RESUMO
A characteristic feature of COVID-19, the disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, is the dysregulated immune response with impaired type I and III interferon (IFN) expression and an overwhelming inflammatory cytokine storm. RIG-I-like receptors (RLRs) and cGAS-STING signaling pathways are responsible for sensing viral infection and inducing IFN production to combat invading viruses. Multiple proteins of SARS-CoV-2 have been reported to modulate the RLR signaling pathways to achieve immune evasion. Although SARS-CoV-2 infection also activates the cGAS-STING signaling by stimulating micronuclei formation during the process of syncytia, whether SARS-CoV-2 modulates the cGAS-STING pathway requires further investigation. Here, we screened 29 SARS-CoV-2-encoded viral proteins to explore the viral proteins that affect the cGAS-STING signaling pathway and found that SARS-CoV-2 open reading frame 10 (ORF10) targets STING to antagonize IFN activation. Overexpression of ORF10 inhibits cGAS-STING-induced interferon regulatory factor 3 phosphorylation, translocation, and subsequent IFN induction. Mechanistically, ORF10 interacts with STING, attenuates the STING-TBK1 association, and impairs STING oligomerization and aggregation and STING-mediated autophagy; ORF10 also prevents the endoplasmic reticulum (ER)-to-Golgi trafficking of STING by anchoring STING in the ER. Taken together, these findings suggest that SARS-CoV-2 ORF10 impairs the cGAS-STING signaling by blocking the translocation of STING and the interaction between STING and TBK1 to antagonize innate antiviral immunity.
Assuntos
COVID-19 , Interferon Tipo I , Autofagia , Humanos , Imunidade Inata , Interferon Tipo I/genética , Interferons , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Nucleotidiltransferases/genética , Fases de Leitura Aberta , Proteínas Serina-Treonina Quinases/genética , SARS-CoV-2 , Proteínas Virais/metabolismoRESUMO
As a highly pathogenic human coronavirus, SARS-CoV-2 has to counteract an intricate network of antiviral host responses to establish infection and spread. The nucleic acid-induced stress response is an essential component of antiviral defense and is closely related to antiviral innate immunity. However, whether SARS-CoV-2 regulates the stress response pathway to achieve immune evasion remains elusive. In this study, SARS-CoV-2 NSP5 and N protein were found to attenuate antiviral stress granule (avSG) formation. Moreover, NSP5 and N suppressed IFN expression induced by infection of Sendai virus or transfection of a synthetic mimic of dsRNA, poly (I:C), inhibiting TBK1 and IRF3 phosphorylation, and restraining the nuclear translocalization of IRF3. Furthermore, HEK293T cells with ectopic expression of NSP5 or N protein were less resistant to vesicular stomatitis virus infection. Mechanistically, NSP5 suppressed avSG formation and disrupted RIG-I-MAVS complex to attenuate the RIG-I-mediated antiviral immunity. In contrast to the multiple targets of NSP5, the N protein specifically targeted cofactors upstream of RIG-I. The N protein interacted with G3BP1 to prevent avSG formation and to keep the cofactors G3BP1 and PACT from activating RIG-I. Additionally, the N protein also affected the recognition of dsRNA by RIG-I. This study revealed the intimate correlation between SARS-CoV-2, the stress response, and innate antiviral immunity, shedding light on the pathogenic mechanism of COVID-19.
Assuntos
Proteases 3C de Coronavírus/genética , Proteínas do Nucleocapsídeo de Coronavírus/genética , Proteína DEAD-box 58/genética , DNA Helicases/genética , Proteínas de Ligação a Poli-ADP-Ribose/genética , RNA Helicases/genética , Proteínas com Motivo de Reconhecimento de RNA/genética , Proteínas de Ligação a RNA/genética , Receptores Imunológicos/genética , SARS-CoV-2/genética , Grânulos de Estresse/genética , Animais , Chlorocebus aethiops , Proteases 3C de Coronavírus/imunologia , Proteínas do Nucleocapsídeo de Coronavírus/imunologia , Proteína DEAD-box 58/imunologia , DNA Helicases/imunologia , Regulação da Expressão Gênica , Células HEK293 , Células HeLa , Humanos , Evasão da Resposta Imune , Fosfoproteínas/genética , Fosfoproteínas/imunologia , Poli I-C/farmacologia , Proteínas de Ligação a Poli-ADP-Ribose/imunologia , Ligação Proteica , RNA Helicases/imunologia , Proteínas com Motivo de Reconhecimento de RNA/imunologia , RNA de Cadeia Dupla/genética , RNA de Cadeia Dupla/imunologia , Proteínas de Ligação a RNA/imunologia , Receptores Imunológicos/imunologia , SARS-CoV-2/imunologia , SARS-CoV-2/patogenicidade , Vírus Sendai/genética , Vírus Sendai/imunologia , Transdução de Sinais , Grânulos de Estresse/efeitos dos fármacos , Grânulos de Estresse/imunologia , Grânulos de Estresse/virologia , Células Vero , Vesiculovirus/genética , Vesiculovirus/imunologiaRESUMO
The global epidemic caused by the coronavirus severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has resulted in the infection of over 200 million people. To extend the knowledge of interactions between SARS-CoV-2 and humans, we systematically investigate the interactome of 29 viral proteins in human cells by using an antibody-based TurboID assay. In total, 1,388 high-confidence human proximal proteins with biotinylated sites are identified. Notably, we find that SARS-CoV-2 manipulates the antiviral and immune responses. We validate that the membrane protein ITGB1 associates angiotensin-converting enzyme 2 (ACE2) to mediate SARS-CoV-2 entry. Moreover, we reveal that SARS-CoV-2 proteins inhibit activation of the interferon pathway through the mitochondrial protein mitochondrial antiviral-signaling protein (MAVS) and the methyltransferase SET domain containing 2, histone lysine methyltransferase (SETD2). We propose 111 potential drugs for the clinical treatment of coronavirus disease 2019 (COVID-19) and identify three compounds that significantly inhibit the replication of SARS-CoV-2. The proximity labeling map of SARS-CoV-2 and humans provides a resource for elucidating the mechanisms of viral infection and developing drugs for COVID-19 treatment.
Assuntos
Anticorpos/imunologia , Antivirais/imunologia , SARS-CoV-2/imunologia , Enzima de Conversão de Angiotensina 2/imunologia , Antivirais/farmacologia , COVID-19/imunologia , Humanos , Integrina beta1/imunologia , Testes de Sensibilidade Microbiana , Tratamento Farmacológico da COVID-19RESUMO
The suppression of types I and III interferon (IFN) responses by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contributes to the pathogenesis of coronavirus disease 2019 (COVID-19). The strategy used by SARS-CoV-2 to evade antiviral immunity needs further investigation. Here, we reported that SARS-CoV-2 ORF9b inhibited types I and III IFN production by targeting multiple molecules of innate antiviral signaling pathways. SARS-CoV-2 ORF9b impaired the induction of types I and III IFNs by Sendai virus and poly (I:C). SARS-CoV-2 ORF9b inhibited the activation of types I and III IFNs induced by the components of cytosolic dsRNA-sensing pathways of RIG-I/MDA5-MAVS signaling, including RIG-I, MDA-5, MAVS, TBK1, and IKKε, rather than IRF3-5D, which is the active form of IRF3. SARS-CoV-2 ORF9b also suppressed the induction of types I and III IFNs by TRIF and STING, which are the adaptor protein of the endosome RNA-sensing pathway of TLR3-TRIF signaling and the adaptor protein of the cytosolic DNA-sensing pathway of cGAS-STING signaling, respectively. A mechanistic analysis revealed that the SARS-CoV-2 ORF9b protein interacted with RIG-I, MDA-5, MAVS, TRIF, STING, and TBK1 and impeded the phosphorylation and nuclear translocation of IRF3. In addition, SARS-CoV-2 ORF9b facilitated the replication of the vesicular stomatitis virus. Therefore, the results showed that SARS-CoV-2 ORF9b negatively regulates antiviral immunity and thus facilitates viral replication. This study contributes to our understanding of the molecular mechanism through which SARS-CoV-2 impairs antiviral immunity and provides an essential clue to the pathogenesis of COVID-19.
Assuntos
Proteína DEAD-box 58/imunologia , Evasão da Resposta Imune/genética , Interferons/imunologia , Nucleotidiltransferases/imunologia , Receptores Imunológicos/imunologia , SARS-CoV-2/imunologia , Receptor 3 Toll-Like/imunologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Proteínas Adaptadoras de Transporte Vesicular/genética , Proteínas Adaptadoras de Transporte Vesicular/imunologia , Animais , Chlorocebus aethiops , Proteínas do Nucleocapsídeo de Coronavírus/genética , Proteínas do Nucleocapsídeo de Coronavírus/imunologia , Proteína DEAD-box 58/genética , Regulação da Expressão Gênica , Células HEK293 , Células HeLa , Humanos , Quinase I-kappa B/genética , Quinase I-kappa B/imunologia , Imunidade Inata , Fator Regulador 3 de Interferon/genética , Fator Regulador 3 de Interferon/imunologia , Helicase IFIH1 Induzida por Interferon/genética , Helicase IFIH1 Induzida por Interferon/imunologia , Interferons/genética , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Nucleotidiltransferases/genética , Fosfoproteínas/genética , Fosfoproteínas/imunologia , Plasmídeos/química , Plasmídeos/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Receptores Imunológicos/genética , SARS-CoV-2/genética , SARS-CoV-2/patogenicidade , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Receptor 3 Toll-Like/genética , Transfecção , Células Vero , Replicação Viral/imunologiaRESUMO
Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has quickly spread worldwide and has affected more than 10 million individuals. A typical feature of COVID-19 is the suppression of type I and III interferon (IFN)-mediated antiviral immunity. However, the molecular mechanism by which SARS-CoV-2 evades antiviral immunity remains elusive. Here, we reported that the SARS-CoV-2 membrane (M) protein inhibits the production of type I and III IFNs induced by the cytosolic dsRNA-sensing pathway mediated by RIG-I/MDA-5-MAVS signaling. In addition, the SARS-CoV-2 M protein suppresses type I and III IFN induction stimulated by SeV infection or poly (I:C) transfection. Mechanistically, the SARS-CoV-2 M protein interacts with RIG-I, MAVS, and TBK1, thus preventing the formation of the multiprotein complex containing RIG-I, MAVS, TRAF3, and TBK1 and subsequently impeding the phosphorylation, nuclear translocation, and activation of IRF3. Consequently, ectopic expression of the SARS-CoV-2 M protein facilitates the replication of vesicular stomatitis virus. Taken together, these results indicate that the SARS-CoV-2 M protein antagonizes type I and III IFN production by targeting RIG-I/MDA-5 signaling, which subsequently attenuates antiviral immunity and enhances viral replication. This study provides insight into the interpretation of SARS-CoV-2-induced antiviral immune suppression and illuminates the pathogenic mechanism of COVID-19.
Assuntos
COVID-19/metabolismo , Proteína DEAD-box 58/metabolismo , Interferon Tipo I/biossíntese , Helicase IFIH1 Induzida por Interferon/metabolismo , Interferons/biossíntese , SARS-CoV-2/metabolismo , Transdução de Sinais , Proteínas da Matriz Viral/metabolismo , Animais , COVID-19/genética , Chlorocebus aethiops , Proteína DEAD-box 58/genética , Células HEK293 , Células HeLa , Humanos , Interferon Tipo I/genética , Helicase IFIH1 Induzida por Interferon/genética , Interferons/genética , Receptores Imunológicos , SARS-CoV-2/genética , Células Vero , Proteínas da Matriz Viral/genética , Interferon lambdaAssuntos
COVID-19/genética , Proteínas do Nucleocapsídeo/isolamento & purificação , SARS-CoV-2/genética , Proteínas Virais/isolamento & purificação , Animais , COVID-19/virologia , Núcleo Celular/ultraestrutura , Núcleo Celular/virologia , Chlorocebus aethiops , Citoplasma/ultraestrutura , Citoplasma/virologia , Humanos , Proteínas do Nucleocapsídeo/genética , Pandemias , SARS-CoV-2/patogenicidade , Células Vero/virologia , Proteínas Virais/genéticaRESUMO
The ongoing outbreak of a new coronavirus (2019-nCoV, or severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2]) has caused an epidemic of the acute respiratory syndrome known as coronavirus disease (COVID-19) in humans. SARS-CoV-2 rapidly spread to multiple regions of China and multiple other countries, posing a serious threat to public health. The spike (S) proteins of SARS-CoV-1 and SARS-CoV-2 may use the same host cellular receptor, angiotensin-converting enzyme 2 (ACE2), for entering host cells. The affinity between ACE2 and the SARS-CoV-2 S protein is much higher than that of ACE2 binding to the SARS-CoV S protein, explaining why SARS-CoV-2 seems to be more readily transmitted from human to human. Here, we report that ACE2 can be significantly upregulated after infection of various viruses, including SARS-CoV-1 and SARS-CoV-2, or by the stimulation with inflammatory cytokines such as interferons. We propose that SARS-CoV-2 may positively induce its cellular entry receptor, ACE2, to accelerate its replication and spread; high inflammatory cytokine levels increase ACE2 expression and act as high-risk factors for developing COVID-19, and the infection of other viruses may increase the risk of SARS-CoV-2 infection. Therefore, drugs targeting ACE2 may be developed for the future emerging infectious diseases caused by this cluster of coronaviruses.