Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 1.752
Filtrar
1.
Signal Transduct Target Ther ; 5(1): 221, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-33024073
2.
Nat Commun ; 11(1): 3810, 2020 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-32733001

RESUMO

The pandemic of COVID-19 has posed an unprecedented threat to global public health. However, the interplay between the viral pathogen of COVID-19, SARS-CoV-2, and host innate immunity is poorly understood. Here we show that SARS-CoV-2 induces overt but delayed type-I interferon (IFN) responses. By screening 23 viral proteins, we find that SARS-CoV-2 NSP1, NSP3, NSP12, NSP13, NSP14, ORF3, ORF6 and M protein inhibit Sendai virus-induced IFN-ß promoter activation, whereas NSP2 and S protein exert opposite effects. Further analyses suggest that ORF6 inhibits both type I IFN production and downstream signaling, and that the C-terminus region of ORF6 is critical for its antagonistic effect. Finally, we find that IFN-ß treatment effectively blocks SARS-CoV-2 replication. In summary, our study shows that SARS-CoV-2 perturbs host innate immune response via both its structural and nonstructural proteins, and thus provides insights into the pathogenesis of SARS-CoV-2.


Assuntos
Betacoronavirus/fisiologia , Infecções por Coronavirus/virologia , Evasão da Resposta Imune , Interferon Tipo I/metabolismo , Pneumonia Viral/virologia , Transdução de Sinais , Betacoronavirus/genética , Betacoronavirus/imunologia , Betacoronavirus/metabolismo , Linhagem Celular , Infecções por Coronavirus/imunologia , Humanos , Imunidade Inata , Interferon beta/genética , Interferon beta/metabolismo , Interferon beta/farmacologia , Mutação , Fases de Leitura Aberta , Pandemias , Pneumonia Viral/imunologia , Regiões Promotoras Genéticas , Transdução de Sinais/efeitos dos fármacos , Proteínas Virais/genética , Proteínas Virais/metabolismo , Replicação Viral/efeitos dos fármacos
3.
Virology ; 548: 117-123, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32838932

RESUMO

The matrix (M) protein of vesicular stomatitis virus (VSV) plays a key role in immune evasion. While VSV has been thought to suppress the interferon (IFN) response primarily by inhibiting host cell transcription and translation, our recent findings indicate that the M protein also targets NF-κB activation. Therefore, the M protein may utilize two distinct mechanisms to limit expression of antiviral genes, inhibiting both host gene expression and NF-κB activation. Here we characterize a recently reported mutation in the M protein [M(D52G)] of VSV isolate 22-20, which suppressed IFN mRNA and protein production despite activating NF-κB. 22-20 inhibited reporter gene expression from multiple promoters, suggesting that 22-20 suppressed the IFN response via M-mediated inhibition of host cell transcription. We propose that suppression of the IFN response and regulation of NF-κB are independent, genetically separable functions of the VSV M protein.


Assuntos
Interferon beta/imunologia , NF-kappa B/imunologia , Estomatite Vesicular/imunologia , Vírus da Estomatite Vesicular Indiana/imunologia , Proteínas da Matriz Viral/imunologia , Animais , Linhagem Celular , Regulação da Expressão Gênica , Interações Hospedeiro-Patógeno , Humanos , Interferon beta/genética , Camundongos , NF-kappa B/genética , Estomatite Vesicular/genética , Estomatite Vesicular/virologia , Vírus da Estomatite Vesicular Indiana/genética , Vírus da Estomatite Vesicular Indiana/fisiologia , Proteínas da Matriz Viral/genética
4.
Nat Rev Immunol ; 20(10): 585-586, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32788708
5.
Vet Microbiol ; 247: 108793, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32768236

RESUMO

Porcine epidemic diarrhea virus (PEDV) belongs to the Alphacoronavirus genus in the Coronaviridae family. Similar to other coronaviruses, PEDV encodes two papain-like proteases. Papain-like protease (PLP)2 has been proposed to play a key role in antagonizing host innate immunity. However, the function of PLP1 remains unclear. In this study, we found that overexpression of PLP1 significantly promoted PEDV replication and inhibited production of interferon-ß. Immunoprecipitation and mass spectrometry were used to identify cellular interaction partners of PLP1. Host cell poly(C) binding protein 2 (PCBP2) was determined to bind and interact with PLP1. Both endogenous and overexpressed PCBP2 co-localized with PLP1 in the cytoplasm. Overexpression of PLP1 upregulated expression of PCBP2. Furthermore, overexpression of PCBP2 promoted PEDV replication. Silencing of endogenous PCBP2 using small interfering RNAs attenuated PEDV replication. Taken together, these data demonstrated that PLP1 negatively regulated the production of type 1 interferon by interacting with PCBP2 and promoted PEDV replication.


Assuntos
Papaína/metabolismo , Vírus da Diarreia Epidêmica Suína/metabolismo , Proteínas não Estruturais Virais/metabolismo , Replicação Viral/fisiologia , Animais , Chlorocebus aethiops , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Células HEK293 , Humanos , Interferon beta/genética , Interferon beta/metabolismo , Proteína Proteolipídica de Mielina/metabolismo , Papaína/genética , Vírus da Diarreia Epidêmica Suína/genética , Vírus da Diarreia Epidêmica Suína/fisiologia , Interferência de RNA , Proteínas de Ligação a RNA , Fator de Necrose Tumoral alfa/farmacologia , Células Vero , Proteínas não Estruturais Virais/genética
6.
Arch Virol ; 165(10): 2249-2258, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32696270

RESUMO

While infectious bursal disease virus (IBDV) mainly targets immature B cells and causes T cell infiltration in the bursa of Fabricius (BF) of chickens, the effect of IBDV infection on the properties of T cells and relevant cytokine production in avian gut-associated lymphoid tissues (GALTs) remains unknown. Here, we show that while the CD8+ T cell subset is not affected, IBDV infection decreases the percentage of CD4+ T cells in the cecal tonsil (CT), but not in esophagus tonsil, pylorus tonsil, and Meckel's diverticulum of GALTs, in contrast to BF and spleen, in which the proportion of CD4+ cells increases upon IBDV infection. Further, IBDV infection upregulates IFN-γ, IL-10, and the T cell checkpoint receptor LAG-3 mRNA expression in BF. In contrast, in CTs, IBDV infection significantly increases the production of IFN-ß and CTLA-4 mRNA, while no significant effect is seen in the case of IFN-γ, IL-10 and LAG-3. Together, our data reveal differential modulation of T cell subsets and proinflammatory cytokine production in different lymphoid tissues during the course of IBDV infection.


Assuntos
Subpopulações de Linfócitos B/imunologia , Infecções por Birnaviridae/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Regulação da Expressão Gênica/imunologia , Doenças das Aves Domésticas/imunologia , Animais , Antígenos CD/genética , Antígenos CD/imunologia , Subpopulações de Linfócitos B/virologia , Infecções por Birnaviridae/genética , Infecções por Birnaviridae/patologia , Infecções por Birnaviridae/virologia , Bolsa de Fabricius/imunologia , Bolsa de Fabricius/virologia , Linfócitos T CD4-Positivos/virologia , Linfócitos T CD8-Positivos/virologia , Antígeno CTLA-4/genética , Antígeno CTLA-4/imunologia , Galinhas/virologia , Vírus da Doença Infecciosa da Bursa/crescimento & desenvolvimento , Vírus da Doença Infecciosa da Bursa/imunologia , Vírus da Doença Infecciosa da Bursa/patogenicidade , Interferon beta/genética , Interferon beta/imunologia , Interferon gama/genética , Interferon gama/imunologia , Interleucina-10/genética , Interleucina-10/imunologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/virologia , Tonsila Palatina/imunologia , Tonsila Palatina/virologia , Doenças das Aves Domésticas/genética , Doenças das Aves Domésticas/patologia , Doenças das Aves Domésticas/virologia
8.
Science ; 369(6508): 1249-1255, 2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32680882

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the current coronavirus disease 2019 (COVID-19) pandemic. A major virulence factor of SARS-CoVs is the nonstructural protein 1 (Nsp1), which suppresses host gene expression by ribosome association. Here, we show that Nsp1 from SARS-CoV-2 binds to the 40S ribosomal subunit, resulting in shutdown of messenger RNA (mRNA) translation both in vitro and in cells. Structural analysis by cryo-electron microscopy of in vitro-reconstituted Nsp1-40S and various native Nsp1-40S and -80S complexes revealed that the Nsp1 C terminus binds to and obstructs the mRNA entry tunnel. Thereby, Nsp1 effectively blocks retinoic acid-inducible gene I-dependent innate immune responses that would otherwise facilitate clearance of the infection. Thus, the structural characterization of the inhibitory mechanism of Nsp1 may aid structure-based drug design against SARS-CoV-2.


Assuntos
Betacoronavirus/química , Evasão da Resposta Imune , Imunidade Inata , Biossíntese de Proteínas , Proteínas não Estruturais Virais/química , Proteínas não Estruturais Virais/metabolismo , Betacoronavirus/imunologia , Betacoronavirus/metabolismo , Betacoronavirus/fisiologia , Sítios de Ligação , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Microscopia Crioeletrônica , Proteína DEAD-box 58/genética , Proteína DEAD-box 58/metabolismo , Humanos , Interferon beta/genética , Interferon beta/metabolismo , Modelos Moleculares , Pandemias , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Ligação Proteica , Domínios Proteicos , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , RNA Mensageiro/metabolismo , Subunidades Ribossômicas Menores de Eucariotos/química , Subunidades Ribossômicas Menores de Eucariotos/metabolismo
9.
PLoS Pathog ; 16(6): e1008611, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32511263

RESUMO

Human infection with avian influenza A (H5N1) and (H7N9) viruses causes severe respiratory diseases. PB1-F2 protein is a critical virulence factor that suppresses early type I interferon response, but the mechanism of its action in relation to high pathogenicity is not well understood. Here we show that PB1-F2 protein of H7N9 virus is a particularly potent suppressor of antiviral signaling through formation of protein aggregates on mitochondria and inhibition of TRIM31-MAVS interaction, leading to prevention of K63-polyubiquitination and aggregation of MAVS. Unaggregated MAVS accumulated on fragmented mitochondria is prone to degradation by both proteasomal and lysosomal pathways. These properties are proprietary to PB1-F2 of H7N9 virus but not shared by its counterpart in WSN virus. A recombinant virus deficient of PB1-F2 of H7N9 induces more interferon ß in infected cells. Our findings reveal a subtype-specific mechanism for destabilization of MAVS and suppression of interferon response by PB1-F2 of H7N9 virus.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Subtipo H7N9 do Vírus da Influenza A/metabolismo , Influenza Humana/metabolismo , Agregação Patológica de Proteínas/metabolismo , Transdução de Sinais , Proteínas Virais/metabolismo , Células A549 , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Cães , Células HEK293 , Humanos , Subtipo H7N9 do Vírus da Influenza A/genética , Influenza Humana/genética , Influenza Humana/patologia , Interferon beta/genética , Interferon beta/metabolismo , Células Madin Darby de Rim Canino , Mitocôndrias/genética , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Agregação Patológica de Proteínas/genética , Células THP-1 , Proteínas com Motivo Tripartido/genética , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Proteínas Virais/genética
10.
Nat Commun ; 11(1): 3187, 2020 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-32581235

RESUMO

The application of adoptive T cell therapies, including those using chimeric antigen receptor (CAR)-modified T cells, to solid tumors requires combinatorial strategies to overcome immune suppression associated with the tumor microenvironment. Here we test whether the inflammatory nature of oncolytic viruses and their ability to remodel the tumor microenvironment may help to recruit and potentiate the functionality of CAR T cells. Contrary to our hypothesis, VSVmIFNß infection is associated with attrition of murine EGFRvIII CAR T cells in a B16EGFRvIII model, despite inducing a robust proinflammatory shift in the chemokine profile. Mechanistically, type I interferon (IFN) expressed following infection promotes apoptosis, activation, and inhibitory receptor expression, and interferon-insensitive CAR T cells enable combinatorial therapy with VSVmIFNß. Our study uncovers an unexpected mechanism of therapeutic interference, and prompts further investigation into the interaction between CAR T cells and oncolytic viruses to optimize combination therapy.


Assuntos
Imunoterapia Adotiva , Interferon beta/metabolismo , Vírus Oncolíticos/metabolismo , Receptores de Antígenos Quiméricos/metabolismo , Linfócitos T/metabolismo , Animais , Apoptose , Linhagem Celular Tumoral , Quimiocinas/metabolismo , Terapia Combinada , Feminino , Interferon beta/genética , Ativação Linfocitária , Melanoma Experimental/imunologia , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Terapia Viral Oncolítica , Vírus Oncolíticos/genética , Receptor de Interferon alfa e beta/genética , Receptor de Interferon alfa e beta/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Baço/imunologia
11.
Virus Res ; 286: 198074, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32589897

RESUMO

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel human coronavirus causing the pandemic of severe pneumonia (Coronavirus Disease 2019, COVID-19). SARS-CoV-2 is highly pathogenic in human, having posed immeasurable public health challenges to the world. Innate immune response is critical for the host defense against viral infection and the dysregulation of the host innate immune responses probably aggravates SARS-CoV-2 infection, contributing to the high morbidity and lethality of COVID-19. It has been reported that some coronavirus proteins play an important role in modulating innate immunity of the host, but few studies have been conducted on SARS-CoV-2. In this study, we screened the viral proteins of SARS-CoV-2 and found that the viral ORF6, ORF8 and nucleocapsid proteins were potential inhibitors of type I interferon signaling pathway, a key component for antiviral response of host innate immune. All the three proteins showed strong inhibition on type I interferon (IFN-ß) and NF-κB-responsive promoter, further examination revealed that these proteins were able to inhibit the interferon-stimulated response element (ISRE) after infection with Sendai virus, while only ORF6 and ORF8 proteins were able to inhibit the ISRE after treatment with interferon beta. These findings would be helpful for the further study of the detailed signaling pathway and unveil the key molecular player that may be targeted.


Assuntos
Betacoronavirus/genética , Interações Hospedeiro-Patógeno/genética , Interferon beta/genética , NF-kappa B/genética , Proteínas do Nucleocapsídeo/genética , Proteínas Virais/genética , Betacoronavirus/imunologia , Regulação da Expressão Gênica , Genes Reporter , Células HEK293 , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/imunologia , Interferon beta/imunologia , Luciferases/genética , Luciferases/metabolismo , NF-kappa B/imunologia , Proteínas do Nucleocapsídeo/imunologia , Fosfoproteínas , Plasmídeos/química , Plasmídeos/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Elementos de Resposta , Vírus Sendai/genética , Vírus Sendai/imunologia , Transdução de Sinais , Transfecção/métodos , Proteínas Virais/imunologia
12.
Arch Virol ; 165(7): 1557-1567, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32356187

RESUMO

Caprine arthritis-encephalitis (CAE) is a chronic progressive infectious disease caused by caprine arthritis-encephalitis virus (CAEV) that seriously threatens the goat industry. Chronic infection and life-long multi-tissue inflammation are the typical features of the disease. Innate antiviral immunity is essential for the host defense system that rapidly recognizes and eliminates invading viruses. Interferon ß (IFN-ß) is important for innate immunity and regulates immunity against a broad spectrum of viruses. To investigate the details of the IFN-ß response to CAEV infection, the effects of six viral proteins and the molecular mechanisms by which they affect IFN-ß production were analyzed. Overexpression of DU and Vif promote virus proliferation and inhibit the production of IFN-ß. qRT-PCR and luciferase reporter assays showed that overexpression of Vif inhibits the expression of luciferase under the control of the ISRE, NF-κB or IFN-ß promoter but does not affect the expression of IFN-ß activated by IRF3, indicating that Vif negatively regulates IFN-ß production by affecting upstream signal transduction of IRF3. Amino acids 149-164 of Vif were found to be necessary for the inhibitory effect of IFN-ß production. Our results indicate that CAEV evades surveillance and clearance by intracellular innate immunity by downregulating IFN-ß production.


Assuntos
Vírus da Artrite-Encefalite Caprina/imunologia , Produtos do Gene vif/imunologia , Doenças das Cabras/imunologia , Interferon beta/imunologia , Infecções por Lentivirus/veterinária , Animais , Vírus da Artrite-Encefalite Caprina/genética , Produtos do Gene vif/genética , Doenças das Cabras/genética , Doenças das Cabras/virologia , Cabras , Interações Hospedeiro-Patógeno , Imunidade Inata , Interferon beta/genética , Infecções por Lentivirus/genética , Infecções por Lentivirus/imunologia , Infecções por Lentivirus/virologia , NF-kappa B/genética , NF-kappa B/imunologia
13.
Proc Natl Acad Sci U S A ; 117(23): 12674-12685, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32430322

RESUMO

Robust cytotoxic T cell infiltration has proven to be difficult to achieve in solid tumors. We set out to develop a flexible protocol to efficiently transfect tumor and stromal cells to produce immune-activating cytokines, and thus enhance T cell infiltration while debulking tumor mass. By combining ultrasound with tumor-targeted microbubbles, membrane pores are created and facilitate a controllable and local transfection. Here, we applied a substantially lower transmission frequency (250 kHz) than applied previously. The resulting microbubble oscillation was significantly enhanced, reaching an effective expansion ratio of 35 for a peak negative pressure of 500 kPa in vitro. Combining low-frequency ultrasound with tumor-targeted microbubbles and a DNA plasmid construct, 20% of tumor cells remained viable, and ∼20% of these remaining cells were transfected with a reporter gene both in vitro and in vivo. The majority of cells transfected in vivo were mucin 1+/CD45- tumor cells. Tumor and stromal cells were then transfected with plasmid DNA encoding IFN-ß, producing 150 pg/106 cells in vitro, a 150-fold increase compared to no-ultrasound or no-plasmid controls and a 50-fold increase compared to treatment with targeted microbubbles and ultrasound (without IFN-ß). This enhancement in secretion exceeds previously reported fourfold to fivefold increases with other in vitro treatments. Combined with intraperitoneal administration of checkpoint inhibition, a single application of IFN-ß plasmid transfection reduced tumor growth in vivo and recruited efficacious immune cells at both the local and distant tumor sites.


Assuntos
Imunoterapia/métodos , Interferon beta/genética , Neoplasias Experimentais/terapia , Linfócitos T/imunologia , Transfecção/métodos , Ondas Ultrassônicas , Animais , Linhagem Celular Tumoral , Membrana Celular/efeitos da radiação , Movimento Celular , Humanos , Interferon beta/metabolismo , Camundongos , Microbolhas/uso terapêutico , Linfócitos T/fisiologia
14.
J Virol ; 94(15)2020 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-32434887

RESUMO

Hepatitis C virus (HCV) has evolved mechanisms to evade innate immunity that are leading to chronic infections. The immunological function of the HCV frameshift (F) protein, which is a frameshift product of core coding sequences, has not been well characterized. The HCV F protein is produced during natural HCV infections and is found most commonly in genotype 1 HCV. In this study, we investigated whether the F protein plays a role in type I interferon (IFN) induction pathways. We engineered F expression constructs from core coding sequences of 4 genotypes (1a, 2a, 3a, and 4a) of HCV as well as the sequences which would only be able to produce core proteins. The peptide lengths and amino acids sequences of F proteins are highly variable. We hypothesized that F proteins from different genotypes might control the type I IFN production and response differently. We found that both IFN-beta (IFN-ß) promoter activities are significantly higher in genotype 1a F protein (F1a)-expressing cells. Conversely, the IFN-ß promoter activities are lower in genotype 2a F (F2a) protein-expressing cells. We also used real-time PCR to confirm IFN-ß mRNA expression levels. By generating chimera F proteins, we discovered that the effects of F proteins were determined by the amino acid sequence 40 to 57 of genotype 1a. The regulation of type I IFN induction pathway is related but not limited to the activity of F1a to interact with proteasome subunits and to disturb the proteasome activity. Further molecular mechanisms of how F proteins from different genotypes of HCV control these pathways differently remain to be investigated.IMPORTANCE Although naturally present in HCV infection patient serum, the virological or immunological functions of the HCV F protein, which is a frameshift product of core coding sequences, remain unclear. Here, we report the effects of the HCV F protein between genotypes and discuss a potential explanation for the differential responses to type I IFN-based therapy among patients infected with different genotypes of HCV. Our study provides one step forward to understanding the host response during HCV infection and new insights for the prediction of the outcome of IFN-based therapy in HCV patients.


Assuntos
Genótipo , Hepacivirus/metabolismo , Hepatite C/metabolismo , Interferon beta/biossíntese , Transdução de Sinais , Proteínas do Core Viral/metabolismo , Linhagem Celular Tumoral , Hepacivirus/genética , Hepatite C/genética , Humanos , Interferon beta/genética , Proteínas do Core Viral/genética
15.
Virology ; 546: 79-87, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32452419

RESUMO

In a previous study, we have shown that highly-pathogenic PRRSV (HP-PRRSV) nonstructural protein 4 (nsp4) antagonizes type I IFN expression induced by poly(I:C). Here, we demonstrated that the mutation of Aspartic acid 185 (Asp185) impaired the ability of nsp4 to inhibit IFN-I production induced by poly(I:C). Subsequently, we verified that all the mutants at the residue 185, regardless of amino acid size (including Cys and Ser) and charge (including Glu and Lys), impaired nsp4 catalytic activity. However, when Asp185 in nsp4 was replaced by a similar structure amino acid Asparagine 185 (Asn185), nsp4 stayed but with a decreased protease activity. Importantly, the recombinant virus with Asn185 mutation in HP-PRRSV-nsp4 exhibited slower replication rate and higher ability to induce IFN-I expression compared with wild-type (wt) HP-PRRSV.


Assuntos
Ácido Aspártico/metabolismo , Interferon beta/metabolismo , Síndrome Respiratória e Reprodutiva Suína/metabolismo , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/metabolismo , Proteínas não Estruturais Virais/química , Proteínas não Estruturais Virais/metabolismo , Animais , Interações Hospedeiro-Patógeno , Interferon beta/genética , Síndrome Respiratória e Reprodutiva Suína/genética , Vírus da Síndrome Respiratória e Reprodutiva Suína/química , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Vírus da Síndrome Respiratória e Reprodutiva Suína/patogenicidade , Suínos , Proteínas não Estruturais Virais/genética , Virulência
16.
Oncogene ; 39(22): 4344-4357, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32335582

RESUMO

We explore a novel strategy of activating immune signaling through increased micronuclei formation utilizing a cell cycle checkpoint inhibitor to drive cell cycle progression following ionizing radiation. The Chk1/2 inhibitor AZD7762 is used to abrogate radiation therapy (RT)-induced G2/M cell cycle arrest in multiple cell lines and, we find that this therapeutic combination promotes increased micronuclei formation in vitro and subsequently drives increased type I interferon signaling and cytotoxic T-cell activation. In vivo studies using B16-F10 melanoma cancer cells implanted in C57/BL6 mice demonstrate improved rates of tumor control at the abscopal (unirradiated) site, located outside of the radiation field, only in the AZD7762 + RT group, with a corresponding reduction in mean tumor volume, increase in the CD8 T-cell population, and immune activated gene signaling. Our results demonstrate that targeted inhibition of cell cycle checkpoint activation following ionizing radiation drives increased production of immunogenic micronuclei, leading to systemic tumor response with potential future clinical benefit.


Assuntos
Núcleo Celular/efeitos dos fármacos , Núcleo Celular/efeitos da radiação , Quinase 1 do Ponto de Checagem/antagonistas & inibidores , Quinase do Ponto de Checagem 2/antagonistas & inibidores , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos da radiação , Melanoma Experimental/imunologia , Proteínas de Neoplasias/antagonistas & inibidores , Tiofenos/farmacologia , Ureia/análogos & derivados , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Interferon beta/biossíntese , Interferon beta/genética , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/patologia , Melanoma Experimental/radioterapia , Proteínas de Membrana/biossíntese , Proteínas de Membrana/genética , Camundongos Endogâmicos C57BL , Testes para Micronúcleos , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Fator de Transcrição STAT1/biossíntese , Fator de Transcrição STAT1/genética , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/efeitos da radiação , Ureia/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Oncol Rep ; 43(6): 2045-2052, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32236604

RESUMO

Although the effects of stem cells expressing anticancer genes on tumor growth have been demonstrated by many researchers in various types of cancer, relatively few studies have investigated their inhibitory effects on cancer metastasis. In the present study, we examined the inhibitory effects of cytosine deaminase (CD)/5­fluorocytosine (5­FC) and interferon­ß (IFN­ß) using genetically engineered neural stem cells (hNSCs) in a cellular and metastasis model of renal cell carcinoma (RCC). The CD/5­FC method has the advantage of minimizing damage to normal tissues since it selectively targets cancer cells by the CD gene, which converts prodrug 5­FC to the drug 5­fluorouracil. Moreover, we used hNSCs as a tool to effectively deliver the anticancer genes to the tumor site. These stem cells are known to possess tumor­tropism because of chemoattractant factors expressed in cancer cells. Therefore, we ascertained the expression of these factors in A498 cells, a cell line of RCC, and identified the A498­specific migration ability of hNSCs. We also confirmed that the proliferation of A498 cells was significantly reduced by therapeutic hNSCs in the presence of 5­FC. Furthermore, we established an A498 metastasis model. In the animal experiment, the weight of the lungs increased in response to cancer metastasis, but was normalized by hNSCs expressing CD and/or IFN­ß genes, while the incidence of liver metastasis was suppressed by the hNSCs. Overall, the results of this study demonstrate that stem cells expressing anticancer genes have the potential for use as an alternative to conventional therapy for metastatic cancer.


Assuntos
Carcinoma de Células Renais/terapia , Citosina Desaminase/genética , Interferon beta/genética , Neoplasias Hepáticas/terapia , Células-Tronco Neurais/citologia , Transplante de Células-Tronco/métodos , Animais , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/metabolismo , Linhagem Celular Tumoral , Células Cultivadas , Técnicas de Cocultura , Citosina Desaminase/metabolismo , Feminino , Engenharia Genética , Humanos , Interferon beta/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Camundongos , Metástase Neoplásica , Células-Tronco Neurais/metabolismo , Células-Tronco Neurais/transplante , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto
18.
J Virol ; 94(11)2020 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-32238581

RESUMO

Highly pathogenic avian influenza A(H5N8) viruses first emerged in China in 2010 and in 2014 spread throughout Asia and to Europe and the United States via migrating birds. Influenza A(H5N8) viruses were first detected in the Netherlands in 2014 and caused five outbreaks in poultry farms but were infrequently detected in wild birds. In 2016, influenza A(H5N8) viruses were reintroduced into the Netherlands, resulting in eight poultry farm outbreaks. This outbreak resulted in numerous dead wild birds with severe pathology. Phylogenetic analysis showed that the polymerase genes of these viruses had undergone extensive reassortment between outbreaks. Here, we investigated the differences in virulence between the 2014-15 and the 2016-17 outbreaks by characterizing the polymerase complex of influenza A(H5N8) viruses from both outbreaks. We found that viruses from the 2014-15 outbreak had significantly higher polymerase complex activity in both human and avian cell lines than did those from the 2016-17 outbreak. No apparent differences in the balance between transcription and replication of the viral genome were observed. Interestingly, the 2014-15 polymerase complexes induced significantly higher levels of interferon beta (IFN-ß) than the polymerase complexes of the 2016-17 outbreak viruses, mediated via retinoic acid-inducible gene I (RIG-I). Inoculation of primary duck cells with recombinant influenza A(H5N8) viruses, including viruses with reassorted polymerase complexes, showed that the polymerase complexes from the 2014-15 outbreak induced higher levels of IFN-ß despite relatively minor differences in replication capacity. Together, these data suggest that despite the lower levels of polymerase activity, the higher 2016-17 influenza A(H5N8) virus virulence may be attributed to the lower level of activation of the innate immune system.IMPORTANCE Compared to the 2014-15 outbreak, the 2016-17 outbreak of influenza A(H5N8) viruses in the Netherlands and Europe was more virulent; the number of dead or diseased wild birds found and the severity of pathological changes were higher during the 2016-17 outbreak. The polymerase complex plays an important role in influenza virus virulence, and the gene segments of influenza A(H5N8) viruses reassorted extensively between the outbreaks. In this study, the 2014-15 polymerase complexes were found to be more active, which is counterintuitive with the observed higher virulence of the 2016-17 outbreak viruses. Interestingly, the 2014-15 polymerase complexes also induced higher levels of IFN-ß. These findings suggest that the higher virulence of influenza A(H5N8) viruses from the 2016-17 outbreak may be related to the lower induction of IFN-ß. An attenuated interferon response could lead to increased dissemination, pathology, and mortality, as observed in (wild) birds infected during the 2016-2017 outbreak.


Assuntos
Proteínas Aviárias , Surtos de Doenças , Vírus da Influenza A Subtipo H5N8 , Influenza Aviária , Interferon beta , Proteínas Virais , Animais , Proteínas Aviárias/genética , Proteínas Aviárias/imunologia , Coturnix , Cães , Patos , Células HEK293 , Humanos , Vírus da Influenza A Subtipo H5N8/genética , Vírus da Influenza A Subtipo H5N8/imunologia , Influenza Aviária/epidemiologia , Influenza Aviária/genética , Influenza Aviária/imunologia , Interferon beta/genética , Interferon beta/imunologia , Células Madin Darby de Rim Canino , /imunologia , Proteínas Virais/genética , Proteínas Virais/imunologia
19.
Immunity ; 52(4): 668-682.e7, 2020 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-32294407

RESUMO

The primary mechanisms supporting immunoregulatory polarization of myeloid cells upon infiltration into tumors remain largely unexplored. Elucidation of these signals could enable better strategies to restore protective anti-tumor immunity. Here, we investigated the role of the intrinsic activation of the PKR-like endoplasmic reticulum (ER) kinase (PERK) in the immunoinhibitory actions of tumor-associated myeloid-derived suppressor cells (tumor-MDSCs). PERK signaling increased in tumor-MDSCs, and its deletion transformed MDSCs into myeloid cells that activated CD8+ T cell-mediated immunity against cancer. Tumor-MDSCs lacking PERK exhibited disrupted NRF2-driven antioxidant capacity and impaired mitochondrial respiratory homeostasis. Moreover, reduced NRF2 signaling in PERK-deficient MDSCs elicited cytosolic mitochondrial DNA elevation and, consequently, STING-dependent expression of anti-tumor type I interferon. Reactivation of NRF2 signaling, conditional deletion of STING, or blockade of type I interferon receptor I restored the immunoinhibitory potential of PERK-ablated MDSCs. Our findings demonstrate the pivotal role of PERK in tumor-MDSC functionality and unveil strategies to reprogram immunosuppressive myelopoiesis in tumors to boost cancer immunotherapy.


Assuntos
Carcinoma Pulmonar de Lewis/imunologia , Carcinoma Epitelial do Ovário/imunologia , Regulação Neoplásica da Expressão Gênica , Melanoma Experimental/imunologia , Proteínas de Membrana/imunologia , Neoplasias Cutâneas/imunologia , eIF-2 Quinase/imunologia , Animais , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Carcinoma Pulmonar de Lewis/genética , Carcinoma Pulmonar de Lewis/metabolismo , Carcinoma Pulmonar de Lewis/patologia , Carcinoma Epitelial do Ovário/genética , Carcinoma Epitelial do Ovário/metabolismo , Carcinoma Epitelial do Ovário/patologia , Feminino , Humanos , Imunossupressão , Interferon-alfa/genética , Interferon-alfa/imunologia , Interferon beta/genética , Interferon beta/imunologia , Masculino , Melanoma Experimental/genética , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/imunologia , Mitocôndrias/metabolismo , Células Supressoras Mieloides/imunologia , Células Supressoras Mieloides/patologia , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/imunologia , Receptores de Interferon/genética , Receptores de Interferon/imunologia , Transdução de Sinais , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Resposta a Proteínas não Dobradas/imunologia , eIF-2 Quinase/deficiência , eIF-2 Quinase/genética
20.
Emerg Microbes Infect ; 9(1): 714-726, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32196427

RESUMO

Tick-borne encephalitis virus (TBEV) accounts for approximately 10,000 annual cases of severe encephalitis in Europe and Asia and causes encephalitis in humans. In this study, we demonstrate TBEV appears to activate the interferon (IFN)-ß dependent on RIG-I/MDA5. Both the IFN-ß accumulation and the IFN stimulated genes (ISGs) transcription greatly delay. Further studies reveal that TBEV NS4A could block the phosphorylation and dimerization of STAT1/STAT2 to affect type I and II IFN-mediated STAT signalling. Additional data indicate that the residue at K132 of TBEV NS4A could be modified by ubiquitination and this modification is necessary for the interaction of NS4A with STAT1. Dynamic ubiquitination of the NS4 protein during TBEV infection might account for delayed activation of the ISGs. These results define the TBEV NS4A as an antagonist of the IFN response, by demonstrating a correlation between the association and STAT interference. Our findings provide a foundation for further understanding how TBEV evade innate immunity and a potential viral target for intervention.


Assuntos
Vírus da Encefalite Transmitidos por Carrapatos/fisiologia , Interferon Tipo I/antagonistas & inibidores , Proteínas não Estruturais Virais/metabolismo , Linhagem Celular , Proteína DEAD-box 58/metabolismo , Vírus da Encefalite Transmitidos por Carrapatos/metabolismo , Humanos , Fatores Reguladores de Interferon/antagonistas & inibidores , Fatores Reguladores de Interferon/genética , Fatores Reguladores de Interferon/metabolismo , Interferon Tipo I/metabolismo , Helicase IFIH1 Induzida por Interferon/metabolismo , Fator Gênico 3 Estimulado por Interferon, Subunidade gama/metabolismo , Interferon beta/genética , Interferon beta/metabolismo , Lisina/metabolismo , Fosforilação , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT2/metabolismo , Transdução de Sinais , Ubiquitinação , Regulação para Cima , Domínios de Homologia de src
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...