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1.
Vet Microbiol ; 236: 108395, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31500730

RESUMO

Porcine reproductive and respiratory syndrome virus (PRRSV) is an economically significant pathogen that has been recognized for its genetic variation, rapid evolution, and immune suppression. Type I interferons (IFNs) play an important role in host defense against viral infection by inducing many antiviral effectors, which might be a selective pressure driving viral evolution towards IFN resistance. To investigate the IFN resistance-related variation of PRRSV genome under IFN selective pressure and explore the molecular mechanism of IFN sensitivity changes, PRRSV strain JXwn06 was serially propagated in porcine pulmonary alveolar macrophages (PAMs) with IFNα treatment for 45 passages and 3 rounds of purification. Four mutant strains named JX-αP51n (n = 1, 2, 3 and 4) with reduced IFNα sensitivity were selected; the strains showed a 100-fold higher titer than the passaging-control strain JX-P51 in IFNα-treated PAMs. IFNα-resistant strains were found to antagonize the IFNα-activated JAK-STAT signaling pathway to a greater extent than the nonresistant strain by down-regulating the expression level of IFNα-activated pJAK1 through interfering with phosphatase. Furthermore, the PRRSV genetic variations interacting with IFNα were identified by full genomic sequencing and alignment. Among these mutations, amino acid substitutions in nsp1ß (E87 G), GP3 (F143 L) and GP5 (Y136 H) were found to correlate with increased IFNα resistance by enhancing the suppression effect on pJAK1, which could be further increased if these three substitution sites were combined. These findings provide some novel evidence for understanding PRRSV genetic variation under host selective pressure and viral evolution strategies to evade the host innate immune response.


Assuntos
Farmacorresistência Viral/genética , Interferon-alfa/farmacologia , Mutação , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Proteínas Virais/metabolismo , Sequência de Aminoácidos , Animais , Linhagem Celular , Regulação da Expressão Gênica , Haplorrinos , Fosforilação , Vírus da Síndrome Respiratória e Reprodutiva Suína/efeitos dos fármacos , RNA Viral , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT2/genética , Fator de Transcrição STAT2/metabolismo , Transdução de Sinais , Suínos , Proteínas Virais/genética
2.
PLoS Pathog ; 15(8): e1007949, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31374104

RESUMO

Host encounters with viruses lead to an innate immune response that must be rapid and broadly targeted but also tightly regulated to avoid the detrimental effects of unregulated interferon expression. Viral stimulation of host negative regulatory mechanisms is an alternate method of suppressing the host innate immune response. We examined three key mediators of the innate immune response: NF-KB, STAT1 and STAT2 during HCV infection in order to investigate the paradoxical induction of an innate immune response by HCV despite a multitude of mechanisms combating the host response. During infection, we find that all three are repressed only in HCV infected cells but not in uninfected bystander cells, both in vivo in chimeric mouse livers and in cultured Huh7.5 cells after IFNα treatment. We show here that HCV and Flaviviruses suppress the innate immune response by upregulation of PDLIM2, independent of the host interferon response. We show PDLIM2 is an E3 ubiquitin ligase that also acts to stimulate nuclear degradation of STAT2. Interferon dependent relocalization of STAT1/2 to the nucleus leads to PDLIM2 ubiquitination of STAT2 but not STAT1 and the proteasome-dependent degradation of STAT2, predominantly within the nucleus. CRISPR/Cas9 knockout of PDLIM2 results in increased levels of STAT2 following IFNα treatment, retention of STAT2 within the nucleus of HCV infected cells after IFNα stimulation, increased interferon response, and increased resistance to infection by several flaviviruses, indicating that PDLIM2 is a global regulator of the interferon response.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Infecções por Flavivirus/imunologia , Flavivirus/imunologia , Hepacivirus/imunologia , Hepatite C/imunologia , Imunidade Inata/imunologia , Proteínas com Domínio LIM/fisiologia , Fator de Transcrição STAT2/metabolismo , Animais , Antivirais/farmacologia , Flavivirus/efeitos dos fármacos , Infecções por Flavivirus/tratamento farmacológico , Infecções por Flavivirus/virologia , Hepacivirus/efeitos dos fármacos , Hepatite C/tratamento farmacológico , Hepatite C/virologia , Humanos , Imunidade Inata/efeitos dos fármacos , Interferon-alfa/farmacologia , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/metabolismo , Camundongos , Camundongos Knockout , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , NF-kappa B , Fator de Transcrição STAT2/genética , Transdução de Sinais
3.
Nat Commun ; 10(1): 2921, 2019 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-31266943

RESUMO

Cells maintain the balance between homeostasis and inflammation by adapting and integrating the activity of intracellular signaling cascades, including the JAK-STAT pathway. Our understanding of how a tailored switch from homeostasis to a strong receptor-dependent response is coordinated remains limited. Here, we use an integrated transcriptomic and proteomic approach to analyze transcription-factor binding, gene expression and in vivo proximity-dependent labelling of proteins in living cells under homeostatic and interferon (IFN)-induced conditions. We show that interferons (IFN) switch murine macrophages from resting-state to induced gene expression by alternating subunits of transcription factor ISGF3. Whereas preformed STAT2-IRF9 complexes control basal expression of IFN-induced genes (ISG), both type I IFN and IFN-γ cause promoter binding of a complete ISGF3 complex containing STAT1, STAT2 and IRF9. In contrast to the dogmatic view of ISGF3 formation in the cytoplasm, our results suggest a model wherein the assembly of the ISGF3 complex occurs on DNA.


Assuntos
Regulação da Expressão Gênica , Fator Gênico 3 Estimulado por Interferon, Subunidade gama/metabolismo , Fator Gênico 3 Estimulado por Interferon/metabolismo , Interferons/metabolismo , Fator de Transcrição STAT2/metabolismo , Animais , Feminino , Humanos , Fator Gênico 3 Estimulado por Interferon/genética , Fator Gênico 3 Estimulado por Interferon, Subunidade gama/genética , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Regiões Promotoras Genéticas , Células RAW 264.7 , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT2/genética , Transcrição Genética
4.
Viruses ; 11(2)2019 01 22.
Artigo em Inglês | MEDLINE | ID: mdl-30678320

RESUMO

Zika virus (ZIKV) infection can cause severe congenital diseases, such as microcephaly, ocular defects and arthrogryposis in fetuses, and Guillain⁻Barré syndrome in adults. Efficacious therapeutic treatments for infected patients, as well as prophylactic treatments to prevent new infections are needed for combating ZIKV infection. Here, we report that ZIKV-specific human polyclonal antibodies (SAB-155), elicited in transchromosomal bovine (TcB), provide significant protection from infection by ZIKV in STAT2 knockout (KO) golden Syrian hamsters both prophylactically and therapeutically. These antibodies also prevent testicular lesions in this hamster model. Our data indicate that antibody-mediated immunotherapy is effective in treating ZIKV infection. Because suitable quantities of highly potent human polyclonal antibodies can be quickly produced from the TcB system against ZIKV and have demonstrated therapeutic efficacy in a small animal model, they have the potential as an effective countermeasure against ZIKV infection.


Assuntos
Anticorpos Neutralizantes/uso terapêutico , Anticorpos Antivirais/uso terapêutico , Imunização Passiva , Fator de Transcrição STAT2/genética , Infecção por Zika virus/prevenção & controle , Infecção por Zika virus/terapia , Animais , Animais Geneticamente Modificados , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Bovinos , Cricetinae , Modelos Animais de Doenças , Feminino , Técnicas de Inativação de Genes , Humanos , Masculino , Testículo/patologia , Testículo/virologia , Zika virus
5.
Hum Antibodies ; 27(2): 91-98, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30412483

RESUMO

BACKGROUND: Multiple sclerosis (MS) is the most common chronic, inflammatory, autoimmune disease of the central nervous system (CNS) maintained by the secretion of a large number of cytokines [1]. The signal transducer and activator of transcription (STAT) family has an essential role in transmitting many of the cytokine-mediated signals and failure in the signaling process contributes to the etiopathogenesis of MS. METHODS: This study aimed to assess STAT1, STAT2 and STAT3 gene expression in the blood of 50 relapsing-remitting MS (RR-MS) patients and 50 healthy controls by TaqMan Quantitative Real-Time PCR. RESULTS: The results showed that STAT1 gene expression was significantly up-regulated (p= 0.023), whereas STAT2 gene expression was significantly down-regulated (p< 0.0001) in MS patients compared to controls. On the other hand, there was no significant difference between MS patients and controls for STAT3 gene expression (p= 0.837). In addition, there was no significant correlation between the expression of STAT1, STAT2, STAT3 genes and clinical findings, such as the level of physical disability in MS patients (according to the Kurtzke Expanded Disability Status Scale (EDSS) criterion) and disease duration. CONCLUSION: A significant positive correlation was demonstrated between STAT1 and STAT2 and also between STAT1 and STAT3. This study shows for the first time that a comparison of the relative quantitative expression of three different STAT genes in the blood cells of MS patients compared to controls revealed marked differences in the expression of the STAT family genes that might reflect their different roles in the pathogenesis of MS. These transcripts might be useful biomarkers for evaluating the efficacy of IFN treatment of the MS patients.


Assuntos
Esclerose Múltipla/sangue , Esclerose Múltipla/genética , RNA Mensageiro/sangue , RNA Mensageiro/genética , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT2/genética , Fator de Transcrição STAT3/genética , Adolescente , Adulto , Estudos de Casos e Controles , Regulação para Baixo/genética , Feminino , Expressão Gênica/genética , Humanos , Masculino , Pessoa de Meia-Idade , Transdução de Sinais/genética , Regulação para Cima/genética , Adulto Jovem
6.
Front Immunol ; 9: 2879, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30574148

RESUMO

STAT1 has a key role in the regulation of innate and adaptive immunity by inducing transcriptional changes in response to cytokines, such as all types of interferons (IFN). STAT1 exist as two splice isoforms, which differ in regard to the C-terminal transactivation domain (TAD). STAT1ß lacks the C-terminal TAD and has been previously reported to be a weaker transcriptional activator than STAT1α, although this was strongly dependent on the target gene. The mechanism of this context-dependent effects remained unclear. By using macrophages from mice that only express STAT1ß, we investigated the role of the C-terminal TAD during the distinct steps of transcriptional activation of selected target genes in response to IFNγ. We show that the STAT1 C-terminal TAD is absolutely required for the recruitment of RNA polymerase II (Pol II) and for the establishment of active histone marks at the class II major histocompatibility complex transactivator (CIIta) promoter IV, whereas it is dispensable for histone acetylation at the guanylate binding protein 2 (Gbp2) promoter but required for an efficient recruitment of Pol II, which correlated with a strongly reduced, but not absent, transcriptional activity. IFNγ-induced expression of Irf7, which is mediated by STAT1 in complex with STAT2 and IRF9, did not rely on the presence of the C-terminal TAD of STAT1. Moreover, we show for the first time that the STAT1 C-terminal TAD is required for an efficient recruitment of components of the core Mediator complex to the IFN regulatory factor (Irf) 1 and Irf8 promoters, which both harbor an open chromatin state under basal conditions. Our study identified novel functions of the STAT1 C-terminal TAD in transcriptional activation and provides mechanistic explanations for the gene-specific transcriptional activity of STAT1ß.


Assuntos
Proteínas Nucleares/genética , Domínios Proteicos/imunologia , RNA Polimerase II/metabolismo , Fator de Transcrição STAT1/metabolismo , Transativadores/genética , Ativação Transcricional/imunologia , Animais , Células Cultivadas , Proteínas de Ligação ao GTP/genética , Proteínas de Ligação ao GTP/metabolismo , Código das Histonas , Fator Gênico 3 Estimulado por Interferon, Subunidade gama/genética , Fator Gênico 3 Estimulado por Interferon, Subunidade gama/metabolismo , Macrófagos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Nucleares/metabolismo , Cultura Primária de Células , Regiões Promotoras Genéticas/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/imunologia , Isoformas de Proteínas/metabolismo , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/imunologia , Fator de Transcrição STAT2/genética , Fator de Transcrição STAT2/metabolismo , Transativadores/metabolismo
7.
Viruses ; 10(12)2018 12 14.
Artigo em Inglês | MEDLINE | ID: mdl-30558110

RESUMO

Type I interferon (IFN-I) is the first line of mammalian host defense against viral infection. To counteract this, the flaviviruses, like other viruses, have encoded a variety of antagonists, and use a multi-layered molecular defense strategy to establish their infections. Among the most potent antagonists is non-structural protein 5 (NS5), which has been shown for all disease-causing flaviviruses to target different steps and players of the type I IFN signaling pathway. Here, we summarize the type I IFN antagonist mechanisms used by flaviviruses with a focus on the role of NS5 in regulating one key regulator of type I IFN, signal transducer and activator of transcription 2 (STAT2).


Assuntos
Flavivirus/imunologia , Interferon Tipo I/imunologia , Transdução de Sinais , Proteínas não Estruturais Virais/imunologia , Animais , Flavivirus/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata , Interferon Tipo I/antagonistas & inibidores , Camundongos , Fator de Transcrição STAT2/genética , Células Vero
8.
Nat Commun ; 9(1): 4560, 2018 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-30385750

RESUMO

Zika virus is a mosquito-borne flavivirus closely related to dengue virus that can cause severe disease in humans, including microcephaly in newborns and Guillain-Barré syndrome in adults. Specific treatments and vaccines for Zika virus are not currently available. Here, we isolate and characterize four monoclonal antibodies (mAbs) from an infected patient that target the non-structural protein NS1. We show that while these antibodies are non-neutralizing, NS1-specific mAbs can engage FcγR without inducing antibody dependent enhancement (ADE) of infection in vitro. Moreover, we demonstrate that mAb AA12 has protective efficacy against lethal challenges of African and Asian lineage strains of Zika virus in Stat2-/- mice. Protection is Fc-dependent, as a mutated antibody unable to activate known Fc effector functions or complement is not protective in vivo. This study highlights the importance of the ZIKV NS1 protein as a potential vaccine antigen.


Assuntos
Anticorpos Antivirais/metabolismo , Receptores de IgG/metabolismo , Proteínas não Estruturais Virais/imunologia , Vacinas Virais/imunologia , Infecção por Zika virus/prevenção & controle , Zika virus/imunologia , Animais , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais/farmacologia , Anticorpos Antivirais/farmacologia , Modelos Animais de Doenças , Células HEK293 , Humanos , Células Jurkat , Camundongos , Camundongos Knockout , Testes de Neutralização , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia , Fator de Transcrição STAT2/genética , Células Vero , Proteínas não Estruturais Virais/metabolismo , Zika virus/metabolismo
9.
Front Immunol ; 9: 2151, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30337919

RESUMO

Influenza is a common respiratory virus that infects between 5 and 20% of the US population and results in 30,000 deaths annually. A primary cause of influenza-associated death is secondary bacterial pneumonia. We have previously shown that influenza induces type I interferon (IFN)-mediated inhibition of Type 17 immune responses, resulting in exacerbation of bacterial burden during influenza and Staphylococcus aureus super-infection. In this study, we investigated the role of STAT2 signaling during influenza and influenza-bacterial super-infection in mice. Influenza-infected STAT2 -/- mice had increased morbidity, viral burden, and inflammation when compared to wild-type mice. Despite an exaggerated inflammatory response to influenza infection, we found increased bacterial control and survival in STAT2 deficient mice during influenza-MRSA super-infection compared to controls. Further, we found that increased bacterial clearance during influenza-MRSA super-infection is not due to rescue of Type 17 immunity. Absence of STAT2 was associated with increased accumulation of M1, M2 and M1/M2 co-expressing macrophages during influenza-bacterial super-infection. Neutralization of IFNγ (M1) and/or Arginase 1 (M2) impaired bacterial clearance in Stat2 -/- mice during super-infection, demonstrating that pulmonary macrophages expressing a mixed M1/M2 phenotype promote bacterial control during influenza-bacterial super-infection. Together, these results suggest that the STAT2 signaling is involved in suppressing macrophage activation and bacterial control during influenza-bacterial super-infection. Further, these studies reveal novel mechanistic insight into the roles of macrophage subpopulations in pulmonary host defense.


Assuntos
Influenza Humana/imunologia , Macrófagos Alveolares/imunologia , Pneumonia Estafilocócica/imunologia , Fator de Transcrição STAT2/metabolismo , Superinfecção/imunologia , Animais , Transplante de Medula Óssea , Embrião de Galinha , Modelos Animais de Doenças , Feminino , Humanos , Vírus da Influenza A Subtipo H1N1/imunologia , Influenza Humana/diagnóstico , Influenza Humana/microbiologia , Influenza Humana/mortalidade , Ativação de Macrófagos/imunologia , Macrófagos Alveolares/metabolismo , Masculino , Células-Tronco Mesenquimais , Staphylococcus aureus Resistente à Meticilina/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pneumonia Estafilocócica/diagnóstico , Pneumonia Estafilocócica/microbiologia , Pneumonia Estafilocócica/mortalidade , Cultura Primária de Células , Fator de Transcrição STAT2/genética , Fator de Transcrição STAT2/imunologia , Índice de Gravidade de Doença , Transdução de Sinais/imunologia , Superinfecção/diagnóstico , Superinfecção/microbiologia , Superinfecção/mortalidade , Quimeras de Transplante
10.
Proc Natl Acad Sci U S A ; 115(39): E9172-E9181, 2018 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-30206152

RESUMO

Genome-wide investigations of host-pathogen interactions are often limited by analyses of mixed populations of infected and uninfected cells, which lower sensitivity and accuracy. To overcome these obstacles and identify key mechanisms by which Zika virus (ZIKV) manipulates host responses, we developed a system that enables simultaneous characterization of genome-wide transcriptional and epigenetic changes in ZIKV-infected and neighboring uninfected primary human macrophages. We demonstrate that transcriptional responses in ZIKV-infected macrophages differed radically from those in uninfected neighbors and that studying the cell population as a whole produces misleading results. Notably, the uninfected population of macrophages exhibits the most rapid and extensive changes in gene expression, related to type I IFN signaling. In contrast, infected macrophages exhibit a delayed and attenuated transcriptional response distinguished by preferential expression of IFNB1 at late time points. Biochemical and genomic studies of infected macrophages indicate that ZIKV infection causes both a targeted defect in the type I IFN response due to degradation of STAT2 and reduces RNA polymerase II protein levels and DNA occupancy, particularly at genes required for macrophage identity. Simultaneous evaluation of transcriptomic and epigenetic features of infected and uninfected macrophages thereby reveals the coincident evolution of dominant proviral or antiviral mechanisms, respectively, that determine the outcome of ZIKV exposure.


Assuntos
Imunidade Inata , Macrófagos/imunologia , Infecção por Zika virus/imunologia , Zika virus/imunologia , Efeito Espectador , Feminino , Humanos , Interferon beta/genética , Interferon beta/imunologia , Macrófagos/patologia , Masculino , Proteólise , RNA Polimerase II/genética , RNA Polimerase II/imunologia , Fator de Transcrição STAT2/genética , Fator de Transcrição STAT2/imunologia , Infecção por Zika virus/patologia
11.
Int J Mol Sci ; 19(6)2018 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-29857509

RESUMO

As a typical product in the Miallard reaction, research on the quantitative detection of furosine is abundant, while its bioactivities and toxic effects are still unclear. Our own work recently demonstrated the induction of furosine on apoptosis in HepG2 cells, while the related mechanism remained elusive. In this study, the effects of furosine on cell viability and apoptosis were detected to select the proper dosage, and transcriptomics detection and data analysis were performed to screen out the special genes. Additionally, SiRNA fragments of the selected genes were designed and transfected into HepG2 cells to validate the role of these genes in inducing apoptosis. Results showed that furosine inhibited cell viability and induced cell apoptosis in a dose-dependent manner, as well as activated expressions of the selected genes STAT1 (signal transducer and activator of transcription 1), STAT2 (signal transducer and activator of transcription 2), UBA7 (ubiquitin-like modifier activating enzyme 7), and UBE2L6 (ubiquitin-conjugating enzyme E2L6), which significantly affected downstream apoptosis factors Caspase-3 (cysteinyl aspartate specific proteinase-3), Bcl-2 (B-cell lymphoma gene-2), Bax (BCL2-Associated gene X), and Caspase-9 (cysteinyl aspartate specific proteinase-9). For the first time, we revealed furosine induced apoptosis through two transcriptional regulators (STAT1 and STAT2) and two ubiquitination-related enzymes (UBA7 and UBE2L6).


Assuntos
Apoptose/efeitos dos fármacos , Apoptose/genética , Regulação Neoplásica da Expressão Gênica , Lisina/análogos & derivados , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT2/genética , Enzimas de Conjugação de Ubiquitina/genética , Sobrevivência Celular/efeitos dos fármacos , Perfilação da Expressão Gênica , Células Hep G2 , Humanos , Lisina/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Enzimas de Conjugação de Ubiquitina/metabolismo
12.
Proc Natl Acad Sci U S A ; 115(15): 3906-3911, 2018 04 10.
Artigo em Inglês | MEDLINE | ID: mdl-29581268

RESUMO

In response to IFNß, the IL6 gene is activated, modestly at early times by ISGF3 (IRF9 plus tyrosine-phosphorylated STATs 1 and 2), and strongly at late times by U-ISGF3 (IRF9 plus U-STATs 1 and 2, lacking tyrosine phosphorylation). A classical IFN-stimulated response element (ISRE) at -1,513 to -1,526 in the human IL6 promoter is required. Pretreating cells with IFNß or increasing the expression of U-STAT2 and IRF9 exogenously greatly enhances IL6 expression in response to the classical NF-κB activators IL1, TNF, and LPS. U-STAT2 binds tightly to IRF9, the DNA binding subunit of ISGF3, and also to the p65 subunit of NF-κB. Therefore, as shown by ChIP analyses, U-STAT2 can bridge the ISRE and κB elements in the IL6 promoter. In some cancer cells, the protumorigenic activation of STAT3 will be enhanced by the increased synthesis of IL6 that is facilitated by high expression of U-STAT2 and IRF9.


Assuntos
Fator Gênico 3 Estimulado por Interferon, Subunidade gama/metabolismo , Interleucina-6/genética , NF-kappa B/metabolismo , Fator de Transcrição STAT2/metabolismo , Regulação da Expressão Gênica , Humanos , Interferon beta/genética , Interferon beta/metabolismo , Interleucina-6/metabolismo , NF-kappa B/genética , Fosforilação , Regiões Promotoras Genéticas , Elementos de Resposta , Fator de Transcrição STAT2/genética , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Regulação para Cima
13.
Anim Genet ; 49(3): 254-258, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29572894

RESUMO

Among swine reproductive traits, sow lifetime productivity (SLP) is considered a profitable trait in commercial pig farming. Notably, longevity and efficiency in SLP can be adopted as the key phenotype representing SLP. In this study, we conducted a co-association network analysis using results from a genome-wide association study for SLP-related traits. A total of 656 purebred Landrace female pigs were genotyped using a 60K SNP array. Significantly associated SNPs identified from the GWAS were annotated for the specific genes. Then, we constructed an association weight matrix to build a network based on the co-associations between the genes and 10 SLP traits. The entire network consisted of 495 nodes and 37 755 significant edges. We identified three key regulatory transcription factors: STAT2 (signal transducer and activator of transcription 2), MYF6 (myogenic factor 6) and TFCP2L1 (transcription factor CP2 like 1). The network revealed that the STAT2 and MYF6 regulatory modules cooperate with each other and specifically influence the longevity and efficiency of sows, whereas the TFCP2L1 family specifically affects the improvement of litter size.


Assuntos
Fertilidade/genética , Redes Reguladoras de Genes , Sus scrofa/genética , Animais , Cruzamento , Feminino , Estudos de Associação Genética/veterinária , Genótipo , Fatores de Regulação Miogênica/genética , Fenótipo , Polimorfismo de Nucleotídeo Único , Gravidez , Proteínas Repressoras/genética , Fator de Transcrição STAT2/genética , Sus scrofa/fisiologia
14.
Proc Natl Acad Sci U S A ; 115(4): E601-E609, 2018 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-29317535

RESUMO

Cytokine signaling through the JAK/STAT pathway controls multiple cellular responses including growth, survival, differentiation, and pathogen resistance. An expansion in the gene regulatory repertoire controlled by JAK/STAT signaling occurs through the interaction of STATs with IRF transcription factors to form ISGF3, a complex that contains STAT1, STAT2, and IRF9 and regulates expression of IFN-stimulated genes. ISGF3 function depends on selective interaction between IRF9, through its IRF-association domain (IAD), with the coiled-coil domain (CCD) of STAT2. Here, we report the crystal structures of the IRF9-IAD alone and in a complex with STAT2-CCD. Despite similarity in the overall structure among respective paralogs, the surface features of the IRF9-IAD and STAT2-CCD have diverged to enable specific interaction between these family members. We derive a model for the ISGF3 complex bound to an ISRE DNA element and demonstrate that the observed interface between STAT2 and IRF9 is required for ISGF3 function in cells.


Assuntos
Fator Gênico 3 Estimulado por Interferon, Subunidade gama/metabolismo , Fator de Transcrição STAT2/metabolismo , Animais , Regulação da Expressão Gênica , Células HEK293 , Humanos , Fator Gênico 3 Estimulado por Interferon, Subunidade gama/genética , Janus Quinases/metabolismo , Camundongos , Mutação Puntual , Domínios Proteicos , Fator de Transcrição STAT2/genética , Transdução de Sinais
15.
Nat Immunol ; 19(1): 41-52, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29242538

RESUMO

Prolonged activation of interferon-STAT1 signaling is closely related to inflammatory autoimmune disorders, and therefore the identification of negative regulators of these pathways is important. Through high-content screening of 115 mouse RING-domain E3 ligases, we identified the E3 ubiquitin ligase RNF2 as a potent inhibitor of interferon-dependent antiviral responses. RNF2 deficiency substantially enhanced interferon-stimulated gene (ISG) expression and antiviral responses. Mechanistically, nuclear RNF2 directly bound to STAT1 after interferon stimulation and increased K33-linked polyubiquitination of the DNA-binding domain of STAT1 at position K379, in addition to promoting the disassociation of STAT1/STAT2 from DNA and consequently suppressing ISG transcription. Our study provides insight into the regulation of interferon-dependent responses via a previously unrecognized post-translational modification of STAT1 in the nucleus.


Assuntos
DNA/metabolismo , Interferon Tipo I/farmacologia , Lisina/metabolismo , Complexo Repressor Polycomb 1/metabolismo , Fator de Transcrição STAT1/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Antivirais/farmacologia , Linhagem Celular , Expressão Gênica/efeitos dos fármacos , Lisina/genética , Macrófagos/metabolismo , Macrófagos/virologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Complexo Repressor Polycomb 1/genética , Ligação Proteica/efeitos dos fármacos , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT2/genética , Fator de Transcrição STAT2/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitinação/efeitos dos fármacos , Estomatite Vesicular/genética , Estomatite Vesicular/prevenção & controle , Estomatite Vesicular/virologia , Vírus da Estomatite Vesicular Indiana/fisiologia
16.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 33(9): 1261-1265, 2017 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-29089085

RESUMO

Objective To investigate the expressions of the inhibitor of apoptosis protein survivin and signal transducer and activator of transcription 2 (STAT2) in the tissues of the plaque-like lesions of patients with psoriasis vulgaris (PV) in the progressive stage and discuss their correlation. Methods We collected the plaque-like lesion samples and non-lesion samples from 22 PV patients and the normal skin sample from 18 healthy controls. Real-time quantitative PCR was used to evaluate the mRNA levels of survivin and STAT2 in these samples. Western blot analysis was performed to determine the protein expressions of survivin and STAT2. Immunohistochemical staining was also used to detect the expressions of survivin and STAT2. The correlation between survivin and STAT2 in the PV lesions was determined by Pearson's correlation analysis. Results Compared with healthy control skin and PV non-lesion tissues, the mRNA and protein expressions of survivin and STAT2 were significantly elevated in PV lesion tissues, and the mRNA expression of survivin was positively correlated with STAT2 mRNA in PV lesion tissues (r=0.5282). Conclusion The expressions of survivin and STAT2 are up-regulated in skin lesions of PV patients, and their mRNA expressions are positively correlated.


Assuntos
Proteínas Inibidoras de Apoptose/genética , Psoríase/metabolismo , Fator de Transcrição STAT2/genética , Pele/metabolismo , Adulto , Feminino , Regulação da Expressão Gênica , Humanos , Proteínas Inibidoras de Apoptose/análise , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/análise , Fator de Transcrição STAT2/análise , Survivina
17.
J Clin Invest ; 127(12): 4415-4420, 2017 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-29106381

RESUMO

Primary immunodeficiencies are often monogenic disorders characterized by vulnerability to specific infectious pathogens. Here, we performed whole-exome sequencing of a patient with disseminated Mycobacterium abscessus, Streptococcus viridians bacteremia, and cytomegalovirus (CMV) viremia and identified mutations in 2 genes that regulate distinct IFN pathways. The patient had a homozygous frameshift deletion in IFNGR2, which encodes the signal transducing chain of the IFN-γ receptor, that resulted in minimal protein expression and abolished downstream signaling. The patient also harbored a homozygous deletion in IFNAR1 (IFNAR1*557Gluext*46), which encodes the IFN-α receptor signaling subunit. The IFNAR1*557Gluext*46 resulted in replacement of the stop codon with 46 additional codons at the C-terminus. The level of IFNAR1*557Gluext*46 mutant protein expressed in patient fibroblasts was comparable to levels of WT IFNAR1 in control fibroblasts. IFN-α-induced signaling was impaired in the patient fibroblasts, as evidenced by decreased STAT1/STAT2 phosphorylation, nuclear translocation of STAT1, and expression of IFN-α-stimulated genes critical for CMV immunity. Pretreatment with IFN-α failed to suppress CMV protein expression in patient fibroblasts, whereas expression of WT IFNAR1 restored IFN-α-mediated suppression of CMV. This study identifies a human IFNAR1 mutation and describes a digenic immunodeficiency specific to type I and type II IFNs.


Assuntos
Doenças Genéticas Inatas/genética , Doenças Genéticas Inatas/imunologia , Síndromes de Imunodeficiência/genética , Síndromes de Imunodeficiência/imunologia , Mutação , Receptor de Interferon alfa e beta , Receptores de Interferon , Bacteriemia/genética , Bacteriemia/imunologia , Bacteriemia/microbiologia , Citomegalovirus/imunologia , Infecções por Citomegalovirus/genética , Infecções por Citomegalovirus/imunologia , Feminino , Fibroblastos/imunologia , Fibroblastos/microbiologia , Fibroblastos/virologia , Doenças Genéticas Inatas/microbiologia , Doenças Genéticas Inatas/virologia , Humanos , Síndromes de Imunodeficiência/microbiologia , Síndromes de Imunodeficiência/virologia , Masculino , Infecções por Micobactéria não Tuberculosa/genética , Infecções por Micobactéria não Tuberculosa/imunologia , Mycobacterium abscessus/imunologia , Fosforilação/genética , Fosforilação/imunologia , Receptor de Interferon alfa e beta/genética , Receptor de Interferon alfa e beta/imunologia , Receptores de Interferon/genética , Receptores de Interferon/imunologia , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/imunologia , Fator de Transcrição STAT2/genética , Fator de Transcrição STAT2/imunologia , Infecções Estreptocócicas/genética , Infecções Estreptocócicas/imunologia , Viremia/genética , Viremia/imunologia , Viremia/virologia , Estreptococos Viridans/imunologia
18.
J Virol ; 91(21)2017 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-28835499

RESUMO

Henipaviruses, such as Nipah (NiV) and Hendra (HeV) viruses, are highly pathogenic zoonotic agents within the Paramyxoviridae family. The phosphoprotein (P) gene products of the paramyxoviruses have been well characterized for their interferon (IFN) antagonist activity and their contribution to viral pathogenicity. In this study, we demonstrated that the nucleoprotein (N) of henipaviruses also prevents the host IFN signaling response. Reporter assays demonstrated that the NiV and HeV N proteins (NiV-N and HeV-N, respectively) dose-dependently suppressed both type I and type II IFN responses and that the inhibitory effect was mediated by their core domains. Additionally, NiV-N prevented the nuclear transport of signal transducer and activator of transcription 1 (STAT1) and STAT2. However, NiV-N did not associate with Impα5, Impß1, or Ran, which are members of the nuclear transport system for STATs. Although P protein is known as a binding partner of N protein and actively retains N protein in the cytoplasm, the IFN antagonist activity of N protein was not abolished by the coexpression of P protein. This suggests that the IFN inhibition by N protein occurs in the cytoplasm. Furthermore, we demonstrated that the complex formation of STATs was hampered in the N protein-expressing cells. As a result, STAT nuclear accumulation was reduced, causing a subsequent downregulation of interferon-stimulated genes (ISGs) due to low promoter occupancy by STAT complexes. This novel route for preventing host IFN responses by henipavirus N proteins provides new insight into the pathogenesis of these viruses.IMPORTANCE Paramyxoviruses are well known for suppressing interferon (IFN)-mediated innate immunity with their phosphoprotein (P) gene products, and the henipaviruses also possess P, V, W, and C proteins for evading host antiviral responses. There are numerous studies providing evidence for the relationship between viral pathogenicity and antagonistic activities against IFN responses by P gene products. Meanwhile, little attention has been paid to the influence of nucleoprotein (N) on host innate immune responses. In this study, we demonstrated that both the NiV and HeV N proteins have antagonistic activity against the JAK/STAT signaling pathway by preventing the nucleocytoplasmic trafficking of STAT1 and STAT2. This inhibitory effect is due to an impairment of the ability of STATs to form complexes. These results provide new insight into the involvement of N protein in viral pathogenicity via its IFN antagonism.


Assuntos
Núcleo Celular/metabolismo , Vírus Hendra/fisiologia , Infecções por Henipavirus/metabolismo , Vírus Nipah/fisiologia , Nucleoproteínas/metabolismo , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT2/metabolismo , Núcleo Celular/genética , Células HEK293 , Células HeLa , Infecções por Henipavirus/imunologia , Infecções por Henipavirus/virologia , Humanos , Imunidade Inata/imunologia , Nucleoproteínas/genética , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT2/genética , Transdução de Sinais
19.
Biochemistry ; 56(32): 4145-4153, 2017 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-28707474

RESUMO

A significant fraction of the eukaryotic proteome consists of proteins that are either partially or completely disordered under native-like conditions. Intrinsically disordered proteins (IDPs) are common in protein-protein interactions and are involved in numerous cellular processes. Although many proteins have been identified as disordered, much less is known about the binding mechanisms of the coupled binding and folding reactions involving IDPs. Here we have analyzed the rate-limiting transition state for binding between the TAZ1 domain of CREB binding protein and the intrinsically disordered transactivation domain of STAT2 (TAD-STAT2) by site-directed mutagenesis and kinetic experiments (Φ-value analysis) and found that the native protein-protein binding interface is not formed at the transition state for binding. Instead, native hydrophobic binding interactions form late, after the rate-limiting barrier has been crossed. The association rate constant in the absence of electrostatic enhancement was determined to be rather high. This is consistent with the Φ-value analysis, which showed that there are few or no obligatory native contacts. Also, linear free energy relationships clearly demonstrate that native interactions are cooperatively formed, a scenario that has usually been observed for proteins that fold according to the so-called nucleation-condensation mechanism. Thus, native hydrophobic binding interactions at the rate-limiting transition state for association between TAD-STAT2 and TAZ1 are not a requirement, which is generally in agreement with previous findings on other IDP systems and might be a common mechanism for IDPs.


Assuntos
Proteínas Intrinsicamente Desordenadas/química , Modelos Químicos , Fragmentos de Peptídeos/química , Fator de Transcrição STAT2/química , Sialoglicoproteínas/química , Humanos , Interações Hidrofóbicas e Hidrofílicas , Proteínas Intrinsicamente Desordenadas/genética , Proteínas Intrinsicamente Desordenadas/metabolismo , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Ligação Proteica , Domínios Proteicos , Fator de Transcrição STAT2/genética , Fator de Transcrição STAT2/metabolismo , Sialoglicoproteínas/genética , Sialoglicoproteínas/metabolismo
20.
Tumour Biol ; 39(7): 1010428317708546, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28714361

RESUMO

In recent years, abnormal RNA editing has been shown to play an important role in the development of esophageal squamous cell carcinoma, as such abnormal editing is catalyzed by ADAR (adenosine deaminases acting on RNA). However, the regulatory mechanism of ADAR1 in esophageal squamous cell carcinomas remains largely unknown. In this study, we investigated ADAR1 expression and its association with RNA editing in esophageal squamous cell carcinomas. RNA sequencing applied to esophageal squamous cell carcinoma clinical samples showed that ADAR1 expression was correlated with the expression of STAT1, STAT2, and IRF9. In vitro experiments showed that the abundance of ADAR1 protein was associated with the induced activation of the JAK/STAT pathway by type I interferon. RNA sequencing results showed that treatment with type I interferon caused an increase in the number and degree of RNA editing in esophageal squamous cell carcinoma cell lines. In conclusion, the activation of the JAK/STAT pathway is a regulatory mechanism of ADAR1 expression and causes abnormal RNA editing profile in esophageal squamous cell carcinoma. This mechanism may serve as a new target for esophageal squamous cell carcinoma therapy.


Assuntos
Adenosina Desaminase/genética , Carcinoma de Células Escamosas/genética , Neoplasias Esofágicas/genética , Interferon Tipo I/genética , Proteínas de Ligação a RNA/genética , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Fator Gênico 3 Estimulado por Interferon, Subunidade gama/genética , Edição de RNA/genética , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT2/genética , Transdução de Sinais
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