RESUMO
The conventional view posits that E3 ligases function primarily through conjugating ubiquitin (Ub) to their substrate molecules. We report here that RIPLET, an essential E3 ligase in antiviral immunity, promotes the antiviral signaling activity of the viral RNA receptor RIG-I through both Ub-dependent and -independent manners. RIPLET uses its dimeric structure and a bivalent binding mode to preferentially recognize and ubiquitinate RIG-I pre-oligomerized on dsRNA. In addition, RIPLET can cross-bridge RIG-I filaments on longer dsRNAs, inducing aggregate-like RIG-I assemblies. The consequent receptor clustering synergizes with the Ub-dependent mechanism to amplify RIG-I-mediated antiviral signaling in an RNA-length dependent manner. These observations show the unexpected role of an E3 ligase as a co-receptor that directly participates in receptor oligomerization and ligand discrimination. It also highlights a previously unrecognized mechanism by which the innate immune system measures foreign nucleic acid length, a common criterion for self versus non-self nucleic acid discrimination.
Assuntos
Imunidade Inata , RNA de Cadeia Dupla/imunologia , Transdução de Sinais/imunologia , Ubiquitina-Proteína Ligases/imunologia , Ubiquitina/imunologia , Células A549 , Animais , Proteína DEAD-box 58/imunologia , Células HEK293 , Humanos , Camundongos , Receptores ImunológicosRESUMO
Antiviral lignin was produced by acidic microwave glycerolysis of sugarcane bagasse. The lignin exhibited antiviral activity against nonenveloped (encephalomyocarditis virus (EMCV) and Theiler's murine encephalomyelitis virus (TMEV)) and enveloped (vesicular stomatitis virus (VSV), Sindbis virus (SINV), and Newcastle disease virus (NDV)) viruses. A series of lignins with different antiviral activities were prepared by reacting bagasse at 140, 160, 180, and 200 °C to analyze the antiviral mechanism. No difference in ζ-potential was observed among the lignin preparations; however, the lignin prepared at 200 °C (FR200) showed the strongest anti-EMCV activity, smallest hydrodynamic diameter, highest hydrophilicity, and highest affinity for EMCV. FR200 inhibited viral propagation through contact with the virion at the attachment stage to host cells, and the EMCV RNA was intact after treatment. Therefore, the lignin inhibits viral entry to host cells through interactions with the capsid surface. The nonvolatile antiviral substance is potentially useful for preventing the spread of viruses in human living and livestock breeding environments.
Assuntos
Cardiovirus , Saccharum , Animais , Antivirais/farmacologia , Celulose/farmacologia , Vírus da Encefalomiocardite/genética , Humanos , Lignina/farmacologia , Camundongos , Micro-OndasRESUMO
Nucleic acids carrying pathogen-associated molecular patterns trigger innate immune responses and are used to activate host immunity. Although synthetic nucleic acids have been used for that purpose, they have shown limitations for in vivo and clinical applications. To address this issue, we tested a naturally occurring dsRNA extracted from rice bran (rb-dsRNA) and characterized it as a potent ligand of TLR3 and MDA5. In this study, intranasal administration of rb-dsRNA induced production of type I IFNs by alveolar macrophages and protected mice from morbidity and mortality resulting from respiratory virus infection, such as influenza A virus. This protection was completely absent in mice lacking both TRIF and MDA5, indicating the essential role of TLR3- and MDA5-dependent pathways. Interestingly, IFNAR1-deficient mice retained residual antiviral protection, which was abolished by pharmacological inhibition of caspase 1, but not IL-1ß signaling. In fact, rb-dsRNA activated caspase 1 via TRIF, resulting in the release of IL-1ß and LDH. In addition to the direct antiviral activity, rb-dsRNA modulated the immune cell population in the lungs by repopulating virus-depleted alveolar macrophages. Our data demonstrate that rb-dsRNA orchestrates IFN-dependent and -independent direct antiviral protection and that it is a potent immune stimulator modulating antiviral immunity in the lungs. These findings open doors to a range of precise immune-modulating studies and therapeutic options.
Assuntos
Antivirais/isolamento & purificação , Vírus da Influenza A/imunologia , Interferon Tipo I/imunologia , Infecções por Orthomyxoviridae/imunologia , Oryza/genética , RNA de Cadeia Dupla/imunologia , RNA de Cadeia Dupla/isolamento & purificação , Proteínas Adaptadoras de Transporte Vesicular/deficiência , Proteínas Adaptadoras de Transporte Vesicular/genética , Animais , Antivirais/imunologia , Inibidores de Caspase/administração & dosagem , Imunidade Inata , Interferon Tipo I/biossíntese , Helicase IFIH1 Induzida por Interferon/química , Helicase IFIH1 Induzida por Interferon/deficiência , Helicase IFIH1 Induzida por Interferon/genética , Interleucina-1beta/antagonistas & inibidores , Interleucina-1beta/metabolismo , Ligantes , Pulmão/imunologia , Pulmão/virologia , Macrófagos Alveolares/efeitos dos fármacos , Macrófagos Alveolares/imunologia , Camundongos , Infecções por Orthomyxoviridae/prevenção & controle , Oryza/química , Plantas/química , Plantas/genética , RNA de Cadeia Dupla/administração & dosagem , RNA de Cadeia Dupla/farmacologia , Receptor de Interferon alfa e beta/deficiência , Transdução de Sinais/efeitos dos fármacos , Receptor 3 Toll-Like/químicaRESUMO
RIG-I triggers antiviral responses by recognizing viral RNA (vRNA) in the cytoplasm. However, the spatio-temporal dynamics of vRNA sensing and signal transduction remain elusive. We investigated the time course of events in cells infected with Newcastle disease virus (NDV), a non-segmented negative-strand RNA virus. RIG-I was recruited to viral replication complexes (vRC) and triggered minimal primary type I interferon (IFN) production. RIG-I subsequently localized to antiviral stress granules (avSG) induced after vRC formation. The inhibition of avSG attenuated secondary IFN production, suggesting avSG as a platform for efficient vRNA detection. avSG selectively captured positive-strand vRNA, and poly(A)+ RNA induced IFN production. Further investigations suggested that uncapped vRNA derived from read-through transcription was sensed by RIG-I in avSG. These results highlight how viral infections stimulate host stress responses, thereby selectively recruiting uncapped vRNA to avSG, in which RIG-I and other components cooperate in an efficient antiviral program.
Assuntos
RNA Helicases DEAD-box/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Proteína DEAD-box 58 , Humanos , Vírus da Influenza A/genética , Fator Regulador 3 de Interferon/metabolismo , Interferon Tipo I/metabolismo , Interferon beta/efeitos dos fármacos , Interferon beta/genética , Camundongos , Vírus da Doença de Newcastle/genética , RNA Viral/efeitos dos fármacos , Receptores Imunológicos , Estresse FisiológicoRESUMO
RIG-I-like receptors (RLRs), protein kinase R (PKR), and endosomal Toll-like receptor 3 (TLR3) sense viral non-self RNA and are involved in cell fate determination. However, the mechanisms by which intracellular RNA induces apoptosis, particularly the role of each RNA sensor, remain unclear. We performed cytoplasmic injections of different types of RNA and elucidated the molecular mechanisms underlying viral dsRNA-induced apoptosis. The results obtained revealed that short 5'-triphosphate dsRNA, the sole ligand of RIG-I, induced slow apoptosis in a fraction of cells depending on IRF-3 transcriptional activity and IFN-I production. However, intracellular long dsRNA was sensed by PKR and TLR3, which activate distinct signals, and synergistically induced rapid apoptosis. PKR essentially induced translational arrest, resulting in reduced levels of cellular FLICE-like inhibitory protein and functioned in the TLR3/TRIF-dependent activation of caspase 8. The present results demonstrated that PKR and TLR3 were both essential for inducing the viral RNA-mediated apoptosis of infected cells and the arrest of viral production.
Assuntos
Antivirais , Receptor 3 Toll-Like , Antivirais/farmacologia , Apoptose , Interferon beta/genética , RNA de Cadeia Dupla/genética , RNA Viral/metabolismo , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , eIF-2 Quinase/genética , eIF-2 Quinase/metabolismoRESUMO
Type I interferon (IFN) production by the proper activation of nucleic acid sensors is essential for hosts to eliminate invading viruses. Among these sensors, RIG-I-like receptors (RLRs) are well-known viral RNA sensors in the cytoplasm that recognize the nonself signatures of viral RNAs to trigger IFN responses. Recent accumulating evidence has clarified that some specific and atypical self-RNAs also cause activation of RLRs independently of virus infection. Importantly, when RLR-activation by these RNAs or a conformational change via missense mutations is sustained, the resulting continuous production of type I IFN will lead to autoimmune disorders. We, herein, focus on autoimmune diseases caused by chronic activation of RLRs and discuss possible mechanisms of their onset.
Assuntos
Doenças Autoimunes/imunologia , Doenças Autoimunes/fisiopatologia , Proteína DEAD-box 58/imunologia , Interferon Tipo I/imunologia , Animais , Proteína DEAD-box 58/genética , Humanos , Interferon Tipo I/biossíntese , Helicase IFIH1 Induzida por Interferon/genética , Helicase IFIH1 Induzida por Interferon/imunologia , Polimorfismo de Nucleotídeo Único/genética , Polimorfismo de Nucleotídeo Único/imunologia , Receptores ImunológicosRESUMO
BACKGROUND/AIMS: The Model for End-Stage Liver Disease (MELD) consists of serum bilirubin and creatinine levels, International Normalized Ratio (INR) for prothrombin time, and etiology of liver disease. The MELD score is a reliable measurement of mortality risk and is suitable for a disease severity index in patients with end-stage liver disease. We examined the validity of the MELD as a disease severity index for patients with end-stage liver disease. METHODS: We investigated the 379 patients with liver cirrhosis hospitalized between January 1995 and May 2001. We retrospectively reviewed the hospital records to verify the diagnosis of cirrhosis and to collect exact patient information about their demographic data, portal hypertensive complications and laboratory data. The ability to classify patients with liver cirrhosis according to their risk of death was examined using the concordance c-statistic. RESULTS: The MELD score performed well in predicting death within 3 months with a c-statistic of 0.73 with etiology and 0.71 without etiology. The significant clinical, laboratory variables on 3 month survival in patients with liver cirrhosis are serum bilirubin, ascites and hepatic encephalopathy. The addition of portal hypertensive complications to the MELD score did not improve the accuracy of the MELD score. CONCLUSIONS: The MELD score is a useful disease severity index for the patients with end-stage liver disease and provides reliable measurement of short term survival over a wide range of liver disease severity and diverse etiology.