RESUMO
COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a global pandemic and has claimed over 2 million lives worldwide. Although the genetic sequences of SARS-CoV and SARS-CoV-2 have high homology, the clinical and pathological characteristics of COVID-19 differ significantly from those of SARS. How and whether SARS-CoV-2 evades (cellular) immune surveillance requires further elucidation. In this study, we show that SARS-CoV-2 infection leads to major histocompability complex class Ι (MHC-Ι) down-regulation both in vitro and in vivo. The viral protein encoded by open reading frame 8 (ORF8) of SARS-CoV-2, which shares the least homology with SARS-CoV among all viral proteins, directly interacts with MHC-Ι molecules and mediates their down-regulation. In ORF8-expressing cells, MHC-Ι molecules are selectively targeted for lysosomal degradation via autophagy. Thus, SARS-CoV-2-infected cells are much less sensitive to lysis by cytotoxic T lymphocytes. Because ORF8 protein impairs the antigen presentation system, inhibition of ORF8 could be a strategy to improve immune surveillance.
Assuntos
Apresentação de Antígeno , COVID-19/imunologia , Regulação para Baixo/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Evasão da Resposta Imune , SARS-CoV-2/imunologia , Proteínas Virais/imunologia , Animais , Autofagia/genética , Autofagia/imunologia , COVID-19/genética , Chlorocebus aethiops , Células HEK293 , Antígenos de Histocompatibilidade Classe I/genética , Humanos , Lisossomos/genética , Lisossomos/imunologia , Lisossomos/virologia , Camundongos , Camundongos Transgênicos , SARS-CoV-2/genética , Células Vero , Proteínas Virais/genéticaRESUMO
Influenza A virus (IAV) infection is a complicated process. After IAVs spread to the lung, extensive pro-inflammatory cytokines and chemokines are released, which largely determine the outcome of infection. Using a single-cell RNA sequencing (scRNA-seq) assay, we systematically and sequentially analyzed the transcriptome of more than 16,000 immune cells in the pulmonary tissue of infected mice, and demonstrated that two waves of pro-inflammatory factors were released. A group of IAV-infected PD-L1+ neutrophils were the major contributor to the first wave at an earlier stage (day 1-3 post infection). Notably, at a later stage (day 7 post infection) when IAV was hardly detected in the immune cells, a group of platelet factor 4-positive (Pf4+)-macrophages generated another wave of pro-inflammatory factors, which were probably the precursors of alveolar macrophages (AMs). Furthermore, single-cell signaling map identified inter-lineage crosstalk between different clusters and helped better understand the signature of PD-L1+ neutrophils and Pf4+-macrophages. Our data characteristically clarified the infiltrated immune cells and their production of pro-inflammatory factors during the immunopathogenesis development, and deciphered the important mechanisms underlying IAV-driven inflammatory reactions in the lung.
Assuntos
Vírus da Influenza A/imunologia , Pulmão/imunologia , Macrófagos Alveolares/imunologia , Animais , Plaquetas/imunologia , Quimiocinas/imunologia , Citocinas/imunologia , Feminino , Humanos , Inflamação/patologia , Vírus da Influenza A Subtipo H1N1/imunologia , Vírus da Influenza A/metabolismo , Vírus da Influenza A/patogenicidade , Influenza Humana/virologia , Macrófagos/imunologia , Macrófagos Alveolares/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/imunologia , Infecções por Orthomyxoviridae/imunologia , Análise de Sequência de RNA/métodos , Transdução de Sinais/imunologia , Análise de Célula Única/métodosRESUMO
Although antiviral drugs are available for the treatment of influenza infection, it is an urgent requirement to develop new antiviral drugs regarding the emergence of drug-resistant viruses. The nucleoprotein (NP) is conserved among all influenza A viruses (IAVs) and has no cellular equivalent. Therefore, NP is an ideal target for the development of new IAV inhibitors. In this study, we identified a novel anti-influenza compound, ZBMD-1, from a library of 20,000 compounds using cell-based influenza A infection assays. We found that ZBMD-1 inhibited the replication of H1N1 and H3N2 influenza A virus strains in vitro, with an IC50 ranging from 0.41-1.14 µM. Furthermore, ZBMD-1 inhibited the polymerase activity and specifically impaired the nuclear export of NP. Further investigation indicated that ZBMD-1 binds to the nuclear export signal 3 (NES3) domain and the dimer interface of the NP pocket. ZBMD-1 also protected mice that were challenged with lethal doses of A/PR/8/1934 (H1N1) virus, effectively relieving lung histopathology changes, as well as strongly inhibiting the expression of pro-inflammatory cytokines/chemokines, without inducing toxicity effects in mice. These results suggest that ZBMD-1 is a promising anti-influenza compound which can be further investigated as a useful strategy against IAVs in the future.
Assuntos
Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Antivirais/farmacologia , Vírus da Influenza A/efeitos dos fármacos , Proteínas de Ligação a RNA/antagonistas & inibidores , Proteínas do Core Viral/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Animais , Humanos , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus da Influenza A Subtipo H1N1/metabolismo , Vírus da Influenza A Subtipo H3N2/efeitos dos fármacos , Vírus da Influenza A Subtipo H3N2/metabolismo , Vírus da Influenza A/metabolismo , Concentração Inibidora 50 , Masculino , Camundongos Endogâmicos BALB C , Proteínas do Nucleocapsídeo , Infecções por Orthomyxoviridae/prevenção & controle , Infecções por Orthomyxoviridae/virologia , Proteínas de Ligação a RNA/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Proteínas do Core Viral/metabolismoRESUMO
Upon infection, naive CD8+ T cells differentiate into cytotoxic effector cells to eliminate the pathogen-infected cells. Although many mechanisms underlying this process have been demonstrated, the regulatory role of chromatin remodeling system in this process remains largely unknown. Here we show that BRD7, a component of the polybromo-associated BAF complex (PBAF), was required for naive CD8+ T cells to differentiate into functional short-lived effector cells (SLECs) in response to acute infections caused by influenza virus or lymphocytic choriomeningitis virus (LCMV). BRD7 deficiency in CD8+ T cells resulted in profound defects in effector population and functions, thereby impairing viral clearance and host recovery. Further mechanical studies indicate that the expression of BRD7 significantly turned to high from naive CD8+ T cells to effector cells, which bridged BRG1 and PBRM1 to the core module of PBAF complex, consequently facilitating the assembly of PBAF complex rather than BAF complex in the effector cells. The PBAF complex changed the chromatin accessibility at the loci of Tbx21 gene and upregulated its expression, leading to the maturation of effector T cells. Our research demonstrates that BRD7 and the PBAF complex are key in CD8+ T cell development and present a significant target for advancing immune therapies.
Assuntos
Linfócitos T CD8-Positivos , Diferenciação Celular , Proteínas Cromossômicas não Histona , Vírus da Coriomeningite Linfocítica , Animais , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Camundongos , Diferenciação Celular/imunologia , Diferenciação Celular/genética , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Cromossômicas não Histona/genética , Vírus da Coriomeningite Linfocítica/imunologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Camundongos Endogâmicos C57BL , Camundongos Knockout , Montagem e Desmontagem da Cromatina , Proteínas que Contêm BromodomínioRESUMO
BACKGROUND: T-cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT) is an immune checkpoint molecule that suppresses CD8+ T-cell function in cancer. However, the expression profile and functional significance of TIGIT in the immune microenvironment of lung adenocarcinoma (LUAD) remain elusive. Interleukin (IL)-15 has emerged as a promising candidate for enhancing CD8+ T-cell mediated tumour eradication. Exploring therapeutic strategies that combine IL-15 with TIGIT blockade in LUAD is warranted. METHODS: We investigated the regulatory network involving coinhibitory TIGIT and CD96, as well as costimulatory CD226 in LUAD using clinical samples. The potential role of TIGIT in regulating the pathogenesis of LUAD was addressed through a murine model with transplanted tumours constructed in Tigit-/- mice. The therapeutic strategy that combines TIGIT blockade with IL-15 stimulation was verified using a transplanted tumour murine model and a patient-derived organoid (PDO) model. RESULTS: The frequency of TIGIT+ CD8+ T cells was significantly increased in LUAD. Increased TIGIT expression indicated poorer prognosis in LUAD patients. Furthermore, the effector function of TIGIT+ CD8+ tumour-infiltrating lymphocytes (TILs) was impaired in LUAD patients and TIGIT inhibited antitumour immune response of CD8+ TILs in tumour-bearing mice. Mechanistically, IL-15 enhanced the effector function of CD8+ TILs but stimulated the expression of TIGIT on CD8+ TILs concomitantly. The application of IL-15 combined with TIGIT blockade showed additive effects in enhancing the cytotoxicity of CD8+ TILs and thus further increased the antitumour immune response in LUAD. CONCLUSIONS: Our findings identified TIGIT as a promising therapeutic target for LUAD. LUAD could benefit more from the combined therapy of IL-15 stimulation and TIGIT blockade.
Assuntos
Adenocarcinoma de Pulmão , Neoplasias Pulmonares , Animais , Humanos , Camundongos , Adenocarcinoma de Pulmão/terapia , Adenocarcinoma de Pulmão/metabolismo , Linfócitos T CD8-Positivos , Modelos Animais de Doenças , Imunoterapia , Interleucina-15/farmacologia , Neoplasias Pulmonares/terapia , Neoplasias Pulmonares/metabolismo , Receptores Imunológicos/genética , Receptores Imunológicos/metabolismo , Microambiente TumoralRESUMO
Long-lived plasma cells (LLPCs) are robust specialized antibody-secreting cells that mainly stay in the bone marrow and can persist a lifetime. As they can be generated by inducing the differentiation of B-lymphocytes, we investigated the possibility that human LLPCs might be engineered to express α-PD-1 monoclonal antibody to substitute recombinant α-PD-1 antitumor immunotherapy. To this end, we inserted an α-PD-1 cassette into the GAPDH locus through Cas9/sgRNA-guided specific integration in B-lymphocytes, which was mediated by an integrase-defective lentiviral vector. The edited B cells were capable of differentiating into LLPCs both in vitro and in vivo. Transcriptional profiling analysis confirmed that these cells were typical LLPCs. Importantly, these cells secreted de novo antibodies persistently, which were able to inhibit human melanoma growth via an antibody-mediated checkpoint blockade in xenograft-tumor mice. Our work suggests that the engineered LLPCs may be utilized as a vehicle to constantly produce special antibodies for long-term cellular immunotherapy to eradicate tumors and cellular reservoirs for various pathogens including human immunodeficiency virus type 1 (HIV-1) and hepatitis B virus (HBV).
Assuntos
Anticorpos Monoclonais/imunologia , Sistemas CRISPR-Cas , Imunoterapia/métodos , Plasmócitos/imunologia , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/imunologia , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/genética , Linfócitos B/citologia , Linfócitos B/imunologia , Diferenciação Celular/imunologia , Linhagem Celular Tumoral , Células HEK293 , Humanos , Camundongos , Plasmócitos/citologia , Linfócitos T/imunologia , TransgenesRESUMO
The polymerase complex of Ebola virus (EBOV) is the functional unit for transcription and replication of viral genome. Nucleoprotein (NP) is a multifunctional protein with high RNA binding affinity and recruits other viral proteins to form functional polymerase complex. In our study, we investigated host proteins associated with EBOV polymerase complex using NP as bait in a transcription and replication competent minigenome system by mass spectrometry analysis and identified SET and MYND domain-containing protein 3 (SMYD3) as a novel host protein which was required for the replication of EBOV. SMYD3 specifically interacted with NP and was recruited to EBOV inclusion bodies through NP. The depletion of SMYD3 dramatically suppressed EBOV mRNA production. A mimic of non-phosphorylated VP30, which is a transcription activator, could partially rescue the viral mRNA production downregulated by the depletion of SMYD3. In addition, SMYD3 promoted NP-VP30 interaction in a dose-dependent manner. These results revealed that SMYD3 was a novel host factor recruited by NP to supporting EBOV mRNA transcription through increasing the binding of VP30 to NP. Thus, our study provided a new understanding of mechanism underlying the transcription of EBOV genome, and a novel anti-EBOV drug design strategy by targeting SMYD3.
Assuntos
Ebolavirus/fisiologia , Histona-Lisina N-Metiltransferase/metabolismo , Interações Hospedeiro-Patógeno , Proteínas do Nucleocapsídeo/metabolismo , RNA Mensageiro/biossíntese , RNA Viral/biossíntese , Transcrição Gênica , Células HEK293 , Humanos , Espectrometria de Massas , Ligação Proteica , Mapeamento de Interação de ProteínasRESUMO
Objectives. Published data on resistin levels in patients with colorectal cancer (CRC) were conflicting and heterogeneous. We conducted a meta-analysis of observational studies to examine the association of circulating resistin levels with carcinogenesis of the CRC. Methods. Potentially eligible studies published up to November 2015 were searched through MEDLINE, EMBASE, Science Citation Index Expanded database, CNKI, and WanFang database. The pooled weighted mean differences (WMDs) with 95% confidence intervals (CIs) calculated by fixed- or random-effect model were used to estimate the effects. Results. A total of 11 studies involving 965 patients were admitted in our meta-analysis. The pooled effects indicated that resistin levels were higher in CRC patients compared to healthy controls (WMD: 1.47 ng/mL; 95% CI: 0.78 to 2.16), with significant heterogeneity across the studies (I (2) = 72%, p < 0.0001). Subgroup analyses and sensitivity analyses revealed that study quality, design, sample type, and resistin assays may account for this heterogeneity. No publication bias was observed. Conclusions. Our meta-analysis suggests that increased circulating resistin levels are associated with greater risk of colorectal cancer. Given the limited number of available studies and significant heterogeneity, larger well-designed randomized studies are warranted.