characterize the p.(A219V) variant.ResultsWGS identified a rare homozygous single nucleotide transition in STAT2 (c.656C > T), resulting in a p.(A219V) substitution, in a patient displaying developmental delay, intracranial calcification, and up-regulation of interferon-stimulated gene (ISG) expression in blood. In vitro studies revealed that the STAT2 p.(A219V) variant retained the ability to transduce an IFN-I stimulus. Notably, STAT2 p.(A219V) failed to support receptor desensitization, resulting in sustained STAT2 phosphorylation and ISG up-regulation. Mechanistically, STAT2 p.(A219V) showed defective binding to ubiquitin specific protease 18 (USP18), providing a possible explanation for the chronic IFN-I pathway activation seen in the patient.ConclusionOur data indicate an impaired negative regulatory role of STAT2 p.(A219V) in IFN-I signalling and that mutations in STAT2 resulting in a type I interferonopathy state are not limited to the previously reported R148 residue. Indeed, structural modelling highlights at least 3 further residues critical to mediating a STAT2-USP18 interaction, in which mutations might be expected to result in defective negative feedback regulation of IFN-I signalling.

Phosphorylation of interferon regulatory factor 9 (IRF9).

IRF9 is a transcription factor that mediates the expression of interferon-stimulated genes (ISGs) through the Janus kinase-Signal transducer and activator of transcription (JAK-STAT) pathway. The JAK-STAT pathway is regulated through phosphorylation reactions, in which all components of the pathway are known to be phosphorylated except IRF9. The enigma surrounding IRF9 regulation by a phosphorylation event is intriguing. As IRF9 plays a major role in establishing an antiviral state in host cells, the topic of IRF9 regulation warrants deeper investigation. Initially, total lysates of 2fTGH and U2A cells (transfected with recombinant IRF9) were filter-selected and concentrated using phosphoprotein enrichment assay. The phosphoprotein state of IRF9 was further confirmed using Phos-tag™ assay. All protein expression was determined using Western blotting. Tandem mass spectrometry was conducted on immunoprecipitated IRF9 to identify the phosphorylated amino acids. Finally, site-directed mutagenesis was performed and the effects of mutated IRF9 on relevant ISGs (i.e., USP18 and Mx1) was evaluated using qPCR. IRF9 is phosphorylated at S252 and S253 under IFNβ-induced condition and R242 under non-induced condition. Site-directed mutagenesis of S252 and S253 to either alanine or aspartic acid has a modest effect on the upregulation of USP18 gene-a negative regulator of type I interferon (IFN) response-but not Mx1 gene. Our preliminary study shows that IRF9 is phosphorylated and possibly regulates USP18 gene expression. However, further in vivo studies are needed to determine the significance of IRF9 phosphorylation.

Cytokine Receptor-Like Factor 3 Negatively Regulates Antiviral Immunity by Promoting the Degradation of TBK1 in Teleost Fish.

The cytokine receptor-like factor 3 (Crlf3) belongs to the orphan class I cytokine receptors and is identified as a neuroprotective erythropoietin receptor. In previous studies of Crlf3, few focused on its role in innate immunity. Therefore, this study explored the regulatory role of Crlf3 in innate immunity. TANK-binding kinase 1 (TBK1) is a vital adaptor protein for the activation of the RLRs-MVAS-IRF3 antiviral signaling axis; thus, its expression and activity must be tightly regulated to maintain immune homeostasis and avoid undesirable effects. Here, we report that Crlf3 is a negative regulator of type I interferon production. The expression of Crlf3 is induced by poly(I·C) or Siniperca chuatsi rhabdovirus (SCRV) treatment. Silencing of Crlf3 enhanced poly(I·C)- and SCRV-induced type I interferon production, whereas overexpression of Crlf3 suppressed type I interferon production. Mechanistically, Crlf3 interacted with TBK1 via its N domain and then inhibited type I interferon production by promoting TBK1 proteasomal degradation through K48-linked polyubiquitination. Our study shows that Crlf3 is a key factor for viral escape from innate antiviral immunity in fish and provides a new perspective on mammalian resistance to viral invasion. IMPORTANCE The expression of Crlf3 was upregulated with SCRV invasion, which proved that Crlf3 was involved in the regulation of the antiviral immune response. In this study, we found that the existence of Crlf3 promoted the replication of SCRV. Therefore, it is reasonable to believe that SCRV evades innate immune attack with the assistance of Crlf3. In addition, we report that Crlf3 negatively regulates interferon (IFN) induction by promoting the degradation of TBK1 in fish. We showed that Crlf3 is evenly distributed in the cytoplasm and interacts with TBK1. Further studies showed that Crlf3 specifically mediates K48-linked ubiquitination of TBK1 and promotes TBK1 degradation, resulting in a marked inhibition of retinoic acid-inducible gene I (RIG-I) downstream signaling.

Three IRF4 paralogs act as negative regulators of type Ⅰ IFN responses in yellow catfish (Pelteobagrus fulvidraco)

IRF4 is a master member of the interferon regulatory factor (IRF) family playing vital regulatory roles in immune system development and function. Tetrapods have a single-copy IRF4 gene, while teleosts harbor duplicated IRF4 genes. This work describes three IRF4 paralogs from yellow catfish (Pelteobagrus fulvidraco), designated PfIRF4A, PfIRF4B and PfIRF4B-like. These genes all contain a typical IRF structural architecture. Phylogenic and synteny analyses indicate that they should arise from the teleost-specific whole-genome duplication. PfIRF4 genes are abundantly expressed in the immune-related tissues and upregulated by PolyI:C, LPS, and Edwardsiella ictaluri. Ectopic expression of these genes inhibits the activation of fish type Ⅰ IFN promoters and downregulates the transcription levels of IFN-responsive genes, thus allowing the efficient replication of a fish rhabdovirus, spring viremia of carp virus (SVCV). PfIRF4s possess a repressive effect on MyD88-mediated activation of IFN and NF-κB. Some differences are observed between each individual paralog. PfIRF4B is the main form expressed across the tissues and the most up-regulated one after pathogen induction. It exerts a stronger inhibitory effect on IFN antiviral response than the other two paralogs. PfIRF4A and PfIRF4B-like are primarily present in the nucleus, while PfIRF4B displays colocalization and direct associations with MyD88 in the cytoplasm. Overall, the data demonstrate that three PfIRF4 paralogs show shared and individual functional properties in the negative regulation of type Ⅰ IFN response.

Vimentin inhibits type I interferon production by disrupting the TBK1-IKKε-IRF3 axis.

Cytoskeleton proteins have been reported to be involved in the host antiviral immune responses. However, how cytoskeleton proteins regulate host antiviral immune responses is not fully understood. Here we report that the cytoskeletal protein vimentin is a negative regulator of type I interferon (IFN-I) production upon viral infection. Ectopic expression of vimentin suppresses RNA- and DNA viruses-induced IFN-I production, whereas knockout of vimentin expression enhances IFN-I production. Viral infection increases vimentin expression and ultimately inhibits IFN-I production. Mechanistically, upregulated vimentin interacts with TBK1 and IKKε to disrupt the interactions of TBK1-IRF3 and IKKε-IRF3, resulting in inhibition of IRF3 phosphorylation and nuclear translocation. Furthermore, we generate vimentin knockout mice to confirm that deficiency of vimentin gene in mice suppressed encephalomyocarditis virus replication invivo. Our findings demonstrates that vimentin plays an important role in regulating IFN-I production, revealing its antiviral function of the cytoskeletal protein vimentin.

MST4 negatively regulates type I interferons production via targeting MAVS-mediated pathway

BackgroundCytosolic RNA sensing can elicit immune responses against viral pathogens. However, antiviral responses must be tightly regulated to avoid the uncontrolled production of type I interferons (IFN) that might have deleterious effects on the host. Upon bacterial infection, the germinal center kinase MST4 can directly phosphorylate the adaptor TRAF6 to limit the inflammatory responses, thereby avoiding the damage caused by excessive immune activation. However, the molecular mechanism of how MST4 regulates virus-mediated type I IFN production remains unknown.MethodsThe expression levels of IFN-β, IFIT1, and IFIT2 mRNA were determined by RT-PCR. The expression levels of p-IRF3, IRF3, RIG-I, MAVS, and MST4 proteins were determined by Western blot. The effect of secreted level of IFN-β was measured by ELISA. The relationship between MST4 and MAVS was investigated by immunofluorescence staining and coimmunoprecipitation.ResultsIn this study, we reported that MST4 can act as a negative regulator of type I IFN production. Ectopic expression of MST4 suppressed the Poly (I:C) (polyino-sinic-polycytidylic acid)- and Sendai virus (SeV)-triggered production of type I IFN, while the knockdown of MST4 enhanced the production of type I IFN. Mechanistically, upon SeV infection, the MST4 competed with TRAF3 to bind to the 360–540 domain of MAVS, thereby inhibiting the TRAF3/MAVS association. Additionally, MST4 facilitated the interaction between the E3 ubiquitin ligase Smurf1 and MAVS. This promoted the K48-linked ubiquitination of MAVS, thereby accelerating the ubiquitin-mediated proteasome degradation of MAVS.ConclusionsOur findings showed that MST4 acted as a crucial negative regulator of RLR-mediated type I IFN production.-szD4SjHvt2BGoxb8QjZ8LVideo

NLRP6 negatively regulates type 2 immune responses in mice.

Inflammasomes are large protein complexes that assemble in the cytosol in response to danger such as tissue damage or infection. Following activation, inflammasomes trigger cell death and the release of biologically active forms of pro-inflammatory cytokines interleukin (IL)-1β and IL-18. NOD-like receptor family pyrin domain containing 6 (NLRP6) inflammasome is required for IL-18 secretion by intestinal epithelial cells, macrophages, and T cells, contributing to homeostasis and self-defense against pathogenic microbes. However, the involvement of NLRP6 in type 2 lung inflammation remains elusive. Wild-type (WT) and Nlrp6-/- mice were used. Birch pollen extract (BPE)-induced allergic lung inflammation, eosinophil recruitment, Th2-related cytokine and chemokine production, airway hyperresponsiveness, and lung histopathology, Th2 cell differentiation, GATA3, and Th2 cytokines expression, were determined. Nippostrongylus brasiliensis (Nb) infection, worm count in intestine, type 2 innate lymphoid cell (ILC2), and Th2 cells in lungs were evaluated. We demonstrate in Nlrp6-/- mice that a mixed Th2/Th17 immune responses prevailed following birch pollen challenge with increased eosinophils, ILC2, Th2, and Th17 cell induction and reduced IL-18 production. Nippostrongylus brasiliensis infected Nlrp6-/- mice featured enhanced early expulsion of the parasite due to enhanced type 2 immune responses compared to WT hosts. In vitro, NLRP6 repressed Th2 polarization, as shown by increased Th2 cytokines and higher expression of the transcription factor GATA3 in the absence of NLRP6. Exogenous IL-18 administration partially reduced the enhanced airways inflammation in Nlrp6-/- mice. In summary, our data identify NLRP6 as a negative regulator of type 2 immune responses.

A partial form of inherited human USP18 deficiency underlies infection and inflammation.

Human USP18 is an interferon (IFN)-stimulated gene product and a negative regulator of type I IFN (IFN-I) signaling. It also removes covalently linked ISG15 from proteins, in a process called deISGylation. In turn, ISG15 prevents USP18 from being degraded by the proteasome. Autosomal recessive complete USP18 deficiency is life-threatening in infancy owing to uncontrolled IFN-I-mediated autoinflammation. We report three Moroccan siblings with autoinflammation and mycobacterial disease who are homozygous for a new USP18 variant. We demonstrate that the mutant USP18 (p.I60N) is normally stabilized by ISG15 and efficient for deISGylation but interacts poorly with the receptor-anchoring STAT2 and is impaired in negative regulation of IFN-I signaling. We also show that IFN-γ-dependent induction of IL-12 and IL-23 is reduced owing to IFN-I-mediated impairment of myeloid cells to produce both cytokines. Thus, insufficient negative regulation of IFN-I signaling by USP18-I60N underlies a specific type I interferonopathy, which impairs IL-12 and IL-23 production by myeloid cells, thereby explaining predisposition to mycobacterial disease.

DDX19 inhibits RLR/IRF3 mediated type I interferon signaling of black carp Mylopharyngodon piceus by restricting IRF3 from entering nucleus

Members of DExD/H-box helicase family are important receptors for detecting viral nucleic acids. Mammalian DExD/H-box RNA helicase 19 (DDX19) has been reported to play as a negative regulator of type I interferon (IFN). However, the role of teleost DDX19 during the innate immune response is still obscure. In this study, the DDX19 homolog of black carp (bcDDX19) has been cloned and characterized. The open reading frame (ORF) of bcDDX19 consists of 1440 nucleotides and encodes contains 479 amino acids. bcDDX19 migrates around 54 kDa in immunoblotting assay and is identified as a cytosolic protein by immunofluorescence staining. The qPCR results show that the transcription of bcDDX19 in host cells rises in response to Lipopolysaccharide (LPS), Poly (I:C), spring viremia of carp virus (SVCV) or grass carp reovirus (GCRV) stimulation. In the report assay, bcDDX19 suppressed bcIRF3-, but not bcIRF7-mediated transcription activities of bcIFNa and DrIFNφ1 promoter. Co-immunoprecipitation assays identify that bcDDX19 interacts with bcIRF3 but not bcIRF7. According, plaque assay results demonstrate that bcDDX19 dampens bcIRF3- but not bcIRF7-induced antiviral activity of EPC cells. Knockdown of bcDDX19 offers host cells the increase of mRNA levels of bcIFNa, bcMx1 and bcViperin, and the improved antiviral ability against SVCV. In addition, the nuclear translocation of bcIRF3 is dampened when co-expressed with bcDDX19. All of these findings demonstrate that bcDDX19 negatively regulates RLR/IRF3 mediated type I IFN signaling in the innate immune activation through restricting IRF3 from entering nucleus.

OASL1-Mediated Inhibition of Type I IFN Reduces Influenza A Infection-Induced Airway Inflammation by Regulating ILC2s.

PurposeThree observations drove this study. First, 2′-5′-oligoadenylate synthetase-like protein (OASL) is a negative regulator of type I interferon (IFN). Second, type I IFN plays a central role during virus infections and the pathogenesis of various diseases, including asthma. Third, influenza A virus (IAV) causes non-eosinophilic asthma. To evaluate the potential relationships between OASL, type I IFN, and pulmonary innate immune cells in IAV-induced acute airway inflammation by using Oasl1-/- mice.MethodsAsthma was induced in wild-type (WT) and Oasl1-/- mice with IAV or ovalbumin (OVA). Airway hyperreactivity (AHR) and immune cell infiltration in the bronchoalveolar lavage (BAL) fluids were measured. The immune cells in the lungs were analyzed by flow cytometry. To investigate the ability of type I IFN to shape the response of lung type 2 innate lymphoid cells (ILC2s), IFN-α was treated intratracheally. Plasmacytoid dendritic cells (pDCs) sorted from bone marrow and ILC2s sorted from lungs of naive mice were co-cultured with/without interferon-alpha receptor subunit 1 (IFNAR-1)-blocking antibodies.ResultsIn the IAV-induced asthma model, Oasl1-/- mice developed greater AHR and immune cell infiltration in the BAL fluids than WT mice. This was not observed in OVA-induced asthma, a standard model of allergen-induced asthma. The lungs of infected Oasl1-/- mice also had elevated DC numbers and Ifna expression and depressed IAV-induced ILC2 responses, namely, proliferation and type 2 cytokine and amphiregulin production. Intratracheal administration of type I IFN in naïve mice suppressed lung ILC2 production of type 2 cytokines and amphiregulin. Co-culture of ILC2s with pDCs showed that pDCs inhibit the function of ILC2s by secreting type I IFN.ConclusionsOASL1 may impede the IAV-induced acute airway inflammation that drives AHR by inhibiting IAV-induced type I IFN production from lung DCs, thereby preserving the functions of lung ILC2s, including their amphiregulin production.

A Novel Familial Case Report of Genetic Syndrome Mimicking Congenital TORCH infections; Pseudo-TORCH Syndrome 2

Background:Pseudo-TORCH syndrome (PTS) is a group of autosomal recessive disorders that clinically and radiologically mimic TORCH congenital infections. The prevalence of pseudo-TORCH syndrome 2 is 1 in 1,000,000 cases worldwide. This novel disorder is extremely rare, and is generally detected by prenatal diagnosis through next generation sequencing (NGS) during pregnancy. In this study, a familial case of pseudo-TORCH syndrome 2 with novel non-sense mutation in the ubiquitin-specific peptidase 18 (USP 18) gene in the parents was reported, who are heterozygous asymptomatic carriers; however, all children have inherited a homozygous pathogenic form of USP18, which is an important negative regulator of type I interferon (IFN) signal transduction. To the best of our knowledge, this is the first case of a novel mutation of USP18 seen in a family with pseudo-TORCH syndrome 2 (PTS 2) from India.Case Presentation:A 23-year-old pregnant woman with bad obstetric history, including intrauterine and neonatal mortality was referred to the Institute of Genetics in the year 2021 for clinical and genetic evaluation. Advanced clinical exome sequencing of the parents and the fetus revealed heterozygous carrier status in parents and homozygous mutation in USP 18 gene in the progeny leading to pseudo-TORCH-2 syndrome.Conclusion:The present case highlights the significance of carrier screening, prenatal diagnosis, and genetic counseling in couples with bad obstetric history for the detection of rare genetic disorders with poor prognosis.

Selective autophagy controls the stability of TBK1 via NEDD4 to balance host defense.

As a core kinase of antiviral immunity, the activity and stability of TANK-binding kinase 1 (TBK1) is tightly controlled by multiple post-translational modifications. Although it has been demonstrated that TBK1 stability can be regulated by ubiquitin-dependent proteasome pathway, it is unclear whether another important protein degradation pathway, autophagosome pathway, can specifically affect TBK1 degradation by cargo receptors. Here we report that E3 ubiquitin ligase NEDD4 functions as a negative regulator of type I interferon (IFN) signaling by targeting TBK1 for degradation at the late stage of viral infection, to prevent the host from excessive immune response. Mechanically NEDD4 catalyzes the K27-linked poly-ubiquitination of TBK1 at K344, which serves as a recognition signal for cargo receptor NDP52-mediated selective autophagic degradation. Taken together, our study reveals the regulatory role of NEDD4 in balancing TBK1-centered type I IFN activation and provides insights into the crosstalk between selective autophagy and antiviral signaling.

Mechanisms underlying genetic susceptibility to multisystem inflammatory syndrome in children (MIS-C)

BackgroundMultisystem inflammatory syndrome in children (MIS-C) is a pediatric complication of severe acute respiratory syndrome coronavirus 2 infection that is characterized by multiorgan inflammation and frequently by cardiovascular dysfunction. It occurs predominantly in otherwise healthy children. We previously reported haploinsufficiency of suppressor of cytokine signaling 1 (SOCS1), a negative regulator of type I and II interferons, as a genetic risk factor for MIS-C.ObjectivesWe aimed to identify additional genetic mechanisms underlying susceptibility to severe acute respiratory syndrome coronavirus 2–associated MIS-C.MethodsIn a single-center, prospective cohort study, whole exome sequencing was performed on patients with MIS-C. The impact of candidate variants was tested by using patients’ PBMCs obtained at least 7 months after recovery.ResultsWe enrolled 18 patients with MIS-C (median age = 8 years; interquartile range = 5-12.25 years), of whom 89% had no conditions other than obesity. In 2 boys with no significant infection history, we identified and validated hemizygous deleterious defects in XIAP, encoding X-linked inhibitor of apoptosis, and CYBB, encoding cytochrome b-245, beta subunit. Including the previously reported SOCS1 haploinsufficiency, a genetic diagnosis was identified in 3 of 18 patients (17%). In contrast to patients with mild COVID-19, patients with defects in SOCS1, XIAP, or CYBB exhibit an inflammatory immune cell transcriptome with enrichment of differentially expressed genes in pathways downstream of IL-18, oncostatin M, and nuclear factor κB, even after recovery.ConclusionsAlthough inflammatory disorders are rare in the general population, our cohort of patients with MIS-C was enriched for monogenic susceptibility to inflammation. Our results support the use of next-generation sequencing in previously healthy children who develop MIS-C.