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1.
Cell Rep ; 36(2): 109364, 2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34214467

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) variants govern transmissibility, responsiveness to vaccination, and disease severity. In a screen for new models of SARS-CoV-2 infection, we identify human H522 lung adenocarcinoma cells as naturally permissive to SARS-CoV-2 infection despite complete absence of angiotensin-converting enzyme 2 (ACE2) expression. Remarkably, H522 infection requires the E484D S variant; viruses expressing wild-type S are not infectious. Anti-S monoclonal antibodies differentially neutralize SARS-CoV-2 E484D S in H522 cells as compared to ACE2-expressing cells. Sera from vaccinated individuals block this alternative entry mechanism, whereas convalescent sera are less effective. Although the H522 receptor remains unknown, depletion of surface heparan sulfates block H522 infection. Temporally resolved transcriptomic and proteomic profiling reveal alterations in cell cycle and the antiviral host cell response, including MDA5-dependent activation of type I interferon signaling. These findings establish an alternative SARS-CoV-2 host cell receptor for the E484D SARS-CoV-2 variant, which may impact tropism of SARS-CoV-2 and consequently human disease pathogenesis.


Assuntos
COVID-19/imunologia , COVID-19/metabolismo , Receptores Virais , Glicoproteína da Espícula de Coronavírus/metabolismo , Substituição de Aminoácidos , Enzima de Conversão de Angiotensina 2 , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Ciclo Celular , Linhagem Celular Tumoral , Chlorocebus aethiops , Perfilação da Expressão Gênica , Heparitina Sulfato/metabolismo , Humanos , Interferon Tipo I/metabolismo , Helicase IFIH1 Induzida por Interferon/metabolismo , Modelos Biológicos , Ligação Proteica , Domínios Proteicos , Proteômica , Receptores Virais/metabolismo , SARS-CoV-2 , Serina Endopeptidases/metabolismo , Transdução de Sinais , Glicoproteína da Espícula de Coronavírus/genética , Células Vero , Internalização do Vírus , Replicação Viral
2.
Nat Commun ; 12(1): 4175, 2021 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-34234126

RESUMO

Although we can now measure single-cell signaling responses with multivariate, high-throughput techniques our ability to interpret such measurements is still limited. Even interpretation of dose-response based on single-cell data is not straightforward: signaling responses can differ significantly between cells, encompass multiple signaling effectors, and have dynamic character. Here, we use probabilistic modeling and information-theory to introduce fractional response analysis (FRA), which quantifies changes in fractions of cells with given response levels. FRA can be universally performed for heterogeneous, multivariate, and dynamic measurements and, as we demonstrate, quantifies otherwise hidden patterns in single-cell data. In particular, we show that fractional responses to type I interferon in human peripheral blood mononuclear cells are very similar across different cell types, despite significant differences in mean or median responses and degrees of cell-to-cell heterogeneity. Further, we demonstrate that fractional responses to cytokines scale linearly with the log of the cytokine dose, which uncovers that heterogeneous cellular populations are sensitive to fold-changes in the dose, as opposed to additive changes.


Assuntos
Ensaios de Triagem em Larga Escala/métodos , Interferon Tipo I/metabolismo , Leucócitos Mononucleares/metabolismo , Modelos Imunológicos , Células 3T3 , Animais , Voluntários Saudáveis , Humanos , Interferon Tipo I/imunologia , Leucócitos Mononucleares/imunologia , Camundongos , Modelos Estatísticos , Cultura Primária de Células , Transdução de Sinais/imunologia , Análise de Célula Única , Software
3.
Int J Mol Sci ; 22(14)2021 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-34299101

RESUMO

Children with the new coronavirus disease 2019 (COVID-19) have milder symptoms and a better prognosis than adult patients. Several investigations assessed type I, II, and III interferon (IFN) signatures in SARS-CoV-2 infected adults, however no data are available for pediatric patients. TRIM28 and SETDB1 regulate the transcription of multiple genes involved in the immune response as well as of human endogenous retroviruses (HERVs). Exogenous viral infections can trigger the activation of HERVs, which in turn can induce inflammatory and immune reactions. Despite the potential cross-talks between SARS-CoV-2 infection and TRIM28, SETDB1, and HERVs, information on their expressions in COVID-19 patients is lacking. We assessed, through a PCR real time Taqman amplification assay, the transcription levels of six IFN-I stimulated genes, IFN-II and three of its sensitive genes, three IFN-lIIs, as well as of TRIM28, SETDB1, pol genes of HERV-H, -K, and -W families, and of env genes of Syncytin (SYN)1, SYN2, and multiple sclerosis-associated retrovirus (MRSV) in peripheral blood from COVID-19 children and in control uninfected subjects. Higher expression levels of IFN-I and IFN-II inducible genes were observed in 36 COVID-19 children with mild or moderate disease as compared to uninfected controls, whereas their concentrations decreased in 17 children with severe disease and in 11 with multisystem inflammatory syndrome (MIS-C). Similar findings were found for the expression of TRIM-28, SETDB1, and every HERV gene. Positive correlations emerged between the transcriptional levels of type I and II IFNs, TRIM28, SETDB1, and HERVs in COVID-19 patients. IFN-III expressions were comparable in each group of subjects. This preserved induction of IFN-λs could contribute to the better control of the infection in children as compared to adults, in whom IFN-III deficiency has been reported. The upregulation of IFN-I, IFN-II, TRIM28, SETDB1, and HERVs in children with mild symptoms, their declines in severe cases or with MIS-C, and the positive correlations of their transcription in SARS-CoV-2-infected children suggest that they may play important roles in conditioning the evolution of the infection.


Assuntos
COVID-19/epidemiologia , COVID-19/metabolismo , Retrovirus Endógenos/metabolismo , SARS-CoV-2/isolamento & purificação , Índice de Gravidade de Doença , COVID-19/patologia , COVID-19/virologia , Estudos de Casos e Controles , Criança , Retrovirus Endógenos/genética , Feminino , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Interferon Tipo I/genética , Interferon Tipo I/metabolismo , Interferon gama/genética , Interferon gama/metabolismo , Interferons/genética , Interferons/metabolismo , Itália/epidemiologia , Masculino , Proteína 28 com Motivo Tripartido/genética , Proteína 28 com Motivo Tripartido/metabolismo
4.
Front Immunol ; 12: 658428, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34149696

RESUMO

SARS-CoV-2 virus causes upper and lower respiratory diseases including pneumonia, and in some cases, leads to lethal pulmonary failure. Angiotensin converting enzyme-2 (ACE2), the receptor for cellular entry of SARS-CoV-2 virus, has been shown to protect against severe acute lung failure. Here, we provide evidence that SARS-CoV-2 spike protein S1 reduced the mRNA expression of ACE2 and type I interferons in primary cells of lung bronchoalveolar lavage (BAL) from naïve rhesus macaques. The expression levels of ACE2 and type I interferons were also found to be correlated with each other, consistent with the recent finding that ACE2 is an interferon-inducible gene. Furthermore, induction of ACE2 and type I interferons by poly I:C, an interferon inducer, was suppressed by S1 protein in primary cells of BAL. These observations suggest that the downregulation of ACE2 and type I interferons induced by S1 protein may directly contribute to SARS-CoV-2-associated lung diseases.


Assuntos
Enzima de Conversão de Angiotensina 2/metabolismo , COVID-19 , Interferon Tipo I/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Animais , Líquido da Lavagem Broncoalveolar/citologia , Macaca mulatta , SARS-CoV-2
5.
Proc Natl Acad Sci U S A ; 118(26)2021 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-34108245

RESUMO

Patients with severe COVID-19 infection exhibit a low level of oxygen in affected tissue and blood. To understand the pathophysiology of COVID-19 infection, it is therefore necessary to understand cell function during hypoxia. We investigated aspects of human monocyte activation under hypoxic conditions. HMGB1 is an alarmin released by stressed cells. Under normoxic conditions, HMGB1 activates interferon regulatory factor (IRF)5 and nuclear factor-κB in monocytes, leading to expression of type I interferon (IFN) and inflammatory cytokines including tumor necrosis factor α, and interleukin 1ß, respectively. When hypoxic monocytes are activated by HMGB1, they produce proinflammatory cytokines but fail to produce type I IFN. Hypoxia-inducible factor-1α, induced by hypoxia, functions as a direct transcriptional repressor of IRF5 and IRF3. As hypoxia is a stressor that induces secretion of HMGB1 by epithelial cells, hypoxia establishes a microenvironment that favors monocyte production of inflammatory cytokines but not IFN. These findings have implications for the pathogenesis of COVID-19.


Assuntos
Hipóxia Celular/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Monócitos/imunologia , COVID-19/imunologia , Células Cultivadas , Citocinas/imunologia , Humanos , Fatores Reguladores de Interferon/metabolismo , Interferon Tipo I/imunologia , Interferon Tipo I/metabolismo , Interleucina-1beta/metabolismo , Monócitos/metabolismo , NF-kappa B/imunologia , NF-kappa B/metabolismo , Oxigênio/metabolismo , SARS-CoV-2/imunologia , Fator de Necrose Tumoral alfa/metabolismo
6.
J Virol ; 95(13): e0026621, 2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-34110264

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the ongoing coronavirus disease 2019 (COVID-19) pandemic. While previous studies have shown that several SARS-CoV-2 proteins can antagonize the interferon (IFN) response, some of the mechanisms by which they do so are not well understood. In this study, we describe two novel mechanisms by which SARS-CoV-2 blocks the IFN pathway. Type I IFNs and IFN-stimulated genes (ISGs) were poorly induced during SARS-CoV-2 infection, and once infection was established, cells were highly resistant to ectopic induction of IFNs and ISGs. Levels of two key IFN signaling pathway components, Tyk2 and STAT2, were significantly lower in SARS-CoV-2-infected cells. Expression of nonstructural protein 1 (NSP1) or nucleocapsid in the absence of other viral proteins was sufficient to block IFN induction, but only NSP1 was able to inhibit IFN signaling. Mapping studies suggest that NSP1 prevents IFN induction in part by blocking IRF3 phosphorylation. In addition, NSP1-induced depletion of Tyk2 and STAT2 dampened ISG induction. Together, our data provide new insights into how SARS-CoV-2 successfully evades the IFN system to establish infection. IMPORTANCE SARS-CoV-2 is the causative agent of COVID-19, a serious disease that can have a myriad of symptoms from loss of taste and smell to pneumonia and hypercoagulation. The rapid spread of SARS-CoV-2 can be attributed in part to asymptomatic transmission, where infected individuals shed large amounts of virus before the onset of disease. This is likely due to the ability of SARS-CoV-2 to effectively suppress the innate immune system, including the IFN response. Indeed, we show that the IFN response is efficiently blocked during SARS-CoV-2 infection, a process that is mediated in large part by nonstructural protein 1 and nucleocapsid. Our study provides new insights on how SARS-CoV-2 evades the IFN response to successfully establish infection. These findings should be considered for the development and administration of therapeutics against SARS-CoV-2.


Assuntos
Interferon Tipo I/antagonistas & inibidores , SARS-CoV-2/metabolismo , Transdução de Sinais , Proteínas não Estruturais Virais/metabolismo , Animais , COVID-19/imunologia , COVID-19/virologia , Chlorocebus aethiops , Proteínas do Nucleocapsídeo de Coronavírus/metabolismo , Células HEK293 , Humanos , Imunidade Inata , Fator Regulador 3 de Interferon/metabolismo , Interferon Tipo I/metabolismo , Fosfoproteínas/metabolismo , SARS-CoV-2/patogenicidade , Fator de Transcrição STAT2/metabolismo , TYK2 Quinase/metabolismo , Células Vero
7.
Cell Host Microbe ; 29(7): 1052-1062, 2021 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-34022154

RESUMO

COVID-19 can result in severe disease characterized by significant immunopathology that is spurred by an exuberant, yet dysregulated, innate immune response with a poor adaptive response. A limited and delayed interferon I (IFN-I) and IFN-III response results in exacerbated proinflammatory cytokine production and in extensive cellular infiltrates in the respiratory tract, resulting in lung pathology. The development of effective therapeutics for patients with severe COVID-19 depends on our understanding of the pathological elements of this unbalanced innate immune response. Here, we review the mechanisms by which SARS-CoV-2 both activates and antagonizes the IFN and inflammatory response following infection, how a dysregulated cytokine and cellular response contributes to immune-mediated pathology in COVID-19, and therapeutic strategies that target elements of the innate response.


Assuntos
COVID-19/imunologia , Imunidade Inata/imunologia , Interferons/uso terapêutico , SARS-CoV-2/imunologia , Animais , Anti-Inflamatórios/uso terapêutico , COVID-19/tratamento farmacológico , Citocinas/metabolismo , Modelos Animais de Doenças , Humanos , Evasão da Resposta Imune , Interferon Tipo I/metabolismo , Interferons/metabolismo , Cinética
8.
PLoS Pathog ; 17(5): e1009229, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-34029358

RESUMO

While MERS-CoV (Middle East respiratory syndrome Coronavirus) provokes a lethal disease in humans, camelids, the main virus reservoir, are asymptomatic carriers, suggesting a crucial role for innate immune responses in controlling the infection. Experimentally infected camelids clear infectious virus within one week and mount an effective adaptive immune response. Here, transcription of immune response genes was monitored in the respiratory tract of MERS-CoV infected alpacas. Concomitant to the peak of infection, occurring at 2 days post inoculation (dpi), type I and III interferons (IFNs) were maximally transcribed only in the nasal mucosa of alpacas, while interferon stimulated genes (ISGs) were induced along the whole respiratory tract. Simultaneous to mild focal infiltration of leukocytes in nasal mucosa and submucosa, upregulation of the anti-inflammatory cytokine IL10 and dampened transcription of pro-inflammatory genes under NF-κB control were observed. In the lung, early (1 dpi) transcription of chemokines (CCL2 and CCL3) correlated with a transient accumulation of mainly mononuclear leukocytes. A tight regulation of IFNs in lungs with expression of ISGs and controlled inflammatory responses, might contribute to virus clearance without causing tissue damage. Thus, the nasal mucosa, the main target of MERS-CoV in camelids, seems central in driving an efficient innate immune response based on triggering ISGs as well as the dual anti-inflammatory effects of type III IFNs and IL10.


Assuntos
Camelídeos Americanos , Infecções por Coronavirus/imunologia , Interferon Tipo I/metabolismo , Interferons/metabolismo , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Animais , Antivirais/metabolismo , Antivirais/farmacologia , Camelídeos Americanos/imunologia , Camelídeos Americanos/metabolismo , Camelídeos Americanos/virologia , Chlorocebus aethiops , Infecções por Coronavirus/metabolismo , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/veterinária , Reservatórios de Doenças/veterinária , Resistência à Doença/efeitos dos fármacos , Resistência à Doença/genética , Resistência à Doença/imunologia , Regulação da Expressão Gênica , Imunidade Inata/fisiologia , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/veterinária , Inflamação/virologia , Interferon Tipo I/genética , Interferon Tipo I/farmacologia , Interferons/genética , Interferons/farmacologia , Coronavírus da Síndrome Respiratória do Oriente Médio/efeitos dos fármacos , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Mucosa Nasal/efeitos dos fármacos , Mucosa Nasal/imunologia , Mucosa Nasal/metabolismo , Mucosa Nasal/virologia , Sistema Respiratório/efeitos dos fármacos , Sistema Respiratório/imunologia , Sistema Respiratório/metabolismo , Sistema Respiratório/virologia , Células Vero , Carga Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos
9.
Cell Rep ; 35(7): 109126, 2021 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-33974846

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) evades most innate immune responses but may still be vulnerable to some. Here, we systematically analyze the impact of SARS-CoV-2 proteins on interferon (IFN) responses and autophagy. We show that SARS-CoV-2 proteins synergize to counteract anti-viral immune responses. For example, Nsp14 targets the type I IFN receptor for lysosomal degradation, ORF3a prevents fusion of autophagosomes and lysosomes, and ORF7a interferes with autophagosome acidification. Most activities are evolutionarily conserved. However, SARS-CoV-2 Nsp15 antagonizes IFN signaling less efficiently than the orthologs of closely related RaTG13-CoV and SARS-CoV-1. Overall, SARS-CoV-2 proteins counteract autophagy and type I IFN more efficiently than type II or III IFN signaling, and infection experiments confirm potent inhibition by IFN-γ and -λ1. Our results define the repertoire and selected mechanisms of SARS-CoV-2 innate immune antagonists but also reveal vulnerability to type II and III IFN that may help to develop safe and effective anti-viral approaches.


Assuntos
COVID-19/virologia , SARS-CoV-2/imunologia , Proteínas Virais/imunologia , Animais , Antivirais/farmacologia , Autofagossomos/imunologia , Autofagia/imunologia , COVID-19/imunologia , Linhagem Celular , Chlorocebus aethiops , Exorribonucleases/imunologia , Células HEK293 , Células HeLa , Humanos , Evasão da Resposta Imune , Imunidade Inata , Interferon Tipo I/metabolismo , Interferons/metabolismo , Receptor de Interferon alfa e beta/antagonistas & inibidores , Receptor de Interferon alfa e beta/imunologia , SARS-CoV-2/patogenicidade , Células Vero , Proteínas não Estruturais Virais/imunologia
10.
Nat Commun ; 12(1): 2970, 2021 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-34016972

RESUMO

Activation of MAVS, an adaptor molecule in Rig-I-like receptor (RLR) signaling, is indispensable for antiviral immunity, yet the molecular mechanisms modulating MAVS activation are not completely understood. Ubiquitination has a central function in regulating the activity of MAVS. Here, we demonstrate that a mitochondria-localized deubiquitinase USP18 specifically interacts with MAVS, promotes K63-linked polyubiquitination and subsequent aggregation of MAVS. USP18 upregulates the expression and production of type I interferon following infection with Sendai virus (SeV) or Encephalomyocarditis virus (EMCV). Mice with a deficiency of USP18 are more susceptible to RNA virus infection. USP18 functions as a scaffold protein to facilitate the re-localization of TRIM31 and enhances the interaction between TRIM31 and MAVS in mitochondria. Our results indicate that USP18 functions as a post-translational modulator of MAVS-mediated antiviral signaling.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Infecções por Cardiovirus/imunologia , Infecções por Respirovirus/imunologia , Ubiquitina Tiolesterase/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/isolamento & purificação , Animais , Infecções por Cardiovirus/virologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Vírus da Encefalomiocardite/imunologia , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Imunidade Inata , Interferon Tipo I/metabolismo , Lisina/metabolismo , Masculino , Camundongos , Camundongos Knockout , Processamento de Proteína Pós-Traducional/imunologia , Células RAW 264.7 , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Infecções por Respirovirus/virologia , Vírus Sendai/imunologia , Transdução de Sinais/imunologia , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina Tiolesterase/genética , Ubiquitina Tiolesterase/isolamento & purificação , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação/imunologia
11.
Nat Commun ; 12(1): 2681, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33976210

RESUMO

Innate immune cells are critical in protective immunity against viral infections, involved in sensing foreign viral nucleic acids. Here we report that the poly(ADP-ribose) polymerase 9 (PARP9), a member of PARP family, serves as a non-canonical sensor for RNA virus to initiate and amplify type I interferon (IFN) production. We find knockdown or deletion of PARP9 in human or mouse dendritic cells and macrophages inhibits type I IFN production in response to double strand RNA stimulation or RNA virus infection. Furthermore, mice deficient for PARP9 show enhanced susceptibility to infections with RNA viruses because of the impaired type I IFN production. Mechanistically, we show that PARP9 recognizes and binds viral RNA, with resultant recruitment and activation of the phosphoinositide 3-kinase (PI3K) and AKT3 pathway, independent of mitochondrial antiviral-signaling (MAVS). PI3K/AKT3 then activates the IRF3 and IRF7 by phosphorylating IRF3 at Ser385 and IRF7 at Ser437/438 mediating type I IFN production. Together, we reveal a critical role for PARP9 as a non-canonical RNA sensor that depends on the PI3K/AKT3 pathway to produce type I IFN. These findings may have important clinical implications in controlling viral infections and viral-induced diseases by targeting PARP9.


Assuntos
Células Dendríticas/enzimologia , Proteínas de Neoplasias/metabolismo , Poli(ADP-Ribose) Polimerases/metabolismo , Infecções por Vírus de RNA/enzimologia , RNA Viral/metabolismo , Animais , Chlorocebus aethiops , Células Dendríticas/virologia , Humanos , Fator Regulador 3 de Interferon/metabolismo , Fator Regulador 7 de Interferon/metabolismo , Interferon Tipo I/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Proteínas de Neoplasias/genética , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Poli(ADP-Ribose) Polimerases/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Infecções por Vírus de RNA/virologia , Vírus de RNA/genética , Vírus de RNA/fisiologia , Transdução de Sinais , Células THP-1 , Células Vero
12.
Blood ; 138(1): 91-103, 2021 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-33881503

RESUMO

Intracerebral hemorrhage associated with thrombolytic therapy with tissue plasminogen activator (tPA) in acute ischemic stroke continues to present a major clinical problem. Here, we report that infusion of tPA resulted in a significant increase in markers of neutrophil extracellular traps (NETs) in the ischemic cortex and plasma of mice subjected to photothrombotic middle cerebral artery occlusion. Peptidylarginine deiminase 4 (PAD4), a critical enzyme for NET formation, is also significantly upregulated in the ischemic brains of tPA-treated mice. Blood-brain barrier (BBB) disruption after ischemic challenge in an in vitro model of BBB was exacerbated after exposure to NETs. Importantly, disruption of NETs by DNase I or inhibition of NET production by PAD4 deficiency restored tPA-induced loss of BBB integrity and consequently decreased tPA-associated brain hemorrhage after ischemic stroke. Furthermore, either DNase I or PAD4 deficiency reversed tPA-mediated upregulation of the DNA sensor cyclic GMP-AMP (cGAMP) synthase (cGAS). Administration of cGAMP after stroke abolished DNase I-mediated downregulation of the STING pathway and type 1 interferon production and blocked the antihemorrhagic effect of DNase I in tPA-treated mice. We also show that tPA-associated brain hemorrhage after ischemic stroke was significantly reduced in cGas-/- mice. Collectively, these findings demonstrate that NETs significantly contribute to tPA-induced BBB breakdown in the ischemic brain and suggest that targeting NETs or cGAS may ameliorate thrombolytic therapy for ischemic stroke by reducing tPA-associated hemorrhage.


Assuntos
Armadilhas Extracelulares/metabolismo , Hemorragias Intracranianas/complicações , Hemorragias Intracranianas/patologia , Nucleotidiltransferases/metabolismo , Acidente Vascular Cerebral/complicações , Animais , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Desoxirribonuclease I/metabolismo , Humanos , Interferon Tipo I/metabolismo , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos C57BL , Infiltração de Neutrófilos , Proteína-Arginina Desiminase do Tipo 4/deficiência , Proteína-Arginina Desiminase do Tipo 4/metabolismo , Transdução de Sinais , Ativador de Plasminogênio Tecidual , Regulação para Cima
14.
Nat Commun ; 12(1): 1980, 2021 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-33790300

RESUMO

The majority of patients with systemic lupus erythematosus (SLE) have high expression of type I IFN-stimulated genes. Mitochondrial abnormalities have also been reported, but the contribution of type I IFN exposure to these changes is unknown. Here, we show downregulation of mitochondria-derived genes and mitochondria-associated metabolic pathways in IFN-High patients from transcriptomic analysis of CD4+ and CD8+ T cells. CD8+ T cells from these patients have enlarged mitochondria and lower spare respiratory capacity associated with increased cell death upon rechallenge with TCR stimulation. These mitochondrial abnormalities can be phenocopied by exposing CD8+ T cells from healthy volunteers to type I IFN and TCR stimulation. Mechanistically these 'SLE-like' conditions increase CD8+ T cell NAD+ consumption resulting in impaired mitochondrial respiration and reduced cell viability, both of which can be rectified by NAD+ supplementation. Our data suggest that type I IFN exposure contributes to SLE pathogenesis by promoting CD8+ T cell death via metabolic rewiring.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Perfilação da Expressão Gênica/métodos , Interferon Tipo I/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Adulto , Idoso , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Feminino , Humanos , Interferon Tipo I/metabolismo , Interferon Tipo I/farmacologia , Lúpus Eritematoso Sistêmico/genética , Lúpus Eritematoso Sistêmico/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Redes e Vias Metabólicas/genética , Pessoa de Meia-Idade , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Adulto Jovem
15.
Cells ; 10(3)2021 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-33801464

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the coronavirus disease 2019 (COVID-19) that has resulted in the current pandemic. The lack of highly efficacious antiviral drugs that can manage this ongoing global emergency gives urgency to establishing a comprehensive understanding of the molecular pathogenesis of SARS-CoV-2. We characterized the role of the nucleocapsid protein (N) of SARS-CoV-2 in modulating antiviral immunity. Overexpression of SARS-CoV-2 N resulted in the attenuation of retinoic acid inducible gene-I (RIG-I)-like receptor-mediated interferon (IFN) production and IFN-induced gene expression. Similar to the SARS-CoV-1 N protein, SARS-CoV-2 N suppressed the interaction between tripartate motif protein 25 (TRIM25) and RIG-I. Furthermore, SARS-CoV-2 N inhibited polyinosinic: polycytidylic acid [poly(I:C)]-mediated IFN signaling at the level of Tank-binding kinase 1 (TBK1) and interfered with the association between TBK1 and interferon regulatory factor 3 (IRF3), subsequently preventing the nuclear translocation of IRF3. We further found that both type I and III IFN production induced by either the influenza virus lacking the nonstructural protein 1 or the Zika virus were suppressed by the SARS-CoV-2 N protein. Our findings provide insights into the molecular function of the SARS-CoV-2 N protein with respect to counteracting the host antiviral immune response.


Assuntos
Proteínas do Nucleocapsídeo de Coronavírus/metabolismo , Proteína DEAD-box 58/metabolismo , Interferons/metabolismo , Receptores Imunológicos/metabolismo , SARS-CoV-2/metabolismo , Proteína DEAD-box 58/genética , Interações Hospedeiro-Patógeno/genética , Humanos , Fator Regulador 3 de Interferon/genética , Fator Regulador 3 de Interferon/metabolismo , Interferon Tipo I/genética , Interferon Tipo I/metabolismo , Interferon gama/genética , Interferon gama/metabolismo , Interferons/genética , Orthomyxoviridae/genética , Orthomyxoviridae/metabolismo , Fosfoproteínas/metabolismo , Poli C/farmacologia , Poli I/farmacologia , Regiões Promotoras Genéticas , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Receptores Imunológicos/genética , SARS-CoV-2/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Fatores de Transcrição/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Regulação para Cima , Zika virus/genética , Zika virus/metabolismo
16.
Exp Mol Med ; 53(4): 483-494, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33828231

RESUMO

The zoonotic coronavirus SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2), which causes COVID-19 (coronavirus disease-2019), has resulted in a pandemic. This has led to an urgent need to understand the molecular determinants of SARS-CoV-2 infection, factors associated with COVID-19 heterogeneity and severity, and therapeutic options for these patients. In this review, we discuss the role of host factors in SARS-CoV-2 infection and describe variations in host factor expression as mechanisms underlying the symptoms and severity of COVID-19. We focus on two host factors, angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2), implicated in SARS-CoV-2 infection. We also discuss genetic variants associated with COVID-19 severity revealed in selected patients and based on genome-wide association studies (GWASs). Furthermore, we highlight important advances in cell and chromatin biology, such as single-cell RNA and chromatin sequencing and chromosomal conformation assays, as methods that may aid in the discovery of viral-host interactions in COVID-19. Understanding how regulation of host factor genes varies in physiological and pathological states might explain the heterogeneity observed in SARS-CoV-2 infection, help identify pathways for therapeutic development, and identify patients most likely to progress to severe COVID-19.


Assuntos
Enzima de Conversão de Angiotensina 2/genética , COVID-19/genética , Interações Hospedeiro-Patógeno/fisiologia , Serina Endopeptidases/genética , Enzima de Conversão de Angiotensina 2/metabolismo , COVID-19/etiologia , Expressão Gênica , Variação Genética , Humanos , Interferon Tipo I/genética , Interferon Tipo I/metabolismo , Pulmão/patologia , Pulmão/virologia , Serina Endopeptidases/metabolismo
17.
Int J Mol Sci ; 22(8)2021 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-33919546

RESUMO

One of the most powerful and multifaceted cytokines produced by immune cells are type I interferons (IFNs), the basal secretion of which contributes to the maintenance of immune homeostasis, while their activation-induced production is essential to effective immune responses. Although, each cell is capable of producing type I IFNs, plasmacytoid dendritic cells (pDCs) possess a unique ability to rapidly produce large amounts of them. Importantly, type I IFNs have a prominent role in the pathomechanism of various pDC-associated diseases. Deficiency in type I IFN production increases the risk of more severe viral infections and the development of certain allergic reactions, and supports tumor resistance; nevertheless, its overproduction promotes autoimmune reactions. Therefore, the tight regulation of type I IFN responses of pDCs is essential to maintain an adequate level of immune response without causing adverse effects. Here, our goal was to summarize those endogenous factors that can influence the type I IFN responses of pDCs, and thus might serve as possible therapeutic targets in pDC-associated diseases. Furthermore, we briefly discuss the current therapeutic approaches targeting the pDC-type I IFN axis in viral infections, cancer, autoimmunity, and allergy, together with their limitations defined by the Janus-faced nature of pDC-derived type I IFNs.


Assuntos
Células Dendríticas/metabolismo , Interferon Tipo I/metabolismo , Animais , Autoimunidade/genética , Autoimunidade/fisiologia , Células Dendríticas/imunologia , Humanos , Imunidade Inata/imunologia , Imunidade Inata/fisiologia , Interferon Tipo I/genética , Transdução de Sinais/genética , Transdução de Sinais/fisiologia
18.
Am J Pathol ; 191(6): 1036-1048, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33753025

RESUMO

Type I interferon (IFN-I) has a well-known function in controlling viral infections, but its contribution in hepatocyte proliferation and hepatocellular carcinoma (HCC) formation remains unclear. Mice deficient in IFN-α receptor expression in whole mice or only in hepatocytes (Ifnar-/- and IfnarΔliver) were used to investigate the role of IFN-I signaling in cell proliferation and cancer formation in the liver. Ifnar-/- mice were resistant to chemical-induced HCC formation in the absence of infection. The results show that low grade of IFN-I and interferon-stimulated gene were expressed substantially in naïve mouse liver. The low level of IFN-I activation is constantly present in mouse liver after weaning and negatively modulates forkhead box O hepatic expression. The IFN-I signaling can be partially blocked by the clearance of lipopolysaccharide. Mice lacking IFN-I signaling have lower basal proliferation activity and delayed liver regeneration processes after two-thirds partial hepatectomy. The activation of IFN-I signaling on hepatocyte controls glucose homeostasis and lipid metabolism to support proliferation potency and long-term tumorigenesis. Our results reveal a positive role of low-grade IFN-I singling to hepatocyte proliferation and HCC formation by modulating glucose homeostasis and lipid metabolism.


Assuntos
Carcinoma Hepatocelular/metabolismo , Hepatócitos/metabolismo , Interferon Tipo I/metabolismo , Neoplasias Hepáticas/metabolismo , Regeneração Hepática/fisiologia , Animais , Proliferação de Células/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais/fisiologia
19.
PLoS Pathog ; 17(3): e1009387, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33690673

RESUMO

The skin innate immune response to methicillin-resistant Staphylococcus aureus (MRSA) culminates in the formation of an abscess to prevent bacterial spread and tissue damage. Pathogen recognition receptors (PRRs) dictate the balance between microbial control and injury. Therefore, intracellular brakes are of fundamental importance to tune the appropriate host defense while inducing resolution. The intracellular inhibitor suppressor of cytokine signaling 1 (SOCS-1), a known JAK/STAT inhibitor, prevents the expression and actions of PRR adaptors and downstream effectors. Whether SOCS-1 is a molecular component of skin host defense remains to be determined. We hypothesized that SOCS-1 decreases type I interferon production and IFNAR-mediated antimicrobial effector functions, limiting the inflammatory response during skin infection. Our data show that MRSA skin infection enhances SOCS-1 expression, and both SOCS-1 inhibitor peptide-treated and myeloid-specific SOCS-1 deficient mice display decreased lesion size, bacterial loads, and increased abscess thickness when compared to wild-type mice treated with the scrambled peptide control. SOCS-1 deletion/inhibition increases phagocytosis and bacterial killing, dependent on nitric oxide release. SOCS-1 inhibition also increases the levels of type I and type II interferon levels in vivo. IFNAR deletion and antibody blockage abolished the beneficial effects of SOCS-1 inhibition in vivo. Notably, we unveiled that hyperglycemia triggers aberrant SOCS-1 expression that correlates with decreased overall IFN signatures in the infected skin. SOCS-1 inhibition restores skin host defense in the highly susceptible hyperglycemic mice. Overall, these data demonstrate a role for SOCS-1-mediated type I interferon actions in host defense and inflammation during MRSA skin infection.


Assuntos
Interferon Tipo I/imunologia , Staphylococcus aureus Resistente à Meticilina/imunologia , Infecções Cutâneas Estafilocócicas/imunologia , Proteína 1 Supressora da Sinalização de Citocina/imunologia , Animais , Interferon Tipo I/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Pele/imunologia , Pele/microbiologia , Infecções Cutâneas Estafilocócicas/microbiologia , Proteína 1 Supressora da Sinalização de Citocina/metabolismo
20.
Nat Immunol ; 22(4): 434-448, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33649580

RESUMO

T cells dynamically interact with multiple, distinct cellular subsets to determine effector and memory differentiation. Here, we developed a platform to quantify cell location in three dimensions to determine the spatial requirements that direct T cell fate. After viral infection, we demonstrated that CD8+ effector T cell differentiation is associated with positioning at the lymph node periphery. This was instructed by CXCR3 signaling since, in its absence, T cells are confined to the lymph node center and alternatively differentiate into stem-like memory cell precursors. By mapping the cellular sources of CXCR3 ligands, we demonstrated that CXCL9 and CXCL10 are expressed by spatially distinct dendritic and stromal cell subsets. Unlike effector cells, retention of stem-like memory precursors in the paracortex is associated with CCR7 expression. Finally, we demonstrated that T cell location can be tuned, through deficiency in CXCL10 or type I interferon signaling, to promote effector or stem-like memory fates.


Assuntos
Infecções por Arenaviridae/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Diferenciação Celular , Quimiocina CXCL10/metabolismo , Quimiocina CXCL9/metabolismo , Memória Imunológica , Linfonodos/metabolismo , Células Precursoras de Linfócitos T/metabolismo , Receptores CXCR3/metabolismo , Animais , Infecções por Arenaviridae/genética , Infecções por Arenaviridae/imunologia , Infecções por Arenaviridae/virologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/virologia , Linhagem da Célula , Células Cultivadas , Quimiocina CXCL10/genética , Quimiocina CXCL9/genética , Quimiotaxia de Leucócito , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Interações Hospedeiro-Patógeno , Interferon Tipo I/metabolismo , Ligantes , Linfonodos/imunologia , Linfonodos/virologia , Vírus da Coriomeningite Linfocítica/imunologia , Vírus da Coriomeningite Linfocítica/patogenicidade , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Células Precursoras de Linfócitos T/imunologia , Células Precursoras de Linfócitos T/virologia , Receptor de Interferon alfa e beta/genética , Receptor de Interferon alfa e beta/metabolismo , Receptores CCR7/metabolismo , Receptores CXCR3/genética , Transdução de Sinais , Nicho de Células-Tronco , Células Estromais/imunologia , Células Estromais/metabolismo
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