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.

Alpha-Emitter Radium-223 Induces STING-Dependent Pyroptosis to Trigger Robust Antitumor Immunity.

Radium-223 (223 Ra) is the first-in-class alpha-emitter to mediate tumor eradication, which is commonly thought to kill tumor cells by directly cleaving double-strand DNA. However, the immunogenic characteristics and cell death modalities triggered by 223 Ra remain unclear. Here, it is reported that the 223 Ra irradiation induces the pro-inflammatory damage-associated molecular patterns including calreticulin, HMGB1, and HSP70, hallmarks of tumor immunogenicity. Moreover, therapeutic 223 Ra retards tumor progression by triggering pyroptosis, an immunogenic cell death. Mechanically, 223 Ra-induced DNA damage leads to the activation of stimulator of interferon genes (STING)-mediated DNA sensing pathway, which is critical for NLRP3 inflammasome-dependent pyroptosis and subsequent DCs maturation as well as T cell activation. These findings establish an essential role of STING in mediating alpha-emitter 223 Ra-induced antitumor immunity, which provides the basis for the development of novel cancer therapeutic strategies and combinatory therapy.

STING-targeted PET tracer for early assessment of tumor immunogenicity in colorectal cancer after chemotherapy.

PURPOSE: To optimize chemotherapy regimens and improve the effectiveness of chemotherapy combined with immunotherapy, a PET tracer specifically targeting the stimulator of interferon genes (STING), denoted as [18F]FBTA was used to monitor the early changes in tumor immunogenicity after chemotherapy in colorectal cancer (CRC) mice. METHODS: The toluene sulfonate precursor was labeled with 18F to produce the STING targeted probe-[18F]FBTA. [18F]FBTA-PET imaging and biodistribution were performed using CRC mice treated with oxaliplatin (OXA) or cisplatin (CDDP). CRC mice were also treated with low (CDDP-LD: 1 mg/kg) or medium (CDDP-MD: 2.5 mg/kg) doses of CDDP, and subjected to PET imaging and biodistribution. The effects of different chemotherapeutic agents and different doses of CDDP on tumor innate immunity were verified by flow cytometry and immunohistochemistry. RESULTS: PET imaging of CRC mice exhibited notably enhanced tumor uptake in the early phase of chemotherapy with treatment with OXA (3.09 ± 0.25%ID/g) and CDDP (4.01 ± 0.18%ID/g), especially in the CDDP group. The PET-derived tumor uptake values have strong correlations with STING immunohistochemical score. Flow cytometry showed both agents led to DCs and macrophages infiltration in tumors. Compared with OXA, CDDP treatment recruits more DCs and macrophages in CRC tumors. Both CDDP-LD and CDDP-MD treatment elevated uptake in CRC tumors, especially in CDDP-MD group. Immunohistochemistry and flow cytometry confirmed CDDP-MD treatment recruits more DCs and macrophages than CDDP-LD treatment. CONCLUSION: Overall, the STING-targeted tracer-[18F]FBTA was demonstrated to monitor early changes in tumor immunogenicity in CRC mice after chemotherapy. Besides, the STING-targeted strategy may help to select the appropriate chemotherapy regimen, including chemotherapeutic agents and doses, which further improve clinical decision making for combination immunotherapy after chemotherapy for CRC.

The role of chemerin in the regulation of cGAS-STING pathway in gestational diabetes mellitus placenta.

Recent studies already confirmed that placenta mitochondrial dysfunction is associated with the progression of gestational diabetes mellitus (GDM). Besides, a possible relationship between adipokine chemerin and disulfide-bond A oxidoreductase-like protein (DsbA-L) had been revealed, whereas the potential interaction remains unclear. In addition, very little is still known about the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) signaling pathway and its mechanisms of action in the context of GDM. The present study aims to investigate the underlying mechanism of cGAS-STING pathway and its regulatory relationship with chemerin in GDM. A total of 50 participants, including 25 cases of GDM patients and 25 pregnant women with normal glucose tolerance, were enrolled, and their placenta tissues at term labor were collected. Besides, an insulin resistance cell model was established on the human trophoblastic cell line to explore the molecular mechanism of chemerin on cGAS-STING pathway. Results showed that there were mitochondrial pathological changes in GDM placenta, accompanied by the decreased expression of DsbA-L, increased level of chemerin, and the activation of cGAS-STING pathway. In the insulin resistant cell model, overexpression of chemerin upregulated protein expression of DsbA-L, and recombinant chemerin presented time-dependent inhibition on the cGAS-STING pathway, but this effect was not dependent on DsbA-L. In conclusion, elevated chemerin is probably a protective mechanism, which may be a potential therapeutic strategy for GDM.

STING-Targeted PET Imaging for Specific Detection and Therapeutic Monitoring of Myocarditis.

Imaging strategies for the specific detection and therapeutic monitoring of myocarditis are still lacking. Stimulator of interferon genes (STING) is a signal transduction molecule involved in an innate immune response. Here, we evaluated the feasibility of the recently developed STING-targeted radiotracer [18F]FBTA for positron emission tomography (PET) imaging to detect myocardial inflammation and monitor treatment in myocarditis mice. [18F]FBTA-PET imaging was performed in myocarditis mice and normal mice to verify the specificity of [18F]FBTA for the diagnosis of myocarditis. We also performed PET imaging in mice with myocarditis treated to verify the ability of [18F]FBTA in therapeutic monitoring. The expression of STING and inflammatory cell types was confirmed by flow cytometry and immunohistochemistry. [18F]FDG-PET imaging of myocarditis was used as a contrast. [18F]FBTA-PET imaging showed that the average radioactive uptake was significantly higher in the hearts of the myocarditis group than in the control group. STING was highly overexpressed in cardiac inflammatory cells, including macrophages, dendritic cells (DCs), and T cells. However, there was no significant difference in cardiac radiotracer uptake of [18F]FDG between the myocarditis group and the control group. Moreover, cardiac uptake of [18F]FBTA was significantly reduced in cyclosporin A-treated myocarditis mice and myocardial STING expression was also significantly reduced after the treatment. Overall, we showed that a STING-targeted PET tracer [18F]FBTA can be used to monitor changes in the inflammatory microenvironment in myocarditis. Besides, [18F]FBTA-PET is also suitable for real-time monitoring of myocarditis treatment, representing a promising diagnostic and therapeutic monitoring approach for myocarditis.

STING contributes to trauma-induced heterotopic ossification through NLRP3-dependent macrophage pyroptosis.

Trauma-induced heterotopic ossification (HO) is featured by aberrant bone formation at extra-skeletal site. STING is a master adaptor protein linking cellular damage to immune responses, while its role in HO remains elusive. A murine burn/tenotomy model was used to mimic trauma-induced HO in vivo. We demonstrated elevated STING expression in macrophages in inflammatory stage after burn/tenotomy, and STING inhibition significantly alleviated HO formation. Activated NLRP3-dependent macrophage pyroptosis was also found in inflammatory stage after burn/tenotomy. Either STING or NLRP3 suppression reduced mature HO by weakening macrophage pyroptotic inflammation, while protective effects of STING were abolished by NLRP3 overexpression. Further, in vitro, we also found a prominent STING level in pyroptotic BMDMs. STING suppression relieved macrophage pyroptotic inflammation, while abolished by NLRP3 overexpression. Our results reveal that STING poses regulatory effects on trauma-induced HO formation, via modulating NLRP3-dependent macrophage pyroptosis. Targeting STING-NLRP3 axis represents an attractive approach for trauma-induced HO prevention.

Systemic Delivery of a STING Agonist-Loaded Positively Charged Liposome Selectively Targets Tumor Immune Microenvironment and Suppresses Tumor Angiogenesis.

Although stimulator of interferon genes (STING) agonists has shown great promise in preclinical studies, the clinical development of STING agonist therapy is challenged by its limited systemic delivery. Here, positively charged fusogenic liposomes loaded with a STING agonist (PoSTING) are designed for systemic delivery and to preferentially target the tumor microenvironment. When PoSTING is administered intravenously, it selectively targets not only tumor cells but also immune and tumor endothelial cells (ECs). In particular, delivery of STING agonists to tumor ECs normalizes abnormal tumor vasculatures, induces intratumoral STING activation, and elicits robust anti-tumor T cell immunity within the tumor microenvironment. Therefore, PoSTING can be used as a systemic delivery platform to overcome the limitations of using STING agonists in clinical trials.

Fluvoxamine alleviates bleomycin-induced lung fibrosis via regulating the cGAS-STING pathway.

Idiopathic pulmonary fibrosis (IPF) is a fatal disease with high mortality and limited effective therapy. Herein, we reported that fluvoxamine, a selective serotonin reuptake inhibitor (SSRI), used in depression and anxiety treatment, also exhibited therapeutic activities in IPF. Fluvoxamine inhibited cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING), restrained the activation of their downstream targets, including PERK/ eIF2α/ c-Myc/ miR-9-5p/ TBPL1 and TBK1/ YAP/ JNK1/2/ Bnip3/ CaMKII/ cofilin signaling, thus attenuated the activation and migration of fibroblasts upon TGF-ß1 challenge. Fluvoxamine dose-dependently improved pulmonary function, decreased the expression of inflammatory factors, reduced excessive production of extracellular matrix, and thus alleviated bleomycin (BLM)-induced lung fibrosis in mice. Moreover, fluvoxamine at a dose of 10 mg/ kg showed similar efficacy as pirfenidone (PFD) at a dose of 30 mg/kg in a mice model of lung fibrosis. In summary, our results suggest that fluvoxamine is an effective anti-fibrotic agent for IPF.

Antibody drug conjugates beyond cytotoxic payloads.

For many years, antibody drug conjugates (ADC) have teased with the promise of targeted payload delivery to diseased cells, embracing the targeting of the antibody to which a cytotoxic payload is conjugated. During the past decade this promise has started to be realised with the approval of more than a dozen ADCs for the treatment of various cancers. Of these ADCs, brentuximab vedotin really laid the foundations of a template for a successful ADC with lysosomal payload release from a cleavable dipeptide linker, measured DAR by conjugation to the Cys-Cys interchain bonds of the antibody and a cytotoxic payload. Using this ADC design model oncology has now expanded their repertoire of payloads to include non-cytotoxic compounds. These new payload classes have their origins in prior medicinal chemistry programmes aiming to design selective oral small molecule drugs. While this may not have been achieved, the resulting compounds provide excellent starting points for ADC programmes with some compounds amenable to immediate linker attachment while for others extensive SAR and structural information offer invaluable design insights. Many of these new oncology payload classes are of interest to other therapeutic areas facilitating rapid access to drug-linkers for exploration as non-oncology ADCs. Other therapeutic areas have also pursued unique payload classes with glucocorticoid receptor modulators (GRM) being the most clinically advanced in immunology. Here, ADC payloads come full circle, as oncology is now investigating GRM payloads for the treatment of cancer. This chapter aims to cover all these new ADC approaches while describing the medicinal chemistry origins of the new non-cytotoxic payloads.

Neoantigen Immunotherapeutic-Gel Combined with TIM-3 Blockade Effectively Restrains Orthotopic Hepatocellular Carcinoma Progression.

Herein, we integrate the Hepa1-6 liver cancer-specific neoantigen, toll-like receptor 9 agonist and stimulator of interferon genes agonist by silk-hydrogel package, and combine with TIM-3 blockade to elicit robust antitumor immunity for effectively suppressing orthotopic hepatocellular carcinoma (HCC) progression. Unlike intradermal injection of simple mixed components with short-term immune protection, the neoantigen immunotherapeutic-gels evoke long-term immune protection to achieve significant prophylactic and therapeutic activity against HCC through only one-shot administration without any side effects. Notably, the synergized immunotherapy by further combining NGC-gels with TIM-3 antibody significantly reduces regulatory T-cells and increases the IFN-γ and IL-12p70 levels in tumor tissues for promoting the infiltration of IFN-γ+CD8+T-cells and 41BB+CD8+T-cells to achieve complete remission (4/7) and prevent pulmonary metastasis in orthotopic HCC, and establish long-term memory against tumor rechallenge with remarkably longer survival time (180 days). Overall, this study provides an attractive and promising synergistic strategy for HCC immunotherapy with possible clinical translation prospects.

Radiation Induces Bone Microenvironment Disruption by Activating the STING-TBK1 Pathway.

Background and Objectives: Damage to normal bone tissue following therapeutic irradiation (IR) represents a significant concern, as IR-induced bone microenvironment disruption can cause bone loss and create a more favorable environment for tumor metastases. The aim of the present study was to explore the cellular regulatory mechanism of IR-induced bone microenvironment disruption to effectively prevent radiotherapy-associated adverse effects in the future. Materials and Methods: In this study, a mouse model of local IR was established via local irradiation of the left hind limb of BALB/c mice with 12 Gy X-rays, and an in vitro osteocyte (OCY) model was established by exposing osteocyte-like MLO-Y4 cells to 2, 4, and 8 Gy irradiation to analyze multicellular biological injuries and cellular senescence. Small interfering RNA (siRNA) transfection at the cellular level and a selective antagonist intervention C-176 at the animal level were used to explore the potential role of the stimulator of interferon genes (STING) on IR-induced bone microenvironment disruption. Results: The results showed that 12 Gy local IR induces multicellular dysfunction, manifested as ascension of OCYs exfoliation, activation of osteoclastogenesis, degeneration of osteogenesis and fate conversion of adipogenesis, as well as cellular senescence and altered senescence-associated secretory phenotype (SASP) secretion. Furthermore, the expression of STING was significantly elevated, both in the primary OCYs harvested from locally irradiated mice and in vitro irradiated MLO-Y4 cells, accompanied by the markedly upregulated levels of phosphorylated TANK-binding kinase 1 (P-TBK1), RANKL and sclerostin (SOST). STING-siRNA transfection in vitro restored IR-induced upregulated protein expression of P-TBK1 and RANKL, as well as the mRNA expression levels of inflammatory cytokines, such as IL-1α, IL-6 and NF-κB, accompanied by the alleviation of excessive osteoclastogenesis. Finally, administration of the STING inhibitor C-176 mitigated IR-induced activation of osteoclastogenesis and restraint of osteogenesis, ameliorating the IR-induced biological damage of OCYs, consistent with the inhibition of P-TBK1, RANKL and SOST. Conclusions: The STING-P-TBK1 signaling pathway plays a crucial role in the regulation of the secretion of inflammatory cytokines and osteoclastogenesis potential in IR-induced bone microenvironment disruption. The selective STING antagonist can be used to intervene to block the STING pathway and, thereby, repair IR-induced multicellular biological damage and mitigate the imbalance between osteoclastogenesis and osteoblastgenesis.

COVID-19 Molecular Pathophysiology: Acetylation of Repurposing Drugs.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) induces immune-mediated type 1 interferon (IFN-1) production, the pathophysiology of which involves sterile alpha motif and histidine-aspartate domain-containing protein 1 (SAMHD1) tetramerization and the cytosolic DNA sensor cyclic-GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) signaling pathway. As a result, type I interferonopathies are exacerbated. Aspirin inhibits cGAS-mediated signaling through cGAS acetylation. Acetylation contributes to cGAS activity control and activates IFN-1 production and nuclear factor-κB (NF-κB) signaling via STING. Aspirin and dapsone inhibit the activation of both IFN-1 and NF-κB by targeting cGAS. We define these as anticatalytic mechanisms. It is necessary to alleviate the pathologic course and take the lag time of the odds of achieving viral clearance by day 7 to coordinate innate or adaptive immune cell reactions.

BioID screening of biotinylation sites using the avidin-like protein Tamavidin 2-REV identifies global interactors of stimulator of interferon genes (STING).

Stimulator of interferon genes (STING) mediates cytosolic DNA-induced innate immune signaling via membrane trafficking. The global identification of proteins that spatiotemporally interact with STING will provide a better understanding of its trafficking mechanisms and of STING signaling pathways. Proximity-dependent biotin identification (BioID) is a powerful technology to identify physiologically relevant protein-protein interactions in living cells. However, biotinylated peptides are rarely detected in the conventional BioID method, which uses streptavidin beads to pull down biotinylated proteins, because the biotin-streptavidin interaction is too strong. As a result, only nonbiotinylated peptides are identified, which cannot be distinguished from peptides of nonspecifically pull-downed proteins. Here, we developed a simple method to efficiently and specifically enrich biotinylated peptides using Tamavidin 2-REV, an engineered avidin-like protein with reversible biotin-binding capability. Using RAW264.7 macrophages stably expressing TurboID-fused STING, we identified and quantified >4,000 biotinylated peptides of STING-proximal proteins. Various endoplasmic reticulum-associated proteins were biotinylated in unstimulated cells, and STING activation caused biotinylation of many proteins located in the Golgi and endosomes. These proteins included those known to interact with activated STING, such as TANK-binding kinase 1 (TBK1), several palmitoyl transferases, and p62/sequestosome 1 (SQSTM1). Furthermore, interferon-induced transmembrane protein 3 (IFITM3), an endolysosome-localized antiviral protein, bound to STING at the late activation stage. These dynamic interaction profiles will provide detailed insights into STING signaling; we propose that our approach using Tamavidin 2-REV would be useful for BioID-based and other biotinylation-based peptide identification methods.

STING agonist and IDO inhibitor combination therapy inhibits tumor progression in murine models of colorectal cancer.

Despite impressive clinical success, cancer immunotherapy based on immune checkpoint blockade remains ineffective in colorectal cancer (CRC). Stimulator of interferon genes (STING) is a novel potential target and STING agonists have shown potential anti-tumor efficacy. Combined therapy based on synergistic mechanism can overcome the resistance. However, STING agonists-based combination therapies are deficient. We designed different immunotherapy combinations, including STING agonist, indoleamine 2,3 dioxygenase (IDO) inhibitor and PD-1 blockade, with purpose of exploring which option can effectively inhibit CRC growth. To further explore the possible reasons of therapeutic effectiveness, we observed the combination therapy in C57BL/6Tmem173gt mice. Our findings demonstrated that STING agonist diABZI combined with IDO inhibitor 1-MT significantly inhibited tumor growth, even better than the three-drug combination, promoted the recruitment of CD8+ T cells and dendritic cells, and decreased the infiltration of myeloid-derived suppressor cells. We conclude that diABZI combined with 1-MT is a promising option for CRC.

Long non-coding RNA MALAT1 targeting STING transcription promotes bronchopulmonary dysplasia through regulation of CREB.

Bronchopulmonary dysplasia (BPD) is a severe complication of preterm infants characterized by increased alveolarization and inflammation. Premature exposure to hyperoxia is believed to be a key contributor to the pathogenesis of BPD. No effective preventive or therapeutic agents have been created. Stimulator of interferon gene (STING) is associated with inflammation and apoptosis in various lung diseases. Long non-coding RNA MALAT1 has been reported to be involved in BPD. However, how MALAT1 regulates STING expression remains unknown. In this study, we assessed that STING and MALAT1 were up-regulated in the lung tissue from BPD neonates, hyperoxia-based rat models and lung epithelial cell lines. Then, using the flow cytometry and cell proliferation assay, we found that down-regulating of STING or MALAT1 inhibited the apoptosis and promoted the proliferation of hyperoxia-treated cells. Subsequently, qRT-PCR, Western blotting and dual-luciferase reporter assays showed that suppressing MALAT1 decreased the expression and promoter activity of STING. Moreover, transcription factor CREB showed its regulatory role in the transcription of STING via a chromatin immunoprecipitation. In conclusion, MALAT1 interacts with CREB to regulate STING transcription in BPD neonates. STING, CREB and MALAT1 may be promising therapeutic targets in the prevention and treatment of BPD.

Nanomedicine-mediated alteration of the pharmacokinetic profile of small molecule cancer immunotherapeutics.

The advent of immunotherapy is a game changer in cancer therapy with monoclonal antibody- and T cell-based therapeutics being the current flagships. Small molecule immunotherapeutics might offer advantages over the biological drugs in terms of complexity, tissue penetration, manufacturing cost, stability, and shelf life. However, small molecule drugs are prone to rapid systemic distribution, which might induce severe off-target side effects. Nanotechnology could aid in the formulation of the drug molecules to improve their delivery to specific immune cell subsets. In this review we summarize the current efforts in changing the pharmacokinetic profile of small molecule immunotherapeutics with a strong focus on Toll-like receptor agonists. In addition, we give our vision on limitations and future pathways in the route of nanomedicine to the clinical practice.

Stimulator of interferon genes (STING) provides insect antiviral immunity by promoting Dredd caspase-mediated NF-κB activation.

The antiviral cGMP-AMP (cGAMP)-stimulator of interferon genes (STING) pathway is well characterized in mammalian cells. However, whether this pathway also plays a role in insect antiviral immunity is unknown. In this study, we found that cGAMP is produced in silkworm (Bombyx mori) cells infected with nucleopolyhedrovirus (NPV). In searches for STING-related sequences, we identified BmSTING, a potential cGAMP sensor in B. mori We observed that BmSTING overexpression effectively inhibits NPV replication in silkworm larvae, whereas dsRNA-mediated BmSTING knockdown resulted in higher viral load. Cleavage and nuclear translocation of BmRelish, a NF-κB-related transcription factor, was also observed when BmSTING was overexpressed and was enhanced by cGAMP stimulation or viral infection of B. mori larvae. Moreover, we identified a caspase-8-like protein (BmCasp8L) as a BmSTING-interacting molecule and as a suppressor of BmSTING-mediated BmRelish activation. Interestingly, cGAMP stimulation decreased BmCasp8L binding to BmSTING and increased BmRelish activity. Of note, an interaction between death-related ced-3/Nedd2-like caspase (BmDredd) and BmSTING promoted BmRelish cleavage for efficient antiviral signaling and protection of insect cells from viral infection. Our findings have uncovered BmSTING as a critical mediator of antiviral immunity in the model insect B. mori and have identified several BmSTING-interacting proteins that control antiviral defenses.

STING positively regulates human ORMDL3 expression through TBK1-IRF3-STAT6 complex mediation.

Orosomucoid 1-like protein 3 (ORMDL3) is an asthma candidate gene associated with virus-triggered recurrent wheeze. Stimulator of interferon gene (STING) controls TLR-independent cytosolic responses to viruses. However, the association of STING with ORMDL3 is unclear. Here, we have shown that ORMDL3 expression shows a linear correlation with STING in recurrent wheeze patients. In elucidating the molecular mechanisms of the ORMDL3-STING relationship, we found that STING promoted the transcriptional activity of ORMDL3, which was significantly associated with increased levels of interferon regulatory factor 3 (IRF3) and signal transducer and activator of transcription 6 (STAT6). Further study showed that via activation of TANK binding kinase 1 (TBK1), STING enhanced the phosphorylation and binding of IRF3 and STAT6, which upregulated ORMDL3 by binding to the promoter. Our results showed that STING positively regulated ORMDL3 through the TBK1-IRF3-STAT6 complex.

Disease-associated mutations identify a novel region in human STING necessary for the control of type I interferon signaling.

BACKGROUND: Gain-of-function mutations in transmembrane protein 173 (TMEM173) encoding stimulator of interferon genes (STING) underlie a recently described type I interferonopathy called STING-associated vasculopathy with onset in infancy (SAVI). OBJECTIVES: We sought to define the molecular and cellular pathology relating to 3 individuals variably exhibiting the core features of the SAVI phenotype including systemic inflammation, destructive skin lesions, and interstitial lung disease. METHODS: Genetic analysis, conformational studies, in vitro assays and ex vivo flow-cytometry were performed. RESULTS: Molecular and in vitro data demonstrate that the pathology in these patients is due to amino acid substitutions at positions 206, 281, and 284 of the human STING protein. These mutations confer cGAMP-independent constitutive activation of type I interferon signaling through TBK1 (TANK-binding kinase), independent from the alternative STING pathway triggered by membrane fusion of enveloped RNA viruses. This constitutive activation was abrogated by ex vivo treatment with the janus kinase 1/2 inhibitor ruxolitinib. CONCLUSIONS: Structural analysis indicates that the 3 disease-associated mutations at positions 206, 281, and 284 of the STING protein define a novel cluster of amino acids with functional importance in the regulation of type I interferon signaling.