Toll-Like Receptor 3 Is Involved in Detection of Enterovirus A71 Infection and Targeted by Viral 2A Protease

Enterovirus A71 (EV-A71) has emerged as a major pathogen causing hand, foot, and mouth disease, as well as neurological disorders. The host immune response affects the outcomes of EV-A71 infection, leading to either resolution or disease progression. However, the mechanisms of how the mammalian innate immune system detects EV-A71 infection to elicit antiviral immunity remain elusive. Here, we report that the Toll-like receptor 3 (TLR3) is a key viral RNA sensor for sensing EV-A71 infection to trigger antiviral immunity. Expression of TLR3 in HEK293 cells enabled the cells to sense EV-A71 infection, leading to type I, IFN-mediated antiviral immunity. Viral double-stranded RNA derived from EV-A71 infection was a key ligand for TLR3 detection. Silencing of TLR3 in mouse and human primary immune cells impaired the activation of IFN-β upon EV-A71 infection, thus reinforcing the importance of the TLR3 pathway in defending against EV-A71 infection. Our results further demonstrated that TLR3 was a target of EV-A71 infection. EV-A71 protease 2A was implicated in the downregulation of TLR3. Together, our results not only demonstrate the importance of the TLR3 pathway in response to EV-A71 infection, but also reveal the involvement of EV-A71 protease 2A in subverting TLR3-mediated antiviral defenses.

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DDX3 directly facilitates IKKα activation and regulates downstream signalling pathways

Biochemical Journal ◽

10.1042/bcj20180163 ◽

2018 ◽

Vol 475 (22) ◽

pp. 3595-3607 ◽

Cited By ~ 5

Author(s):

Anthony Fullam ◽

Lili Gu ◽

Yvette Höhn ◽

Martina Schröder

Keyword(s):

Nuclear Factor Κb ◽

Innate Immune ◽

Type I Interferons ◽

Signalling Pathways ◽

Type I ◽

Nuclear Factor ◽

Toll Like Receptor ◽

Disease States ◽

Dead Box ◽

Iκb Kinase

DDX3 is a DEAD-box RNA helicase that we and others have previously implicated in antiviral immune signalling pathways leading to type I interferon (IFN) induction. We previously demonstrated that it directly interacts with the kinase IKKε (IκB kinase ε), enhances it activation, and then facilitates phosphorylation of the transcription factor IRF3 by IKKε. However, the TLR7/9 (Toll-like receptor 7/9)-mediated pathway, one of the most physiologically relevant IFN induction pathways, proceeds independently of IKKε or the related kinase TBK1 (TANK-binding kinase 1). This pathway induces type I IFN production via the kinases NIK (NF-κB-inducing kinase) and IKKα and is activated when plasmacytoid dendritic cells sense viral nucleic acids. In the present study, we demonstrate that DDX3 also directly interacts with IKKα and enhances its autophosphorylation and -activation. Modulation of DDX3 expression consequently affected NIK/IKKα-mediated IRF7 phosphorylation and induction of type I interferons. In addition, alternative NF-κB (nuclear factor-κB) activation, another pathway regulated by NIK and IKKα, was also down-regulated in DDX3 knockdown cells. This substantially broadens the effects of DDX3 in innate immune signalling to pathways beyond TBK1/IKKε and IFN induction. Dysregulation of these pathways is involved in disease states, and thus, our research might implicate DDX3 as a potential target for their therapeutic manipulation.

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Toll-Like Receptor 7 Agonist GS-9620 Induces HIV Expression and HIV-Specific Immunity in Cells from HIV-Infected Individuals on Suppressive Antiretroviral Therapy

Journal of Virology ◽

10.1128/jvi.02166-16 ◽

2017 ◽

Vol 91 (8) ◽

Cited By ~ 60

Author(s):

Angela Tsai ◽

Alivelu Irrinki ◽

Jasmine Kaur ◽

Tomas Cihlar ◽

George Kukolj ◽

...

Keyword(s):

Antiretroviral Therapy ◽

Mononuclear Cells ◽

Antiviral Immunity ◽

Type I ◽

Toll Like Receptor ◽

Peripheral Blood Mononuclear ◽

Effector Functions ◽

Latently Infected ◽

Infected Cells ◽

Latent Hiv

ABSTRACT Antiretroviral therapy can suppress HIV replication to undetectable levels but does not eliminate latent HIV, thus necessitating lifelong therapy. Recent efforts to target this persistent reservoir have focused on inducing the expression of latent HIV so that infected cells may be recognized and eliminated by the immune system. Toll-like receptor (TLR) activation stimulates antiviral immunity and has been shown to induce HIV from latently infected cells. Activation of TLR7 leads to the production of several stimulatory cytokines, including type I interferons (IFNs). In this study, we show that the selective TLR7 agonist GS-9620 induced HIV in peripheral blood mononuclear cells (PBMCs) from HIV-infected individuals on suppressive antiretroviral therapy. GS-9620 increased extracellular HIV RNA 1.5- to 2-fold through a mechanism that required type I IFN signaling. GS-9620 also activated HIV-specific T cells and enhanced antibody-mediated clearance of HIV-infected cells. Activation by GS-9620 in combination with HIV peptide stimulation increased CD8 T cell degranulation, production of intracellular cytokines, and cytolytic activity. T cell activation was again dependent on type I IFNs produced by plasmacytoid dendritic cells. GS-9620 induced phagocytic cell maturation and improved effector-mediated killing of HIV-infected CD4 T cells by the HIV envelope-specific broadly neutralizing antibody PGT121. Collectively, these data show that GS-9620 can activate HIV production and improve the effector functions that target latently infected cells. GS-9620 may effectively complement orthogonal therapies designed to stimulate antiviral immunity, such as therapeutic vaccines or broadly neutralizing antibodies. Clinical studies are under way to determine if GS-9620 can target HIV reservoirs. IMPORTANCE Though antiretroviral therapies effectively suppress viral replication, they do not eliminate integrated proviral DNA. This stable intermediate of viral infection is persistently maintained in reservoirs of latently infected cells. Consequently, lifelong therapy is required to maintain viral suppression. Ultimately, new therapies that specifically target and eliminate the latent HIV reservoir are needed. Toll-like receptor agonists are potent enhancers of innate antiviral immunity that can also improve the adaptive immune response. Here, we show that a highly selective TLR7 agonist, GS-9620, activated HIV from peripheral blood mononuclear cells isolated from HIV-infected individuals with suppressed infection. GS-9620 also improved immune effector functions that specifically targeted HIV-infected cells. Previously published studies on the compound in other chronic viral infections show that it can effectively induce immune activation at safe and tolerable clinical doses. Together, the results of these studies suggest that GS-9620 may be useful for treating HIV-infected individuals on suppressive antiretroviral therapy.

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Stages of Meningococcal Sepsis Simulated In Vitro, with Emphasis on Complement and Toll-Like Receptor Activation

Infection and Immunity ◽

10.1128/iai.00195-08 ◽

2008 ◽

Vol 76 (9) ◽

pp. 4183-4189 ◽

Cited By ~ 32

Author(s):

Bernt Christian Hellerud ◽

Jørgen Stenvik ◽

Terje Espevik ◽

John D. Lambris ◽

Tom Eirik Mollnes ◽

...

Keyword(s):

Inflammatory Response ◽

Complement Activation ◽

Bacterial Load ◽

Receptor Activation ◽

Innate Immune ◽

Cytokine Secretion ◽

Hek293 Cells ◽

Toll Like Receptor ◽

Bacterial Concentration

ABSTRACT The clinical presentation of meningococcal disease is closely related to the number of meningococci in the circulation. This study aimed to examine the activation of the innate immune system after being exposed to increasing and clinically relevant concentrations of meningococci. We incubated representative Neisseria meningitidis serogroup B (ST-32) and serogroup C (ST-11) strains and a lipopolysaccharide (LPS)-deficient mutant (the 44/76 lpxA mutant) in human serum and whole blood and measured complement activation and cytokine secretion and the effect of blocking these systems. HEK293 cells transfected with Toll-like receptors (TLRs) were examined for activation of NF-κB. The threshold for cytokine secretion and activation of NF-κB was 103 to 104 meningococci/ml. LPS was the sole inflammation-inducing molecule at concentrations up to 105 to 106 meningococci/ml. The activation was dependent on TLR4-MD2-CD14. Complement contributed to the inflammatory response at ≥105 to 106 meningococci/ml, and complement activation increased exponentially at ≥107 bacteria/ml. Non-LPS components initiated TLR2-mediated activation at ≥107 bacteria/ml. As the bacterial concentration exceeded 107/ml, TLR4 and TLR2 were increasingly activated, independent of CD14. In this model mimicking human disease, the inflammatory response to N. meningitidis was closely associated with the bacterial concentration. Therapeutically, CD14 inhibition alone was most efficient at a low bacterial concentration, whereas addition of a complement inhibitor may be beneficial when the bacterial load increases.

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Genome-encoded cytoplasmic double-stranded RNAs, found in C9ORF72 ALS-FTD brain, propagate neuronal loss

Science Translational Medicine ◽

10.1126/scitranslmed.aaz4699 ◽

2021 ◽

Vol 13 (601) ◽

pp. eaaz4699

Author(s):

Steven Rodriguez ◽

Asli Sahin ◽

Benjamin R. Schrank ◽

Hawra Al-Lawati ◽

Isabel Costantino ◽

...

Keyword(s):

Neuronal Loss ◽

Innate Immune ◽

Neural Cells ◽

Type I ◽

Double Stranded Rna ◽

Repeat Sequences ◽

Innate Immune Signaling ◽

Human Neural Cells ◽

Human Brains ◽

Lateral Sclerosis

Triggers of innate immune signaling in the CNS of patients with amyotrophic lateral sclerosis and frontotemporal degeneration (ALS/FTD) remain elusive. We report the presence of cytoplasmic double-stranded RNA (cdsRNA), an established trigger of innate immunity, in ALS-FTD brains carrying C9ORF72 intronic hexanucleotide expansions that included genomically encoded expansions of the G4C2 repeat sequences. The presence of cdsRNA in human brains was coincident with cytoplasmic TAR DNA binding protein 43 (TDP-43) inclusions, a pathologic hallmark of ALS/FTD. Introducing cdsRNA into cultured human neural cells induced type I interferon (IFN-I) signaling and death that was rescued by FDA-approved JAK inhibitors. In mice, genomically encoded dsRNAs expressed exclusively in a neuronal class induced IFN-I and death in connected neurons non–cell-autonomously. Our findings establish that genomically encoded cdsRNAs trigger sterile, viral-mimetic IFN-I induction and propagated death within neural circuits and may drive neuroinflammation and neurodegeneration in patients with ALS/FTD.

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Toll-Like Receptor-3 Ligation-Induced Indoleamine 2, 3-Dioxygenase Expression in Human Trophoblasts

Endocrinology ◽

10.1210/en.2011-0278 ◽

2011 ◽

Vol 152 (12) ◽

pp. 4984-4992 ◽

Cited By ~ 19

Author(s):

Bo Wang ◽

Kaori Koga ◽

Yutaka Osuga ◽

Ingrid Cardenas ◽

Gentaro Izumi ◽

...

Keyword(s):

T Cells ◽

Mrna Expression ◽

Conditioned Media ◽

Poly I:C ◽

Type I ◽

Toll Like Receptor ◽

Human Trophoblast ◽

Double Stranded Rna ◽

Indoleamine 2 ◽

Polycytidylic Acid

Indoleamine 2,3-dioxygenase (IDO) is an enzyme that degrades an essential amino acid, tryptophan, and plays a role in inhibiting the proliferation of T cells and intracellular pathogens. Inhibiting IDO in mice leads to fetal rejection, suggesting its significance in establishing pregnancy. Toll-like receptor 3 (TLR-3) is a key component of the innate immune system that recognizes viral double-stranded RNA and triggers immune reactions by producing type I interferon. Using a human trophoblast cell culture system, we studied the effect of TLR-3 ligation on IDO expression and function by treating trophoblasts with polyinosinic-polycytidylic acid [poly(I:C)] (a synthetic double stranded RNA, which mimics viral RNA). Real-time PCR and Western blot analysis revealed that IDO mRNA and protein expression was significantly induced by poly(I:C). The activity of IDO was also increased by poly(I:C) given that the l-kynurenine concentrations were elevated in conditioned media. Conditioned media from poly(I:C)-treated trophoblasts were found to inhibit the proliferation of human T cells significantly. Poly(I:C) was also shown to induce interferon (IFN)-β mRNA expression in trophoblasts. Recombinant human IFN-β increased IDO mRNA expression in trophoblasts more rapidly than poly(I:C). Pretreating with neutralizing antibody against IFN-β significantly suppressed IDO induction by poly(I:C). Collectively we have demonstrated that ligation of TLR-3 by poly(I:C) induces IDO expression in human first-trimester trophoblasts via an IFN-β-dependent pathway. These findings suggest that upon viral infection, trophoblasts induce IDO and in turn contribute to antimicrobial activity and maintenance of fetomaternal tolerance.

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LGP2 binds to PACT to regulate RIG-I– and MDA5-mediated antiviral responses

Science Signaling ◽

10.1126/scisignal.aar3993 ◽

2019 ◽

Vol 12 (601) ◽

pp. eaar3993 ◽

Cited By ~ 8

Author(s):

Raul Y. Sanchez David ◽

Chantal Combredet ◽

Valérie Najburg ◽

Gael A. Millot ◽

Guillaume Beauclair ◽

...

Keyword(s):

Rna Silencing ◽

Rna Binding ◽

Inflammatory Responses ◽

Innate Immune ◽

Type I ◽

Double Stranded Rna ◽

Rna Molecules ◽

Protein Protein Interaction ◽

Antiviral Responses ◽

Inducible Gene

The retinoic acid–inducible gene I (RIG-I)–like receptors (RLRs) RIG-I, MDA5, and LGP2 stimulate inflammatory and antiviral responses by sensing nonself RNA molecules produced during viral replication. Here, we investigated how LGP2 regulates the RIG-I– and MDA5-dependent induction of type I interferon (IFN) signaling and showed that LGP2 interacted with different components of the RNA-silencing machinery. We identified a direct protein-protein interaction between LGP2 and the IFN-inducible, double-stranded RNA binding protein PACT. The LGP2-PACT interaction was mediated by the regulatory C-terminal domain of LGP2 and was necessary for inhibiting RIG-I–dependent responses and for amplifying MDA5-dependent responses. We described a point mutation within LGP2 that disrupted the LGP2-PACT interaction and led to the loss of LGP2-mediated regulation of RIG-I and MDA5 signaling. These results suggest a model in which the LGP2-PACT interaction regulates the inflammatory responses mediated by RIG-I and MDA5 and enables the cellular RNA-silencing machinery to coordinate with the innate immune response.

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Plasmacytoid dendritic cell–derived type I interferon is crucial for the adjuvant activity of Toll-like receptor 7 agonists

Blood ◽

10.1182/blood-2009-08-238543 ◽

2010 ◽

Vol 115 (10) ◽

pp. 1949-1957 ◽

Cited By ~ 82

Author(s):

Deepa Rajagopal ◽

Carine Paturel ◽

Yannis Morel ◽

Satoshi Uematsu ◽

Shizuo Akira ◽

...

Keyword(s):

T Lymphocytes ◽

Cytotoxic T Lymphocytes ◽

Immune Activation ◽

Plasmacytoid Dendritic Cell ◽

Innate Immune ◽

Type I ◽

Toll Like Receptor ◽

Adjuvant Activity ◽

Innate Immune Activation ◽

Tlr7 Agonist

Abstract There is a high demand for the development of adjuvants that induce cytotoxic T lymphocytes, which are crucial for the elimination of intracellular pathogens and tumor cells. Toll-like receptor (TLR) agonists are prime candidates to fulfill this role because they induce innate immune activation and promote adaptive immune responses. The successful application of the TLR7 agonist R837 for treatment of basal cell carcinoma shows the potential for exploiting this pathway in tumor immunotherapy. Imidazoquinolines like R837 and stimulatory ssRNA oligonucleotides both trigger TLR7-mediated immune activation, but little is known about their comparative ability to promote immunity induction. We investigated differences in innate immune activation and adjuvant activity between the imidazoquinoline R848 and the ssRNA TLR7 agonist polyUs21. In contrast to R848, polyUs21 induced detectable levels of intracellular interferon-α (IFN-α) in plasmacytoid dendritic cells (PDCs). In immunization studies, only polyUs21 led to robust priming of type 1 T helper cells and cytotoxic T lymphocytes, and it was more efficient in inducing antitumor immunity than R848. Notably, exogenous IFN-α augmented the adjuvant activity of R848, whereas depletion of PDC abrogated the adjuvanticity of polyUs21. This study, therefore, identifies sufficient IFN-α production by PDC as an important determinant of vaccine efficacy.

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Disruption of MDA5-Mediated Innate Immune Responses by the 3C Proteins of Coxsackievirus A16, Coxsackievirus A6, and Enterovirus D68

Journal of Virology ◽

10.1128/jvi.00546-17 ◽

2017 ◽

Vol 91 (13) ◽

Cited By ~ 24

Author(s):

Yajuan Rui ◽

Jiaming Su ◽

Hong Wang ◽

Junliang Chang ◽

Shaohua Wang ◽

...

Keyword(s):

Immune Responses ◽

Respiratory Disease ◽

Foot And Mouth Disease ◽

Innate Immune ◽

Type I ◽

Type I Ifn ◽

Innate Immune Responses ◽

Key Factors ◽

Coxsackievirus A16 ◽

Enterovirus D68

ABSTRACT Coxsackievirus A16 (CV-A16), CV-A6, and enterovirus D68 (EV-D68) belong to the Picornaviridae family and are major causes of hand, foot, and mouth disease (HFMD) and pediatric respiratory disease worldwide. The biological characteristics of these viruses, especially their interplay with the host innate immune system, have not been well investigated. In this study, we discovered that the 3Cpro proteins from CV-A16, CV-A6, and EV-D68 bind melanoma differentiation-associated gene 5 (MDA5) and inhibit its interaction with MAVS. Consequently, MDA5-triggered type I interferon (IFN) signaling in the retinoic acid-inducible gene I-like receptor (RLR) pathway was blocked by the CV-A16, CV-A6, and EV-D68 3Cpro proteins. Furthermore, the CV-A16, CV-A6, and EV-D68 3Cpro proteins all cleave transforming growth factor β-activated kinase 1 (TAK1), resulting in the inhibition of NF-κB activation, a host response also critical for Toll-like receptor (TLR)-mediated signaling. Thus, our data demonstrate that circulating HFMD-associated CV-A16 and CV-A6, as well as severe respiratory disease-associated EV-D68, have developed novel mechanisms to subvert host innate immune responses by targeting key factors in the RLR and TLR pathways. Blocking the ability of 3Cpro proteins from diverse enteroviruses and coxsackieviruses to interfere with type I IFN induction should restore IFN antiviral function, offering a potential novel antiviral strategy. IMPORTANCE CV-A16, CV-A6, and EV-D68 are emerging pathogens associated with hand, foot, and mouth disease and pediatric respiratory disease worldwide. The pathogenic mechanisms of these viruses are largely unknown. Here we demonstrate that the CV-A16, CV-A6, and EV-D68 3Cpro proteins block MDA5-triggered type I IFN induction. The 3Cpro proteins of these viruses bind MDA5 and inhibit its interaction with MAVS. In addition, the CV-A16, CV-A6, and EV-D68 3Cpro proteins cleave TAK1 to inhibit the NF-κB response. Thus, our data demonstrate that circulating HFMD-associated CV-A16 and CV-A6, as well as severe respiratory disease-associated EV-D68, have developed a mechanism to subvert host innate immune responses by simultaneously targeting key factors in the RLR and TLR pathways. These findings indicate the potential merit of targeting the CV-A16, CV-A6, and EV-D68 3Cpro proteins as an antiviral strategy.

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Roles of Toll-Like Receptor 3 in Human Tumors

Frontiers in Immunology ◽

10.3389/fimmu.2021.667454 ◽

2021 ◽

Vol 12 ◽

Author(s):

Xin Zheng ◽

Song Li ◽

Hui Yang

Keyword(s):

Cell Lines ◽

Human Tumors ◽

Type I Interferons ◽

Type I ◽

Toll Like Receptor ◽

Double Stranded Rna ◽

Important Group ◽

Important Member ◽

Different Types ◽

Molecular Patterns

Toll-like receptor 3 (TLR3) is an important member of the TLR family, which is an important group of pathogen-associated molecular patterns. TLR3 can recognize double-stranded RNA and induce activation of NF-κB and the production of type I interferons. In addition to its immune-associated role, TLR3 has also been detected in some tumors. However TLR3 can play protumor or antitumor roles in different tumors or cell lines. Here, we review the basic signaling associated with TLR3 and the pro- or antitumor roles of TLR3 in different types of tumors and discuss the possible reasons for the opposing roles of TLR3 in tumors.

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Fetal and Maternal Innate Immunity Receptors Have Opposing Effects on the Severity of Experimental Malaria in Pregnancy: Beneficial Roles for Fetus-Derived Toll-Like Receptor 4 and Type I Interferon Receptor 1

Infection and Immunity ◽

10.1128/iai.00708-17 ◽

2018 ◽

Vol 86 (5) ◽

Cited By ~ 10

Author(s):

Lurdes Rodrigues-Duarte ◽

Yash Pandya ◽

Rita Neres ◽

Carlos Penha-Gonçalves

Keyword(s):

Innate Immunity ◽

Type I Interferon ◽

Parasite Burden ◽

Innate Immune ◽

Malaria In Pregnancy ◽

Type I ◽

Toll Like Receptor ◽

Toll Like Receptor 4 ◽

Interferon Receptor ◽

In Pregnancy

ABSTRACTMalaria in pregnancy (MiP) is a distinctive clinical form ofPlasmodiuminfection and is a cause of placental insufficiency leading to poor pregnancy outcomes. Maternal innate immunity responses play a decisive role in the development of placental inflammation, but the action of fetus-derived factors in MiP outcomes has been overlooked. We investigated the role of theTlr4andIfnar1genes, taking advantage of heterogenic mating strategies to dissect the effects mediated by maternally and fetally derived Toll-like receptor 4 (TLR4) or type I interferon receptor 1 (IFNAR1). Using a mouse infection system displaying severe MiP outcomes, we found that the expressions of TLR4 and IFNAR1 in the maternal compartment take part in deleterious MiP outcomes, but their fetal counterparts patently counteract these effects. We uncovered that fetal TLR4 contributes to thein vitrouptake of infected erythrocytes by trophoblasts and to the innate immune response in the placenta, offering robust protection of fetus viability, but had no sensible impact on the placental parasite burden. In contrast, we observed that the expression of IFNAR1 in the fetal compartment was associated with a reduced placental parasite burden but had little beneficial effect on fetus outcomes. Furthermore, the downregulation ofIfnar1expression in infected placentas and in trophoblasts exposed to infected erythrocytes indicated that the interferon-IFNAR1 pathway is involved in the trophoblast response to infection. This work unravels that maternal and fetal counterparts of innate immune pathways drive opposing responses in murine placental malaria and implicates the activation of innate receptors in fetal trophoblast cells in the control of placental infection and in the protection of the fetus.

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