RESUMO
Interleukin-6 (IL-6) plays a crucial role in various cellular functions, including innate and adaptive immune responses. Dysregulated expression of IL-6 is associated with hyperinflammation and chronic inflammatory diseases. In this study, we aimed to identify the enhancer regions responsible for robust Il6 mRNA expression in murine macrophages. Through comprehensive genome-wide ChIP- and ATAC-seq analyses, we identified two distinct clusters, termed E1 and E2 regions, located at -144 to -163 kb relative to the Il6 transcription start site in lipopolysaccharide (LPS)-activated murine macrophages. These clusters exhibited an accumulation of histone modification marks (H3K27ac and H3K4me1), as well as open chromatin, and were found to contain binding sites for the transcription factors PU.1, NF-κB, C/EBPß, and JunB. Upregulation of non-coding RNA (ncRNA) transcripts from the E1 and E2 regions was observed upon LPS stimulation, and repression of these ncRNAs resulted in abrogation of Il6 expression. Additionally, deletion of either E1 or E2 region significantly impaired Il6 expression, while CRISPR/dCas9 activation-mediated recruitment of the co-activator p300 to the E1 and E2 regions facilitated Il6 expression. Collectively, our findings suggest that the E1 and E2 regions serve as putative enhancers for Il6 expression.
Assuntos
Elementos Facilitadores Genéticos , Interleucina-6 , Lipopolissacarídeos , Macrófagos , Animais , Camundongos , Interleucina-6/metabolismo , Interleucina-6/genética , Interleucina-6/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Elementos Facilitadores Genéticos/genética , Lipopolissacarídeos/farmacologia , Transcrição Gênica , Regulação da Expressão Gênica/imunologia , Camundongos Endogâmicos C57BL , Células RAW 264.7RESUMO
Damage-associated molecular patterns (DAMPs) contribute to antitumor immunity during cancer chemotherapy. We previously demonstrated that topotecan (TPT), a topoisomerase I inhibitor, induces DAMP secretion from cancer cells, which activates STING-mediated antitumor immune responses. However, how TPT induces DAMP secretion in cancer cells is yet to be elucidated. Here, we identified RPL15, a 60S ribosomal protein, as a novel TPT target and showed that TPT inhibited preribosomal subunit formation via its binding to RPL15, resulting in the induction of DAMP-mediated antitumor immune activation independent of TOP1. TPT inhibits RPL15-RPL4 interactions and decreases RPL4 stability, which is recovered by CDK12 activity. RPL15 knockdown induced DAMP secretion and increased the CTL population but decreased the regulatory T cell population in a B16-F10 murine melanoma model, which sensitized B16-F10 tumors against PD-1 blockade. Our study identified a novel TPT target protein and showed that ribosomal stress is a trigger of DAMP secretion, which contributes to antitumor immunotherapy.
Assuntos
Neoplasias , Topotecan , Animais , Camundongos , Neoplasias/tratamento farmacológico , Proteínas Ribossômicas , Inibidores da Topoisomerase I/farmacologia , Topotecan/farmacologia , Topotecan/uso terapêuticoRESUMO
Pattern recognition receptors (PRRs) play critical roles in recognizing pathogen-derived nucleic acids and inducing innate immune responses, such as inflammation and type I interferon production. PRRs that recognize nucleic acids include members of endosomal Toll-like receptors, cytosolic retinoic acid inducible gene I-like receptors, cyclic GMP-AMP synthase, absent in melanoma 2-like receptors, and nucleotide binding oligomerization domain-like receptors. Aberrant recognition of self-derived nucleic acids by these PRRs or unexpected activation of downstream signaling pathways results in the constitutive production of type I interferons and inflammatory cytokines, which lead to the development of autoimmune or autoinflammatory diseases. In this review, we focus on the nucleic acid-sensing machinery and its pathophysiological roles in various inflammatory diseases.
Assuntos
Interferon Tipo I , Ácidos Nucleicos , Imunidade Inata , Interferon Tipo I/metabolismo , Ácidos Nucleicos/metabolismo , Receptores de Reconhecimento de Padrão , Receptores Toll-LikeRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has emerged as a pandemic. Paucity of information concerning the virus and therapeutic interventions have made SARS-CoV-2 infection a genuine threat to global public health. Therefore, there is a growing need for understanding the molecular mechanism of SARS-CoV-2 infection at cellular level. To address this, we undertook a systems biology approach by analyzing publicly available RNA-seq datasets of SARS-CoV-2 infection of different cells and compared with other lung pathogenic infections. Our study identified several key genes and pathways uniquely associated with SARS-CoV-2 infection. Genes such as interleukin (IL)-6, CXCL8, CCL20, CXCL1 and CXCL3 were upregulated, which in particular regulate the cytokine storm and IL-17 signaling pathway. Of note, SARS-CoV-2 infection strongly activated IL-17 signaling pathway compared with other respiratory viruses. Additionally, this transcriptomic signature was also analyzed to predict potential drug repurposing and small molecule inhibitors. In conclusion, our comprehensive data analysis identifies key molecular pathways to reveal underlying pathological etiology and potential therapeutic targets in SARS-CoV-2 infection.
Assuntos
COVID-19/imunologia , Interleucina-17/genética , SARS-CoV-2/fisiologia , Biologia de Sistemas/métodos , Antivirais/uso terapêutico , Quimiocina CCL20/genética , Quimiocina CXCL1/genética , Quimiocinas CXC/genética , Reposicionamento de Medicamentos , Humanos , Interleucina-17/metabolismo , Interleucina-6/genética , Interleucina-8/genética , Especificidade de Órgãos , Transdução de Sinais , Transcriptoma , Tratamento Farmacológico da COVID-19RESUMO
The nucleotide-binding oligomerization domain-like receptor (NLR) family pyrin domain containing (NLRP) 3 inflammasome is a multiprotein complex that triggers Caspase-1-mediated IL-1ß production and pyroptosis, and its dysregulation is associated with the pathogenesis of inflammatory diseases. 1'-Acetoxychavicol acetate (ACA) is a natural compound in the rhizome of tropical ginger Alpinia species with anti-microbial, anti-allergic and anti-cancer properties. In this study, we found that ACA suppressed NLRP3 inflammasome activation in mouse bone marrow-derived macrophages and human THP-1 monocytes. ACA inhibited Caspase-1 activation and IL-1ß production by NLRP3 agonists such as nigericin, monosodium urate (MSU) crystals, and ATP. Moreover, it suppressed oligomerization of the adapter molecule, apoptosis-associated speck-like protein containing a CARD (ASC), and Caspase-1-mediated cleavage of pyroptosis executor Gasdermin D. Mechanistically, ACA inhibited generation of mitochondrial reactive oxygen species (ROS) and prevented release of oxidized mitochondrial DNA, which trigger NLRP3 inflammasome activation. ACA also prevented NLRP3 inflammasome activation in vivo, as evidenced in the MSU crystal-induced peritonitis and dextran sodium sulfate-induced colitis mouse models accompanied by decreased Caspase-1 activation. Thus, ACA is a potent inhibitor of the NLRP3 inflammasome for prevention of NLRP3-associated inflammatory diseases.
Assuntos
Álcoois Benzílicos/farmacologia , Inflamassomos/efeitos dos fármacos , Inflamassomos/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Caspase 1/metabolismo , Células Cultivadas , Modelos Animais de Doenças , Humanos , Inflamação/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Peritonite/tratamento farmacológico , Peritonite/metabolismo , Fagocitose/efeitos dos fármacos , Piroptose/efeitos dos fármacos , Células THP-1/efeitos dos fármacos , Células THP-1/metabolismoRESUMO
Interferon regulatory factors (IRFs) are transcription factors found in both vertebrates and invertebrates that were recently identified and found to play an important role in antiviral immunity in black tiger shrimp Penaeus monodon. In this study, we investigated the mechanism by which P. monodon IRF (PmIRF) regulates the immune-related genes downstream of the cytosolic DNA sensing pathway. Depletion of PmIRF by double-stranded RNA-mediated gene silencing significantly reduced the mRNA expression levels of the IFN-like factors PmVago1, PmVago4, and PmVago5 and antilipopolysaccharide factor 6 (ALFPm6) in shrimp. In human embryonic kidney (HEK293T) cells transfected with PmIRF or co-transfected with DEAD-box polypeptide (PmDDX41) and simulator of IFN genes (PmSTING) expression plasmids, the promoter activity of IFN-ß, nuclear factor (NF-κB), and ALFPm6 was synergistically enhanced following stimulation with the nucleic acid mimics deoxyadenylic-deoxythymidylic acid sodium salt [poly(dA:dT)] and high molecular weight (HMW) polyinosinic-polycytidylic acid [poly(I:C)]. Both nucleic acid mimics also significantly induced PmSTING, PmIRF, and ALFPm6 gene expression. Co-immunoprecipitation experiments showed that PmIRF interacted with PmSTING in cells stimulated with poly(dA:dT). PmSTING, PmIRF, and PmDDX41 were localized in the cytoplasm of unstimulated HEK293T cells and PmIRF and PmDDX41 were translocated to the nucleus upon stimulation with the nucleic acid mimics while PmSTING remained in the cytoplasm. These results indicate that PmIRF transduces the pathogen signal via the PmDDX41-PmSTING DNA sensing pathway to induce downstream production of interferon-like molecules and antimicrobial peptides.
Assuntos
Peptídeos Antimicrobianos/genética , DNA/imunologia , Regulação da Expressão Gênica , Fatores Reguladores de Interferon/metabolismo , Interferons/genética , Proteínas de Membrana/metabolismo , Penaeidae/fisiologia , Animais , Peptídeos Antimicrobianos/metabolismo , Linhagem Celular , Células Cultivadas , Inativação Gênica , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata , Fatores Reguladores de Interferon/farmacologia , Interferons/metabolismo , Transdução de SinaisRESUMO
The innate immune system plays an essential role in initial recognition of pathogen infection by producing inflammatory cytokines and type I interferons. cGAS is a cytoplasmic sensor for DNA derived from DNA viruses. cGAS binding with DNA induces the production of cGAMP, a second messenger that associates with STING in endoplasmic reticulum (ER). STING changes its cellular distribution from ER to perinuclear Golgi, where it activates the protein kinase TBK1 that catalyzes the phosphorylation of IRF3. Here we found that STING trafficking is regulated by myotubularin-related protein (MTMR) 3 and MTMR4, members of protein tyrosine phosphatases that dephosphorylate 3' position in phosphatidylinositol (PtdIns) and generate PtdIns5P from PtdIns3,5P2 and PtdIns from PtdIns3P. We established MTMR3 and MTMR4 double knockout (DKO) RAW264.7 macrophage cells and found that they exhibited increased type I interferon production after interferon-stimulatory DNA (ISD) stimulation and herpes simplex virus 1 infection concomitant with enhanced IRF3 phosphorylation. In DKO cells, STING rapidly trafficked from ER to Golgi after ISD stimulation. Notably, DKO cells exhibited enlarged cytosolic puncta positive for PtdIns3P and STING was aberrantly accumulated in this puncta. Taken together, these results suggest that MTMR3 and MTMR4 regulate the production of PtdIns3P, which plays a critical role in suppressing DNA-mediated innate immune responses via modulating STING trafficking.
Assuntos
DNA Viral/imunologia , Herpesvirus Humano 1/imunologia , Imunidade Inata , Macrófagos/imunologia , Proteínas de Membrana/imunologia , Fosfatos de Fosfatidilinositol/imunologia , Proteínas Tirosina Fosfatases não Receptoras/imunologia , Animais , DNA Viral/genética , Herpesvirus Humano 1/genética , Proteínas de Membrana/genética , Camundongos , Fosfatos de Fosfatidilinositol/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Transporte Proteico/genética , Transporte Proteico/imunologia , Proteínas Tirosina Fosfatases não Receptoras/genética , Células RAW 264.7RESUMO
Toll-like receptor 3 (TLR3) recognizes double-stranded RNA derived from virus and its synthetic analogue, polyinosinic-polycytidylic acid [poly(I:C)]. Upon poly(I:C) binding, TLR3 activates transcription factors to express inflammatory cytokines and type I interferon. TLR3 is located in the endosomes and its recognition of poly(I:C) and activation of downstream signaling is regulated by endosomal acidification. However, the mechanism of post-transcriptional regulation in TLR3-mediated innate responses remains unclear. Here, we focused on Human antigen R (HuR, also known as ELAVL1) that recognizes and binds to the 3' untranslated regions (3'UTRs) of target mRNAs, thereby protecting them from mRNA degradation, and found that HuR-deficient murine macrophage cells showed significantly reduced Ifnb1 mRNA expression after poly(I:C) stimulation. HuR-deficient cells also showed a marked reduction in the expression of Atp6v0d2 mRNA, which encodes a subunit of vacuolar-type H+ ATPase (V-ATPase), and therefore reduced endosomal acidification. HuR associated with the 3'UTR of Atp6v0d2 mRNA and the stability of Atp6v0d2 mRNA was maintained by its association with HuR. Taken together, our results suggest that HuR stabilizes Atp6v0d2 mRNA, which is required for the TLR3-mediated innate immune responses.
Assuntos
Proteína Semelhante a ELAV 1/metabolismo , Imunidade Inata/fisiologia , Macrófagos/imunologia , Receptor 3 Toll-Like/metabolismo , ATPases Vacuolares Próton-Translocadoras/metabolismo , Animais , Regulação da Expressão Gênica , Macrófagos/metabolismo , Camundongos , Estabilidade de RNA/fisiologia , RNA de Cadeia DuplaRESUMO
Retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs), RIG-I, and melanoma differentiation-associated gene 5 (MDA5) play a critical role in inducing antiviral innate immune responses by activating IFN regulatory factor 3 (IRF3) and NF-κB, which regulates the transcription of type I IFN and inflammatory cytokines. Antiviral innate immune responses are also regulated by posttranscriptional and translational mechanisms. In this study, we identified an RNA-binding protein HuR as a regulator for RLR signaling. Overexpression of HuR, but not of other Hu members, increased IFN-ß promoter activity. HuR-deficient macrophage cells exhibited decreased Ifnb1 expression after RLR stimulation, whereas they showed normal induction after stimulation with bacterial LPS or immunostimulatory DNA. Moreover, HuR-deficient cells displayed impaired nuclear translocation of IRF3 after RLR stimulation. In HuR-deficient cells, the mRNA expression of Polo-like kinase (PLK) 2 was markedly reduced. We found that HuR bound to the 3' untranslated region of Plk2 mRNA and increased its stabilization. PLK2-deficient cells also showed reduced IRF3 nuclear translocation and Ifnb mRNA expression during RLR signaling. Together, these findings suggest that HuR bolsters RLR-mediated IRF3 nuclear translocation by controlling the stability of Plk2 mRNA.
Assuntos
Antivirais/imunologia , Proteína Semelhante a ELAV 1/imunologia , Imunidade Inata/imunologia , Proteínas Serina-Treonina Quinases/imunologia , RNA Mensageiro/imunologia , Regiões 3' não Traduzidas/imunologia , Animais , Linhagem Celular , Proteína DEAD-box 58/imunologia , DNA/imunologia , Células HEK293 , Humanos , Fator Regulador 3 de Interferon/imunologia , Interferon Tipo I/imunologia , Helicase IFIH1 Induzida por Interferon/imunologia , Interferon beta/imunologia , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/imunologia , Células RAW 264.7 , Transdução de Sinais/imunologiaRESUMO
TLRs recognize pathogen components and drive innate immune responses. They localize at either the plasma membrane or intracellular vesicles such as endosomes and lysosomes, and proper cellular localization is important for their ligand recognition and initiation of signaling. In this study, we disrupted ATP6V0D2, a component of vacuolar-type H+ adenosine triphosphatase (V-ATPase) that plays a central role in acidification of intracellular vesicles, in a macrophage cell line. ATP6V0D2-deficient cells exhibited reduced cytokine production in response to endosome-localized, nucleic acid-sensing TLR3, TLR7, and TLR9, but enhanced inflammatory cytokine production and NF-κB activation following stimulation with LPS, a TLR4 agonist. Moreover, they had defects in internalization of cell surface TLR4 and exhibited enhanced inflammatory cytokine production after repeated LPS stimulation, thereby failing to induce LPS tolerance. A component of the V-ATPase complex interacted with ARF6, the small GTPase known to regulate TLR4 internalization, and ARF6 deficiency resulted in prolonged TLR4 expression on the cell surface. Taken together, these findings suggest that ATP6V0D2-dependent intravesicular acidification is required for TLR4 internalization, which is associated with prevention from excessive LPS-triggered inflammation and induction of tolerance.
Assuntos
Tolerância Imunológica/imunologia , Inflamação/imunologia , Lipopolissacarídeos/imunologia , Macrófagos/imunologia , Receptor 4 Toll-Like/metabolismo , Animais , Vesículas Citoplasmáticas/imunologia , Vesículas Citoplasmáticas/metabolismo , Células HEK293 , Humanos , Inflamação/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Transporte Proteico/imunologia , Células RAW 264.7 , Receptor 4 Toll-Like/imunologiaRESUMO
The innate immune system serves as the first line of defense to protect the host from pathogen infection. As a first step, the pattern recognition receptors (PRRs) recognize pathogen-associated molecular patterns (PAMPs), such as non-self DNA derived from pathogens, and damage-associated molecular patterns (DAMPs), such as self DNA released from damaged or injured cells. Sensing of such DNAs elicits innate immune responses through the production of type I interferons (IFNs) and proinflammatory cytokines resulting from the activation of interferon regulatory factor 3 (IRF3) and nuclear factor kappa B (NF-κB), respectively. These cytokines are key players in interlinking innate and adaptive immune responses. However, defects in DNA sensors and their signaling cascades lead to dysregulation of immune responses, autoimmune diseases, and cancer progression. Here we provide an update on DNA signaling pathways in response to pathogen infection and cell injury, and on the roles of regulators in governing the immune system and maintaining host homeostasis. We also discuss the evasion of immunosurveillance by pathogens.
Assuntos
DNA/imunologia , Imunidade Inata/imunologia , Receptores de Reconhecimento de Padrão/imunologia , Transdução de Sinais/imunologia , Animais , Citocinas/biossíntese , Citocinas/imunologia , Humanos , Interferon Tipo I/biossíntese , Interferon Tipo I/imunologiaRESUMO
Alveolar macrophages (AMs) are specialized tissue-resident macrophages that orchestrate the immune responses to inhaled pathogens and maintain organ homeostasis of the lung. Dysregulation of AMs is associated with allergic inflammation and asthma. Here, we examined the role of a phosphoinositide kinase PIKfyve in AM development and function. Mice with conditionally deleted PIKfyve in macrophages have altered AM populations. PIKfyve deficiency results in a loss of AKT activation in response to GM-CSF, a cytokine critical for AM development. Upon exposure to house dust mite extract, mutant mice display severe lung inflammation and allergic asthma accompanied by infiltration of eosinophils and lymphoid cells. Moreover, they have defects in production of retinoic acid and fail to support incorporation of Foxp3+ Treg cells in the lung, resulting in exacerbation of lung inflammation. Thus, PIKfyve plays a role in preventing excessive lung inflammation through regulating AM function.
Assuntos
Asma/patologia , Hipersensibilidade/patologia , Inflamação/patologia , Macrófagos Alveolares/fisiologia , Fosfatidilinositol 3-Quinases/metabolismo , Alérgenos/administração & dosagem , Animais , Técnicas de Inativação de Genes , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Pulmão/patologia , Camundongos , Camundongos Knockout , Fosfatidilinositol 3-Quinases/deficiência , Deleção de Sequência , Transdução de Sinais , Linfócitos T Reguladores/imunologiaRESUMO
During viral and bacterial infections, pathogen-derived cytosolic nucleic acids are recognized by the intracellular RNA sensors retinoic acid-inducible gene I and melanoma-differentiated gene 5 and intracellular DNA sensors, including cyclic-di-GMP-AMP synthase, absent in melanoma 2, interferon (IFN)-gamma inducible protein 16, polymerase III, and so on. Binding of intracellular nucleic acids to these sensors activates downstream signaling cascades, resulting in the production of type I IFNs and pro-inflammatory cytokines to induce appropriate systematic immune responses. While these sensors also recognize endogenous nucleic acids and activate immune responses, they can discriminate between self- and non-self-nucleic acids. However, dysfunction of these sensors or failure of regulatory mechanisms causes aberrant activation of immune response and autoimmune disorders. In this review, we focus on how intracellular immune sensors recognize exogenous nucleic acids and activate the innate immune system, and furthermore, how autoimmune diseases result from dysfunction of these sensors.
Assuntos
Adenilil Ciclases/metabolismo , Doenças Autoimunes/imunologia , Proteína DEAD-box 58/metabolismo , Proteínas de Ligação a DNA/metabolismo , Helicase IFIH1 Induzida por Interferon/metabolismo , Ácidos Nucleicos/imunologia , Animais , Citosol/metabolismo , Interações Hospedeiro-Parasita , Humanos , Imunidade Inata , Receptores Imunológicos , Receptores de Reconhecimento de Padrão/metabolismo , Tolerância a Antígenos Próprios , Transdução de SinaisRESUMO
The innate immune system senses RNA viruses by pattern recognition receptors (PRRs) and protects the host from virus infection. PRRs mediate the production of immune modulatory factors and direct the elimination of RNA viruses. Here, we show a unique PRR that mediates antiviral response. Tetrachlorodibenzo-p-dioxin (TCDD)-inducible poly(ADP ribose) polymerase (TIPARP), a Cysteine3 Histidine (CCCH)-type zinc finger-containing protein, binds to Sindbis virus (SINV) RNA via its zinc finger domain and recruits an exosome to induce viral RNA degradation. TIPARP typically localizes in the nucleus, but it accumulates in the cytoplasm after SINV infection, allowing targeting of cytoplasmic SINV RNA. Redistribution of TIPARP is induced by reactive oxygen species (ROS)-dependent oxidization of the nuclear pore that affects cytoplasmic-nuclear transport. BCL2-associated X protein (BAX) and BCL2 antagonist/killer 1 (BAK1), B-cell leukemia/lymphoma 2 (BCL2) family members, mediate mitochondrial damage to generate ROS after SINV infection. Thus, TIPARP is a viral RNA-sensing PRR that mediates antiviral responses triggered by BAX- and BAK1-dependent mitochondrial damage.
Assuntos
Imunidade Inata/genética , Poli(ADP-Ribose) Polimerases/genética , Vírus de RNA/genética , Receptores de Reconhecimento de Padrão/genética , Transporte Ativo do Núcleo Celular/genética , Transporte Ativo do Núcleo Celular/imunologia , Citoplasma/genética , Citoplasma/imunologia , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Mitocôndrias/genética , Mitocôndrias/patologia , Mitocôndrias/virologia , Proteínas de Transporte de Nucleosídeos , Poli(ADP-Ribose) Polimerases/imunologia , Vírus de RNA/imunologia , Espécies Reativas de Oxigênio/metabolismo , Receptores de Reconhecimento de Padrão/imunologia , Sindbis virus/genética , Sindbis virus/imunologia , Sindbis virus/patogenicidade , Proteína Killer-Antagonista Homóloga a bcl-2/genética , Proteína Killer-Antagonista Homóloga a bcl-2/imunologia , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/imunologiaRESUMO
Danger-associated molecular patterns derived from damaged or dying cells elicit inflammation and potentiate antitumor immune responses. In this article, we show that treatment of breast cancer cells with the antitumor agent topotecan (TPT), an inhibitor of topoisomerase I, induces danger-associated molecular pattern secretion that triggers dendritic cell (DC) activation and cytokine production. TPT administration inhibits tumor growth in tumor-bearing mice, which is accompanied by infiltration of activated DCs and CD8+ T cells. These effects are abrogated in mice lacking STING, an essential molecule in cytosolic DNA-mediated innate immune responses. Furthermore, TPT-treated cancer cells release exosomes that contain DNA that activate DCs via STING signaling. These findings suggest that a STING-dependent pathway drives antitumor immunity by responding to tumor cell-derived DNA.
Assuntos
DNA de Neoplasias/imunologia , Exossomos/efeitos dos fármacos , Exossomos/genética , Proteínas de Membrana/metabolismo , Neoplasias/tratamento farmacológico , Inibidores da Topoisomerase I/farmacologia , Topotecan/administração & dosagem , Animais , Antineoplásicos/farmacologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , DNA de Neoplasias/isolamento & purificação , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/fisiologia , Feminino , Imunidade Inata , Ativação Linfocitária , Proteínas de Membrana/deficiência , Proteínas de Membrana/imunologia , Camundongos , Neoplasias/imunologia , Transdução de Sinais/efeitos dos fármacosRESUMO
Pathological conditions such as epilepsy cause misregulation of adult neural stem/progenitor populations in the adult hippocampus in mice, and the resulting abnormal neurogenesis leads to impairment in learning and memory. However, how animals cope with abnormal neurogenesis remains unknown. Here we show that microglia in the mouse hippocampus attenuate convulsive seizure-mediated aberrant neurogenesis through the activation of Toll-like receptor 9 (TLR9), an innate immune sensor known to recognize microbial DNA and trigger inflammatory responses. We found that microglia sense self-DNA from degenerating neurons following seizure, and secrete tumour necrosis factor-α, resulting in attenuation of aberrant neurogenesis. Furthermore, TLR9 deficiency exacerbated seizure-induced cognitive decline and recurrent seizure severity. Our findings thus suggest the existence of bidirectional communication between the innate immune and nervous systems for the maintenance of adult brain integrity.
Assuntos
Disfunção Cognitiva/imunologia , Hipocampo/imunologia , Microglia/imunologia , Neurogênese/imunologia , Convulsões/imunologia , Receptor Toll-Like 9/genética , Animais , Cognição , Disfunção Cognitiva/genética , Disfunção Cognitiva/patologia , Regulação da Expressão Gênica , Hipocampo/crescimento & desenvolvimento , Hipocampo/metabolismo , Hipocampo/patologia , Imunidade Inata/genética , Masculino , Glicoproteínas de Membrana/deficiência , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microglia/metabolismo , Microglia/patologia , Neurogênese/genética , Neurônios/imunologia , Neurônios/metabolismo , Neurônios/patologia , Convulsões/genética , Convulsões/metabolismo , Convulsões/patologia , Índice de Gravidade de Doença , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Receptor 7 Toll-Like/deficiência , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/imunologia , Receptor Toll-Like 9/deficiência , Receptor Toll-Like 9/imunologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
RIG-I-like receptors (RLRs), including retinoic acid-inducible gene-I (RIG-I) and MDA5, constitute a family of cytoplasmic RNA helicases that senses viral RNA and mounts antiviral innate immunity by producing type I interferons and inflammatory cytokines. Despite their essential roles in antiviral host defense, RLR signaling is negatively regulated to protect the host from excessive inflammation and autoimmunity. Here, we identified ADP-ribosylation factor-like protein 5B (Arl5B), an Arl family small GTPase, as a regulator of RLR signaling through MDA5 but not RIG-I. Overexpression of Arl5B repressed interferon ß promoter activation by MDA5 but not RIG-I, and its knockdown enhanced MDA5-mediated responses. Furthermore, Arl5B-deficient mouse embryonic fibroblast cells exhibited increased type I interferon expression in response to MDA5 agonists such as poly(I:C) and encephalomyocarditis virus. Arl5B-mediated negative regulation of MDA5 signaling does not require its GTP binding ability but requires Arl5B binding to the C-terminal domain of MDA5, which prevents interaction between MDA5 and poly(I:C). Our results, therefore, suggest that Arl5B is a negative regulator for MDA5.
Assuntos
Fatores de Ribosilação do ADP/metabolismo , RNA Helicases DEAD-box/metabolismo , Imunidade Inata/genética , Interferon Tipo I/biossíntese , Fatores de Ribosilação do ADP/genética , Fatores de Ribosilação do ADP/imunologia , Animais , Autoimunidade/genética , Proteína DEAD-box 58 , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/imunologia , Humanos , Helicase IFIH1 Induzida por Interferon , Interferon beta/genética , Camundongos , Regiões Promotoras Genéticas/genética , RNA Helicases/imunologia , RNA Viral/imunologia , Receptores Imunológicos , Transdução de SinaisRESUMO
The RIG-I-like receptors, retinoic acid inducible gene-1 (RIG-I), melanoma differentiation-associated protein 5, and laboratory of genetics and physiology-2, are cytoplasmic sensors for RNA viruses that mediate the antiviral innate immune responses. We demonstrate that really interesting new gene-finger domain- and K homology domain-containing MEX3C regulates RIG-I function. MEX3C colocalizes with RIG-I in the stress granules of virally infected cells, and its overexpression causes the lysine-63-linked ubiquitination of RIG-I and activates IFN-ß promoter. Embryonic fibroblast cells, macrophages, and conventional dendritic cells derived from Mex3c-deficient mice showed defective production of type I IFN after infection with RNA viruses that are recognized by RIG-I. These results demonstrate that MEX3C is an E3 ubiquitin ligase that modifies RIG-I in stress granules and plays a critical role in eliciting antiviral immune responses.
Assuntos
Imunidade Inata/imunologia , Proteínas de Ligação a RNA/imunologia , Receptores do Ácido Retinoico/imunologia , Viroses/imunologia , Animais , Citocinas/biossíntese , Primers do DNA/genética , Ensaio de Imunoadsorção Enzimática , Células HEK293 , Humanos , Immunoblotting , Imunoprecipitação , Luciferases , Camundongos , Camundongos Knockout , Mutagênese Sítio-Dirigida , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Receptores do Ácido Retinoico/metabolismo , UbiquitinaçãoRESUMO
The cytoplasmic helicase RIG-I is an established sensor for viral 5'-triphosphorylated RNA species. Recently, RIG-I was also implicated in the detection of intracellular bacteria. However, little is known about the host cell specificity of this process and the bacterial pathogen-associated molecular pattern (PAMP) that activates RIG-I. Here we show that RNA of Salmonella enterica serovar Typhimurium activates production of beta interferon in a RIG-I-dependent fashion only in nonphagocytic cells. In phagocytic cells, RIG-I is obsolete for detection of Salmonella infection. We further demonstrate that Salmonella mRNA reaches the cytoplasm during infection and is thus accessible for RIG-I. The results from next-generation sequencing analysis of RIG-I-associated RNA suggest that coding bacterial mRNAs represent the activating PAMP. IMPORTANCE S. Typhimurium is a major food-borne pathogen. After fecal-oral transmission, it can infect epithelial cells in the gut as well as immune cells (mainly macrophages, dendritic cells, and M cells). The innate host immune system relies on a growing number of sensors that detect pathogen-associated molecular patterns (PAMPs) to launch a first broad-spectrum response to invading pathogens. Successful detection of a given pathogen depends on colocalization of host sensors and PAMPs as well as potential countermeasures of the pathogen during infection. RIG-I-like helicases were mainly associated with detection of RNA viruses. Our work shows that S. Typhimurium is detected by RIG-I during infection specifically in nonimmune cells.
Assuntos
RNA Helicases DEAD-box/imunologia , Interações Hospedeiro-Patógeno , RNA Bacteriano/imunologia , RNA Mensageiro/imunologia , Receptores Imunológicos/imunologia , Salmonella typhimurium/imunologia , Animais , Linhagem Celular , Proteína DEAD-box 58 , RNA Helicases DEAD-box/metabolismo , Humanos , Interferon beta/imunologia , Interferon beta/metabolismo , Camundongos , Ligação Proteica , RNA Bacteriano/metabolismo , RNA Mensageiro/metabolismo , Receptores Imunológicos/metabolismoRESUMO
High-dose ionizing radiation induces severe DNA damage in the epithelial stem cells in small intestinal crypts and causes gastrointestinal syndrome (GIS). Although the tumour suppressor p53 is a primary factor inducing death of crypt cells with DNA damage, its essential role in maintaining genome stability means inhibiting p53 to prevent GIS is not a viable strategy. Here we show that the innate immune receptor Toll-like receptor 3 (TLR3) is critical for the pathogenesis of GIS. Tlr3(-/-) mice show substantial resistance to GIS owing to significantly reduced radiation-induced crypt cell death. Despite showing reduced crypt cell death, p53-dependent crypt cell death is not impaired in Tlr3(-/-) mice. p53-dependent crypt cell death causes leakage of cellular RNA, which induces extensive cell death via TLR3. An inhibitor of TLR3-RNA binding ameliorates GIS by reducing crypt cell death. Thus, we propose blocking TLR3 activation as a novel approach to treat GIS.