Grainyhead-like-2, an epithelial master programmer, promotes interferon induction and suppresses breast cancer recurrence.

Type-I and -III interferons play a central role in immune rejection of pathogens and tumors, thus promoting immunogenicity and suppressing tumor recurrence. Double strand RNA is an important ligand that stimulates tumor immunity via interferon responses. Differentiation of embryonic stem cells to pluripotent epithelial cells activates the interferon response during development, raising the question of whether epithelial vs. mesenchymal gene signatures in cancer potentially regulate the interferon pathway as well. Here, using genomics and signaling approaches, we show that Grainyhead-like-2 (GRHL2), a master programmer of epithelial cell identity, promotes type-I and -III interferon responses to double-strand RNA. GRHL2 enhanced the activation of IRF3 and relA/NF-kB and the expression of IRF1; a functional GRHL2 binding site in the IFNL1 promoter was also identified. Moreover, time to recurrence in breast cancer correlated positively with GRHL2 protein expression, indicating that GRHL2 is a tumor recurrence suppressor, consistent with its enhancement of interferon responses. These observations demonstrate that epithelial cell identity supports interferon responses in the context of cancer.

Mammalian reovirus µ1 protein attenuates RIG-I and MDA5-mediated signaling transduction by blocking IRF3 phosphorylation and nuclear translocation.

Mammalian reovirus (MRV) is a non-enveloped, gene segmented double-stranded RNA (dsRNA) virus. It is an important zoonotic pathogen that infects many mammals and vertebrates that act as natural hosts and causes respiratory and digestive tract diseases. Studies have reported that RIG-I and MDA5 in the innate immune cytoplasmic RNA-sensing RIG-like receptor (RLR) signaling pathway can recognize dsRNA from MRV and promote antiviral type I interferon (IFN) responses. However, the mechanism by which many MRV-encoded proteins evade the host innate immune response remains unclear. Here, we show that exogenous µ1 protein promoted the proliferation of MRV in vitro, while knockdown of MRV µ1 protein expression by shRNA could impair MRV proliferation. Specifically, µ1 protein inhibited MRV or poly(I:C)-induced IFN-ß expression, and attenuated RIG-I/MDA5-mediated signaling axis transduction during MRV infection. Importantly, we found that µ1 protein significantly decreased IFN-ß mRNA expression induced by MDA5, RIG-I, MAVS, TBK1, IRF3(5D), and degraded the protein expression of exogenous MDA5, RIG-I, MAVS, TBK1 and IRF3 via the proteasomal and lysosomal pathways. Additionally, we show that µ1 protein can physically interact with MDA5, RIG-I, MAVS, TBK1, and IRF3 and attenuate the RIG-I/MDA5-mediated signaling cascades by blocking the phosphorylation and nuclear translocation of IRF3. In conclusion, our findings reveal that MRV outer capsid protein µ1 is a key factor in antagonizing RLRs signaling cascades and provide new strategies for effective prevention and treatment of MRV infection.

Fucoidan MF4 from Fucus vesiculosus inhibits Lewis lung cancer via STING-TBK1-IRF3 pathway.

Fucoidan, a sulfated polysaccharide of marine origin found in brown algae and sea cucumbers, has been identified as a neuroprotective compound. In this study, a novel fucoidan MF4 was extracted from Fucus vesiculosus and isolated using Q-Sepharose fast-flow ion-exchange chromatography. The physicochemical properties of MF4 were characterized. MF4 is primarily composed of fucose, xylose, galactose, glucose, and mannose in a molar ratio of 12.3: 4.9: 1.1: 1.0: 1.1, with an average molecular weight of 67.7 kDa. Notably, MF4 demonstrated suppression of LLC tumor growth in vivo. RNA-sequencing analysis revealed that MF4 enhanced the expression of type I interferon-associated downstream genes in macrophages. Furthermore, MF4 increased the levels of phosphorylated TBK1 and IRF3 proteins in vitro. By activating the STING-TBK1-IRF3 signaling pathway, MF4 may enhance the antitumor activity of macrophages. Taken together, MF4 has promising potential as an antitumor and immunomodulatory agent.

NS7a of SADS-CoV promotes viral infection via inducing apoptosis to suppress type III interferon production.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a newly discovered swine coronavirus with potential cross-species transmission risk. Although SADS-CoV-induced host cell apoptosis and innate immunity antagonization has been revealed, underlying signaling pathways remain obscure. Here, we demonstrated that infection of SADS-CoV induced apoptosis in vivo and in vitro, and that viral protein NS7a is mainly responsible for SADS-CoV-induced apoptosis in host cells. Furthermore, we found that NS7a interacted with apoptosis-inducing factor mitochondria associated 1 (AIFM1) to activate caspase-3 via caspase-6 in SADS-CoV-infected cells, and enhanced SADS-CoV replication. Importantly, NS7a suppressed poly(I:C)-induced expression of type III interferon (IFN-λ) via activating caspase-3 to cleave interferon regulatory factor 3 (IRF3), and caspase-3 inhibitor protects piglets against SADS-CoV infection in vivo. These findings reveal how SADS-CoV induced apoptosis to inhibit innate immunity and provide a valuable clue to the development of effective drugs for the clinical control of SADS-CoV infection.IMPORTANCEOver the last 20 years, multiple animal-originated coronaviruses, including severe acute respiratory syndrome coronavirus (SARS-CoV), middle east respiratory syndrome coronavirus (MERS-CoV), and SARS-CoV-2, have caused millions of deaths, seriously jeopardized human health, and hindered social development, indicating that the study of animal-originated coronaviruses with potential for cross-species transmission is particularly important. Bat-originated swine acute diarrhea syndrome coronavirus (SADS-CoV), discovered in 2017, can not only cause fatal diarrhea in piglets, but also infect multiple human cells, with a potential risk of cross-species transmission, but its pathogenesis is unclear. In this study, we demonstrated that NS7a of SADS-CoV suppresses IFN-λ production via apoptosis-inducing factor mitochondria associated 1 (AIFM1)-caspase-6-caspase-3-interferon regulatory factor 3 (IRF3) pathway, and caspase-3 inhibitor (Z-DEVD-FMK) can effectively inhibit SADS-CoV replication and protect infected piglets. Our findings in this study contribute to a better understanding of SADS-CoV-host interactions as a part of the coronaviruses pathogenesis and using apoptosis-inhibitor as a drug as potential therapeutic approaches for prevention and control of SADS-CoV infection.

Oxygen enhances antiviral innate immunity through maintenance of EGLN1-catalyzed proline hydroxylation of IRF3.

Oxygen is essential for aerobic organisms, but little is known about its role in antiviral immunity. Here, we report that during responses to viral infection, hypoxic conditions repress antiviral-responsive genes independently of HIF signaling. EGLN1 is identified as a key mediator of the oxygen enhancement of antiviral innate immune responses. Under sufficient oxygen conditions, EGLN1 retains its prolyl hydroxylase activity to catalyze the hydroxylation of IRF3 at proline 10. This modification enhances IRF3 phosphorylation, dimerization and nuclear translocation, leading to subsequent IRF3 activation. Furthermore, mice and zebrafish with Egln1 deletion, treatment with the EGLN inhibitor FG4592, or mice carrying an Irf3 P10A mutation are more susceptible to viral infections. These findings not only reveal a direct link between oxygen and antiviral responses, but also provide insight into the mechanisms by which oxygen regulates innate immunity.

Mitochondrial (mt)DNA-cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) signaling promotes pyroptosis of macrophages via interferon regulatory factor (IRF)7/IRF3 activation to aggravate lung injury during severe acute pancreatitis.

BACKGROUND: Macrophage proinflammatory activation contributes to the pathology of severe acute pancreatitis (SAP) and, simultaneously, macrophage functional changes, and increased pyroptosis/necrosis can further exacerbate the cellular immune suppression during the process of SAP, where cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) plays an important role. However, the function and mechanism of cGAS-STING in SAP-induced lung injury (LI) remains unknown. METHODS: Lipopolysaccharide (LPS) was combined with caerulein-induced SAP in wild type, cGAS -/- and sting -/- mice. Primary macrophages were extracted via bronchoalveolar lavage and peritoneal lavage. Ana-1 cells were pretreated with LPS and stimulated with nigericin sodium salt to induce pyroptosis in vitro. RESULTS: SAP triggered NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome activation-mediated pyroptosis of alveolar and peritoneal macrophages in mouse model. Knockout of cGAS/STING could ameliorate NLRP3 activation and macrophage pyroptosis. In addition, mitochondrial (mt)DNA released from damaged mitochondria further induced macrophage STING activation in a cGAS- and dose-dependent manner. Upregulated STING signal can promote NLRP3 inflammasome-mediated macrophage pyroptosis and increase serum interleukin (IL)-6, IL-1ß, and tumor necrosis factor (TNF)-α levels and, thus, exacerbate SAP-associated LI (SAP-ALI). Downstream molecules of STING, IRF7, and IRF3 connect the mtDNA-cGAS-STING axis and the NLRP3-pyroptosis axis. CONCLUSIONS: Negative regulation of any molecule in the mtDNA-cGAS-STING-IRF7/IRF3 pathway can affect the activation of NLRP3 inflammasomes, thereby reducing macrophage pyroptosis and improving SAP-ALI in mouse model.

The dsRNA isolated from Escherichia coli infected with the MS2 bacteriophage induces the production of interferons.

Viral infections pose a significant threat to public health, and the production of interferons represents one of the most critical antiviral innate immune responses of the host. Consequently, the screening and identification of compounds or reagents that induce interferon production are of paramount importance. This study commenced with the cultivation of host bacterium 15,597, followed by the infection of Escherichia coli with the MS2 bacteriophage. Utilizing the J2 capture technique, a class of dsRNA mixtures (MS2+15,597) was isolated from the E. coli infected with the MS2 bacteriophage. Subsequent investigations were conducted on the immunostimulatory activity of the MS2+15,597 mixture. The results indicated that the dsRNA mixtures (MS2+15,597) extracted from E. coli infected with the MS2 bacteriophage possess the capability to activate innate immunity, thereby inducing the production of interferon-ß. These dsRNA mixtures can activate the RIG-I and TLR3 pattern recognition receptors, stimulating the expression of interferon stimulatory factors 3/7, which in turn triggers the NF-κB signaling pathway, culminating in the cellular production of interferon-ß to achieve antiviral effects. This study offers novel insights and strategies for the development of broad-spectrum antiviral drugs, potentially providing new modalities for future antiviral therapies.

Computational investigation of the inhibitory interaction of IRF3 and SARS-CoV-2 accessory protein ORF3b.

ORF3b is one of the SARS-CoV-2 accessory proteins. Previous experimental study suggested that ORF3b prevents IRF3 translocating to nucleus. However, the biophysical mechanism of ORF3b-IRF3 interaction is elusive. Here, we explored the conformation ensemble of ORF3b using all-atom replica exchange molecular dynamics simulation. Disordered ORF3b has mixed α-helix, ß-turn and loop conformers. The potential ORF3b-IRF3 binding modes were searched by docking representative ORF3b conformers with IRF3, and 50 ORF3b-IRF3 complex poses were screened using molecular dynamics simulations ranging from 500 to 1000 ns. We found that ORF3b binds IRF3 predominantly on its CBP binding and phosphorylated pLxIS motifs, with CBP binding site has the highest binding affinity. The ORF3b-IRF3 binding residues are highly conserved in SARS-CoV-2. Our results provided biophysics insights into ORF3b-IRF3 interaction and explained its interferon antagonism mechanism.

Salvianolic acid A prevents UV-induced skin damage by inhibiting the cGAS-STING pathway.

DNA damage resulting from UV irradiation on the skin has been extensively documented in numerous studies. In our prior investigations, we demonstrated that UVB-induced DNA breakage from keratinocytes can activate the cGAS-STING pathway in macrophages. The cGAS-STING signaling pathway serves as the principal effector for detecting and responding to abnormal double-stranded DNA in the cytoplasm. Expanding on our previous findings, we have further validated that STING knockout significantly diminishes UVB-induced skin damage, emphasizing the critical role of cGAS-STING activation in this context. Salvianolic acid A, a principal active constituent of Salvia miltiorrhiza Burge, has been extensively studied for its therapeutic effects in conditions such as coronary heart disease, angina pectoris, and diabetic peripheral neuropathy. However, its effect on cGAS-STING pathway and its ability to alleviate skin damage have not been previously reported. In a co-culture system, supernatant from UVB-treated keratinocytes induced IRF3 activation in macrophages, and this activation was inhibited by salvianolic acid A. Our investigation, employing photodamage and photoaging models, establishes that salvianolic acid A effectively mitigates UV-induced epidermal thickening and collagen degeneration. Treatment with salvianolic acid A significantly reduced skin damage, epidermal thickness increase, and keratinocyte hyperproliferation compared to the untreated photo-damage and photoaging model groups. In summary, salvianolic acid A emerges as a promising candidate for preventing UV-induced skin damage by inhibiting cGAS-STING activation. This research enhances our understanding of the intricate mechanisms underlying skin photodamage and provides a potential avenue for the development of therapeutic interventions.

Inhibition of spinal BRD4 alleviates pyroptosis and M1 microglia polarization via STING-IRF3 pathway in morphine-tolerant rats.

BACKGROUND: Morphine tolerance has been a challenging medical issue. Neuroinflammation is considered as a critical mechanism for the development of morphine tolerance. Bromodomain-containing protein 4 (BRD4), a key regulator in cell damage and inflammation, participates in the development of chronic pain. However, whether BRD4 is involved in morphine tolerance and the underlying mechanisms remain unknown. METHODS: The morphine-tolerant rat model was established by intrathecal administration of morphine twice daily for 7 days. Behavior test was assessed by a tail-flick latency test. The roles of BRD4, pyroptosis, microglia polarization and related signaling pathways in morphine tolerance were elucidated by Western blot, real-time quantitative polymerase chain reaction, and immunofluorescence. RESULTS: Repeated morphine administration upregulated BRD4 level, induced pyroptosis, and promoted microglia M1-polarization in spinal cord, accompanied by the release of proinflammatory cytokines, such as TNF-α and IL-1ß. JQ-1, a BRD4 antagonist, alleviated the development of morphine tolerance, diminished pyroptosis and induced the switch of microglia from M1 to M2 phenotype. Mechanistically, stimulator of interferon gene (STING)- interferon regulatory factor 3 (IRF3) pathway was activated and the protective effect of JQ-1 against morphine tolerance was at least partially mediated by inhibition of STING-IRF3 pathway. CONCLUSION: This study demonstrated for the first time that spinal BRD4 contributes to pyroptosis and switch of microglia polarization via STING-IRF3 signaling pathway during the development of morphine tolerance, which extend the understanding of the neuroinflammation mechanism of morphine tolerance and provide an alternative strategy for the precaution against of this medical condition.

HDAC5 enhances IRF3 activation and is targeted for degradation by protein C6 from orthopoxviruses including Monkeypox virus and Variola virus.

Histone deacetylases (HDACs) regulate gene expression and innate immunity. Previously, we showed that HDAC5 is degraded during Vaccinia virus (VACV) infection and is a restriction factor for VACV and herpes simplex virus type 1. Here, we report that HDAC5 promotes interferon regulatory factor 3 (IRF3) activation downstream of Toll-IL-1 receptor (TIR) domain-containing adaptor molecule-1 or Sendai virus-mediated stimulation without requiring HDAC activity. Loss of HDAC5-mediated IRF3 activation is restored by re-introduction of HDAC5 but not HDAC1 or HDAC4. The antiviral activity of HDAC5 is antagonized by VACV protein C6 and orthologs from the orthopoxviruses cowpox, rabbitpox, camelpox, monkeypox, and variola. Infection by many of these viruses induces proteasomal degradation of HDAC5, and expression of C6 alone can induce HDAC5 degradation. Mechanistically, C6 binds to the dimerization domain of HDAC5 and prevents homodimerization and heterodimerization with HDAC4. Overall, this study describes HDAC5 as a positive regulator of IRF3 activation and provides mechanistic insight into how the poxviral protein C6 binds to HDAC5 to antagonize its function.

Type I Interferon, Induced by Adenovirus or Adenoviral Vector Infection, Regulates the Cytokine Response to Lipopolysaccharide in a Macrophage Type-Specific Manner.

INTRODUCTION: While TLR ligands derived from microbial flora and pathogens are important activators of the innate immune system, a variety of factors such as intracellular bacteria, viruses, and parasites can induce a state of hyperreactivity, causing a dysregulated and potentially life-threatening cytokine over-response upon TLR ligand exposure. Type I interferon (IFN-αß) is a central mediator in the induction of hypersensitivity and is strongly expressed in splenic conventional dendritic cells (cDC) and marginal zone macrophages (MZM) when mice are infected with adenovirus. This study investigates the ability of adenoviral infection to influence the activation state of the immune system and underlines the importance of considering this state when planning the treatment of patients. METHODS: Infection with adenovirus-based vectors (Ad) or pretreatment with recombinant IFN-ß was used as a model to study hypersensitivity to lipopolysaccharide (LPS) in mice, murine macrophages, and human blood samples. The TNF-α, IL-6, IFN-αß, and IL-10 responses induced by LPS after pretreatment were measured. Mouse knockout models for MARCO, IFN-αßR, CD14, IRF3, and IRF7 were used to probe the mechanisms of the hypersensitive reaction. RESULTS: We show that, similar to TNF-α and IL-6 but not IL-10, the induction of IFN-αß by LPS increases strongly after Ad infection. This is true both in mice and in human blood samples ex vivo, suggesting that the regulatory mechanisms seen in the mouse are also present in humans. In mice, the scavenger receptor MARCO on IFN-αß-producing cDC and splenic marginal zone macrophages is important for Ad uptake and subsequent cytokine overproduction by LPS. Interestingly, not all IFN-αß-pretreated macrophage types exposed to LPS exhibit an enhanced TNF-α and IL-6 response. Pretreated alveolar macrophages and alveolar macrophage-like murine cell lines (MPI cells) show enhanced responses, while bone marrow-derived and peritoneal macrophages show a weaker response. This correlates with the respective absence or presence of the anti-inflammatory IL-10 response in these different macrophage types. In contrast, Ad or IFN-ß pretreatment enhances the subsequent induction of IFN-αß in all macrophage types. IRF3 is dispensable for the LPS-induced IFN-αß overproduction in infected MPI cells and partly dispensable in infected mice, while IRF7 is required. The expression of the LPS co-receptor CD14 is important but not absolutely required for the elicitation of a TNF-α over-response to LPS in Ad-infected mice. CONCLUSION: Viral infections or application of virus-based vaccines induces type I interferon and can tip the balance of the innate immune system in the direction of hyperreactivity to a subsequent exposure to TLR ligands. The adenoviral model presented here is one example of how multiple factors, both environmental and genetic, affect the physiological responses to pathogens. Being able to measure the current reactivity state of the immune system would have important benefits for infection-specific therapies and for the prevention of vaccination-elicited adverse effects.

Assessing Interferon Regulatory Factor 4 Complex Formation: Differential Behavior of Homocomplexes Versus Heterocomplexes Induced by Mutations.

Interferon regulatory factor 4 (IRF4) is a crucial transcription factor that plays a vital role in lymphocyte development, including in the fate-determining steps in terminal differentiation. It is also implicated in the development of lymphoid tumors such as multiple myeloma and adult T-cell leukemia. IRF4 can form a homodimer and multiple heterocomplexes with other transcription factors such as purine-rich box1 and activator protein 1. Each protein complex binds to specific DNA sequences to regulate a distinct set of genes. However, the precise relationship among these complex formations remains unclear. Herein, we investigated the abilities of IRF4 proteins with functional mutations in the IRF-association domain and autoinhibitory region to form complexes using luciferase reporter assays. The assays allowed us to selectively assess the activity of each complex. Our results revealed that certain IRF-association domain mutants, previously known to have impaired heterocomplex formation, maintained or even enhanced homodimer activity. This discrepancy suggests that the mutated amino acid residues selectively influence homodimer activity. Conversely, a phosphomimetic serine mutation in the autoinhibitory region displayed strong activating effects in all complexes. Furthermore, we observed that partner proteins involved in heterocomplex formation could disrupt the activity of the homodimer, suggesting a potential competition between homocomplexes and heterocomplexes. Our findings provide new insights into the mechanistic function of IRF4.

DDX20 positively regulates the interferon pathway to inhibit viral infection.

The DEAD-box (DDX) family comprises RNA helicases characterized by the conserved sequence D(Asp)-E(Glu)-A(Ala)-D(Asp), participating in various RNA metabolism processes. Some DDX family members have been identified for their crucial roles in viral infections. In this study, RNAi library screening of the DDX family unveiled the antiviral activity of DDX20. Knockdown of DDX20 enhanced the replication of viruses such as vesicular stomatitis virus (VSV) and herpes simplex virus type I (HSV-1), while overexpression of DDX20 significantly diminished the replication level of these viruses. Mechanistically, DDX20 elevated the phosphorylation level of IRF3 induced by external stimuli by facilitating the interaction between TBK1 and IRF3, thereby promoting the expression of IFN-ß. The increased IFN-ß production, in turn, upregulated the expression of interferon-stimulated genes (ISGs), including Cig5 and IFIT1, thereby exerting the antiviral effect. Finally, in an in vivo infection study, Ddx20 gene-deficient mice exhibited increased susceptibility to viral infection. This study provides new evidence that DDX20 positively modulates the interferon pathway and restricts viral infection.

Compound Danshen Dripping Pill effectively alleviates cGAS-STING-triggered diseases by disrupting STING-TBK1 interaction.

BACKGROUND: The cyclic GMP-AMP synthase (cGAS)-stimulator of interferon (IFN) genes (STING) pathway is critical in the innate immune system and can be mobilized by cytosolic DNA. The various inflammatory and autoimmune diseases progression is highly correlated with aberrant cGAS-STING pathway activation. While some cGAS-STING pathway inhibitor were identified, there are no drugs that can be applied to the clinic. Compound Danshen Dripping Pill (CDDP) has been successfully used in clinic around the world, but the most common application is limited to cardiovascular disease. Therefore, the purpose of the present investigation was to examine whether CDDP inhibits the cGAS-STING pathway and could be used as a therapeutic agent for multiple cGAS-STING-triggered diseases. METHODS: BMDMs, THP1 cells or Trex1-/- BMDMs were stimulated with various cGAS-STING-agonists after pretreatment with CDDP to detect the function of CDDP on IFN-ß and ISGs productionn. Next, we detect the influence on IRF3 and P65 nuclear translocation, STING oligomerization and STING-TBK1-IRF3 complex formation of CDDP. Additionally, the DMXAA-mediated activation mice model of cGAS-STING pathway was used to study the effects of CDDP. Trex1-/- mice model and HFD-mediated obesity model were established to clarify the efficacy of CDDP on inflammatory and autoimmune diseases. RESULTS: CDDP efficacy suppressed the IRF3 phosphorylation or the generation of IFN-ß, ISGs, IL-6 and TNF-α. Mechanistically, CDDP did not influence the STING oligomerization and IRF3-TBK1 and STING-IRF3 interaction, but remarkably eliminated the STING-TBK1 interaction, ultimately blocking the downstream responses. In addition, we also clarified that CDDP could suppress cGAS-STING pathway activation triggered by DMXAA, in vivo. Consistently, CDDP could alleviate multi-organ inflammatory responses in Trex1-/- mice model and attenuate the inflammatory disorders, incleding obesity-induced insulin resistance. CONCLUSION: CDDP is a specifically cGAS-STING pathway inhibitor. Furthermore, we provide novel mechanism for CDDP and discovered a clinical agent for the therapy of cGAS-STING-triggered inflammatory and autoimmune diseases.

TRIM21 reduces H1N1-induced inflammation and apoptosis by regulating the TBK1-IRF3 signaling pathway in A549 cells.

Triple motif protein 21 (TRIM21) has an antiviral function that inhibits various viral infections. However, its role in the progress of influenza A virus (IAV) infection is unclear. In this study, we investigated the role and molecular mechanism of TRIM21 in IAV infection. RT-qPCR was used to determine the level of TRIM21 mRNA, and ELISA was used to measure the levels of IFN-α, IFN-ß, IL-6, and TNF-α. The levels of the TRIM21, NP, TBK1, IRF3, p-TBK1, and p-IRF3 proteins were estimated by Western blot. The results showed that, after IAV infection, TRIM21 was upregulated in clinical patient serum and A549 cells, and this was correlated with the IFN response. Overexpression of TRIM21 reduced IAV replication and transcription in in vitro cell experiments. TRIM21 also increased IFN-α and IFN-ß levels and decreased IL-6 and TNF-α levels in A549 cells. In addition, overexpression of TRIM21 inhibited IAV-induced apoptosis. Further experiments demonstrated that TBK1-IRF3 signaling was activated by TRIM21 and was involved in the inhibitory effect of TRIM21 on virus replication. In summary, our study suggests that TRIM21 inhibits viral replication by activating the TBK1-IRF3 signaling pathway, further inhibiting the infection process of IAV.

Flavonoid extracted from Epimedium attenuate cGAS-STING-mediated diseases by targeting the formation of functional STING signalosome.

Hyperactivation of the cyclic-GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) signalling pathway has been shown to be associated with the development of a variety of inflammatory diseases, and the discovery of an inhibitor of the cGAS-STING signalling pathway holds great promise in the therapeutic interventions. Epimedium flavonoid (EF), a major active ingredient isolated from the medicinal plant Epimedium, has been reported to have good anti-inflammatory activity, but its exact mechanism of action remains unclear. In the present study, we found that EF in mouse bone marrow-derived macrophages (BMDMs), THP-1 (Tohoku Hospital Pediatrics-1) as well as in human peripheral blood mononuclear cells (hPBMC) inhibited the activation of the cGAS-STING signalling pathway, which subsequently led to a decrease in the expression of type I interferon (IFN-ß, CXCL10 and ISG15) and pro-inflammatory cytokines (IL-6 and TNF-α). Mechanistically, EF does not affect STING oligomerization, but inhibits the formation of functional STING signalosome by attenuating the interaction of interferon regulatory factor 3 (IRF3) with STING and TANK-binding kinase 1 (TBK1). Importantly, in vivo experiments, EF has shown promising therapeutic effects on inflammatory diseases mediated by the cGAS-STING pathway, which include the agonist model induced by DMXAA stimulation, the autoimmune inflammatory disease model induced by three prime repair exonuclease 1 (Trex1) deficiency, and the non-alcoholic steatohepatitis (NASH) model induced by a pathogenic amino acid and choline deficiency diet (MCD). To summarize, our study suggests that EF is a potent potential inhibitor component of the cGAS-STING signalling pathway for the treatment of inflammatory diseases mediated by the cGAS-STING signalling pathway.

Cordyceps sinensis extract protects against acute kidney injury by inhibiting perforin expression in NK cells via the STING/IRF3 pathway.

Acute kidney injury (AKI) is associated with immune cell activation and inflammation. However, the putative pathogenic mechanisms of this injury have not been thoroughly investigated. Natural killer (NK) cells play an important role in immune regulation; however, whether NK cells regulate AKI remains unclear. Cordyceps sinensis (CS), a modern Chinese patented medicine preparation, has been widely used in treating patients with chronic kidney disease (CKD) owing to its anti-inflammatory effects and maintenance of immune homeostasis. Whether 2'-deoxyadenosine, a major active component in CS, can ameliorate renal AKI by regulating immunity, particularly in NK cells, has not been reported. This study is the first to demonstrate how NK cells promote AKI by releasing perforin, interferon-gamma (IFN-γ) and other inflammatory factors in vivo and in vitro. Differential gene expression between AKI and normal tissues was assessed using bioinformatic analyses. Quantitative real-time PCR, western blotting, and immunohistochemical staining were used to detect target protein mRNA and protein expression. Levels of inflammatory factors were measured using enzyme-linked immunosorbent assay. We found the high doses of the 2'-deoxyadenosine treatment significantly alleviated FA-induced renal damage in vivo, and alleviated the NK cells of renal injury by activating the STING/IRF3 pathway to inhibit perforin release in vitro. The results showed that 2'-deoxyadenosine could mitigate AKI by downregulating the activity of NK cells (by decreasing the expressions of perforin and IFN-γ) and inhibiting the stimulator of interferon genes and phosphorylated IFN regulatory factor 3. This may provide valuable evidence supporting the clinical use of CS in treating patients with AKI.

Interaction between the SFTSV envelope glycoprotein Gn and STING inhibits the formation of the STING-TBK1 complex and suppresses the NF-κB signaling pathway.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is an emerging tick-borne bunyavirus with high pathogenicity. There has been a gradual increase in the number of reported cases in recent years, with high morbidity and mortality rates. The cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) signaling pathway plays an important role in the innate immune defense activated by viral infection; however, the role of the cGAS-STING signaling pathway during SFTSV infection is still unclear. In this study, we investigated the relationship between SFTSV infection and cGAS-STING signaling. We found that SFTSV infection caused the release of mitochondrial DNA into the cytoplasm and inhibits downstream innate immune signaling pathways by activating the cytoplasmic DNA receptor cGAS. We found that the SFTSV envelope glycoprotein Gn was a potent inhibitor of the cGAS-STING pathway and blocked the nuclear accumulation of interferon regulatory factor 3 and p65 to inhibit downstream innate immune signaling. Gn of SFTSV interacted with STING to inhibit STING dimerization and inhibited K27-ubiquitin modification of STING to disrupt the assembly of the STING-TANK-binding kinase 1 complex and downstream signaling. In addition, Gn was found to be involved in inducing STING degradation, further inhibiting the downstream immune response. In conclusion, this study identified the important role of the glycoprotein Gn in the antiviral innate immune response and revealed a novel mechanism of immune escape for SFTSV. Moreover, this study increases the understanding of the pathogenic mechanism of SFTSV and provides new insights for further treatment of SFTS. IMPORTANCE: Severe fever with thrombocytopenia syndrome virus (SFTSV) is a newly discovered virus associated with severe hemorrhagic fever in humans. However, the role of the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) signaling pathway during SFTSV infection is still unclear. We found that SFTSV infection inhibits downstream innate immune signaling pathways by activating the cytoplasmic DNA receptor cGAS. In addition, SFTSV Gn blocked the nuclear accumulation of interferon regulatory factor 3 and p65 to inhibit downstream innate immune signaling. Moreover, we determined that Gn of SFTSV inhibited K27-ubiquitin modification of STING to disrupt the assembly of the STING-TANK-binding kinase 1 complex and downstream signaling. We found that the SFTSV envelope glycoprotein Gn is a potent inhibitor of the cGAS-STING pathway. In conclusion, this study highlights the crucial function of the glycoprotein Gn in the antiviral innate immune response and reveals a new method of immune escape of SFTSV.

The African Swine Fever Virus Virulence Determinant DP96R Suppresses Type I IFN Production Targeting IRF3.

DP96R of African swine fever virus (ASFV), also known as uridine kinase (UK), encodes a virulence-associated protein. Previous studies have examined DP96R along with other genes in an effort to create live attenuated vaccines. While experiments in pigs have explored the impact of DP96R on the pathogenicity of ASFV, the precise molecular mechanism underlying this phenomenon remains unknown. Here, we describe a novel molecular mechanism by which DP96R suppresses interferon regulator factor-3 (IRF3)-mediated antiviral immune responses. DP96R interacts with a crucial karyopherin (KPNA) binding site within IRF3, disrupting the KPNA-IRF3 interaction and consequently impeding the translocation of IRF3 to the nucleus. Under this mechanistic basis, the ectopic expression of DP96R enhances the replication of DNA and RNA viruses by inhibiting the production of IFNs, whereas DP96R knock-down resulted in higher IFNs and IFN-stimulated gene (ISG) transcription during ASFV infection. Collectively, these findings underscore the pivotal role of DP96R in inhibiting IFN responses and increase our understanding of the relationship between DP96R and the virulence of ASFV.