RESUMO
The integration of HIV-1 DNA into the host genome results in single-strand gaps and 2-nt overhangs at the ends of viral DNA, which must be repaired by cellular enzymes. The cellular factors responsible for the DNA damage repair in HIV-1 DNA integration have not yet been well defined. We report here that HIV-1 infection potently activates the Fanconi anemia (FA) DNA repair pathway, and the FA effector proteins FANCI-D2 bind to the C-terminal domain of HIV-1 integrase. Knockout of FANCI blocks productive viral DNA integration and inhibits the replication of HIV-1. Finally, we show that the knockout of DNA polymerases or flap nuclease downstream of FANCI-D2 reduces the levels of integrated HIV-1 DNA, suggesting these enzymes may be responsible for the repair of DNA damages induced by viral DNA integration. These experiments reveal that HIV-1 exploits the FA pathway for the stable integration of viral DNA into host genome.
Assuntos
Anemia de Fanconi , HIV-1 , Dano ao DNA , Reparo do DNA , DNA Viral/genética , DNA Viral/metabolismo , Anemia de Fanconi/genética , Anemia de Fanconi/metabolismo , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/metabolismo , Proteínas de Grupos de Complementação da Anemia de Fanconi/genética , Proteínas de Grupos de Complementação da Anemia de Fanconi/metabolismo , HIV-1/genética , HIV-1/metabolismo , Humanos , UbiquitinaçãoRESUMO
Retinoic acid-inducible gene I (RIG-I) is a pattern recognition receptor and is involved in the innate immune response against RNA viruses infection. Here, we demonstrate that the Ras-GTPase-activating protein SH3-domain-binding protein 1 (G3BP1) serves as a positive regulator of the RIG-I-mediated signaling pathway. G3BP1-deficient cells inhibited RNA virus-triggered induction of downstream antiviral genes. Furthermore, we found that G3BP1 inhibited the replication of Sendai virus and vesicular stomatitis virus, indicating a positive regulation of G3BP1 to cellular antiviral responses. Mechanistically, G3BP1 formed a complex with RNF125 and RIG-I, leading to decreased RNF125 via its auto-ubiquitination; thus, promoting expression of RIG-I. Overall, the results suggest a novel mechanism for G3BP1 in the positive regulation of antiviral signaling mediated by RIG-I.
Assuntos
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 , Infecções por Vírus de RNA/genética , Ubiquitina-Proteína Ligases/genética , Proteína DEAD-box 58/imunologia , Regulação da Expressão Gênica/imunologia , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata/genética , Infecções por Vírus de RNA/imunologia , Infecções por Vírus de RNA/virologia , Vírus de RNA/genética , Vírus de RNA/imunologia , Vírus de RNA/patogenicidade , Receptores Imunológicos , Receptores de Reconhecimento de Padrão/genética , Transdução de Sinais , Ubiquitina-Proteína Ligases/imunologia , Ubiquitinação/genética , Ubiquitinação/imunologia , Replicação Viral/genéticaRESUMO
TANK-binding kinase 1 (TBK1) is essential for interferon beta (IFN-ß) production and innate antiviral immunity. However, other, additional functions of TBK1 have remained elusive. Here, we showed that TBK1 is an E3 ubiquitin ligase that undergoes self-ubiquitylation in vitro in the presence of the E2 enzyme UbcH5c. Further evidence showed that TBK1 could also be self-ubiquitylated in vivo Importantly, multiple picornavirus VP3 proteins were degraded by TBK1 through its kinase and E3 ubiquitin ligase activity. Mechanistically, TBK1 phosphorylated multiple picornavirus VP3 proteins at serine residues and ubiquitinated them via K63-linked ubiquitination at lysine residues. In addition, the C426 and C605 residues of TBK1 were not essential for TBK1 innate immunity activity; however, these residues were required for degradation of multiple picornavirus VP3 proteins and for its E3 ubiquitin ligase activity. Hence, our findings identified a novel role of TBK1 in regulating the virus life cycle and provided new insights into the molecular mechanisms of TBK1-mediated antiviral response.IMPORTANCE TBK1 is an important adaptor protein required for innate immune response to viruses, but its other functions were unknown. In this study, we found that TBK1 is an E3 ubiquitin ligase that undergoes self-ubiquitylation in vitro in the presence of the E2 enzyme UbcH5c. In addition, multiple picornavirus VP3 proteins were degraded by TBK1 through its kinase and E3 ubiquitin ligase activity. Our report provides evidence that TBK1 plays a role in viral protein degradation.
Assuntos
Proteínas do Capsídeo/metabolismo , Picornaviridae/metabolismo , Proteólise , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação , Proteínas Virais/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Imunidade Inata , Lisina/metabolismo , Mutação , Fosforilação , Proteínas Serina-Treonina Quinases , Alinhamento de Sequência , Ubiquitina-Proteína Ligases/genéticaRESUMO
The components of foot-and-mouth disease virus (FMDV) interact with host cellular proteins to promote self-replication and evade the host immune response. Previous studies have shown that FMDV 3A, 2C and 2B proteins interact with host cellular proteins involved in FMDV replication. However, whether the other host proteins have an impact on FMDV replication is less understood. In this study, we identified DDX56 as a positive regulator of FMDV replication. DDX56 overexpression increased FMDV replication, whereas DDX56 knockdown had the opposite effect. DDX56 interacted and cooperated with FMDV 3A to inhibit the type I interferon by reducing the phosphorylation of IRF3. Moreover, the D166 site of DDX56 played a role in increasing FMDV replication and cooperating with FMDV 3A to inhibit the phosphorylation of IRF3. Additionally, knockdown of DDX56 or FMDV 3A results also showed that DDX56 cooperated with FMDV 3A to inhibit the phosphorylation of IRF3. These results suggest that the interaction between FMDV 3A protein and the host protein DDX56 is critical for FMDV replication.
Assuntos
RNA Helicases DEAD-box/metabolismo , Vírus da Febre Aftosa/fisiologia , Febre Aftosa/virologia , Interações entre Hospedeiro e Microrganismos , Fator Regulador 3 de Interferon/metabolismo , Proteínas não Estruturais Virais/metabolismo , Replicação Viral , Animais , Células HEK293 , Hepatócitos , Humanos , Interferon Tipo I/metabolismo , Fosforilação , Ligação Proteica , SuínosRESUMO
Foot-and-mouth disease virus (FMDV) causes a highly contagious and debilitating disease in cloven-hoofed animals, which leads to devastating economic consequences. Previous studies have reported that some FMDV proteins can interact with host proteins to affect FMDV replication. However, the influence of the interactions between FMDV VP0 protein and its partners on FMDV replication remains unknown. In this study, we found that the overexpression of poly (rC) binding protein 2 (PCBP2) promoted FMDV replication, whereas the knockdown of PCBP2 suppressed FMDV replication. Furthermore, PCBP2 can interact with FMDV VP0 protein to promote the degradation of VISA via the apoptotic pathway. Further studies demonstrated that FMDV VP0 protein enhanced the formation of the PCBP2-VISA complex. Ultimately, we found that the degradation of VISA was weaker in PCBP2-knockdown and FMDV VP0-overexpressing cells, or FMDV VP0-knockdown cells than in the control cells. Summarily, our data revealed that the interaction between PCBP2 and VP0 could promote FMDV replication via the apoptotic pathway.
Assuntos
Vírus da Febre Aftosa/fisiologia , Proteínas de Ligação a RNA/metabolismo , Proteínas Virais/metabolismo , Replicação Viral/fisiologia , Animais , Apoptose , Linhagem Celular , Interferon beta/metabolismo , Camundongos , Ligação Proteica , Proteólise , Transdução de Sinais , SuínosRESUMO
Transcription factor IRF3-mediated type I interferon induction plays a role in antiviral innate immunity. However, mechanisms for the control and regulation of IRF3 nuclear import remain largely unknown. We have identified DEAD box polypeptide 56 (DDX56) as a negative regulator of virus-triggered IFN-ß induction. Overexpression of DDX56 suppressed nuclear translocation of IRF3 via disrupting the IRF3-IOP5 interaction, whereas knockdown or knockout of DDX56 had the opposite effect. In addition, the interaction between DDX56 and IRF3 increased during viral infection. We further found that the D166 site of DDX56 was essential for inhibiting IRF3 import into the nucleus. Our findings suggest that DDX56 regulates antiviral innate immunity by inhibiting the nuclear translocation of IRF3, revealing a novel mechanism of the DDX56-mediated innate antiviral response.This article has an associated First Person interview with the first author of the paper.
