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1.
Nat Immunol ; 21(12): 1611, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32681143

RESUMO

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

2.
Cell ; 165(3): 551-65, 2016 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-27040498

RESUMO

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by loss of tolerance to nucleic acids and highly diverse clinical manifestations. To assess its molecular heterogeneity, we longitudinally profiled the blood transcriptome of 158 pediatric patients. Using mixed models accounting for repeated measurements, demographics, treatment, disease activity (DA), and nephritis class, we confirmed a prevalent IFN signature and identified a plasmablast signature as the most robust biomarker of DA. We detected gradual enrichment of neutrophil transcripts during progression to active nephritis and distinct signatures in response to treatment in different nephritis subclasses. Importantly, personalized immunomonitoring uncovered individual correlates of disease activity that enabled patient stratification into seven groups, supported by patient genotypes. Our study uncovers the molecular heterogeneity of SLE and provides an explanation for the failure of clinical trials. This approach may improve trial design and implementation of tailored therapies in genetically and clinically complex autoimmune diseases. PAPERCLIP.


Assuntos
Lúpus Eritematoso Sistêmico/genética , Adolescente , Criança , Feminino , Humanos , Estudos Longitudinais , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/patologia , Lúpus Eritematoso Sistêmico/terapia , Nefrite Lúpica/genética , Nefrite Lúpica/imunologia , Neutrófilos/imunologia , Polimorfismo de Nucleotídeo Único , Medicina de Precisão , Transcriptoma
3.
Nat Immunol ; 17(6): 646-55, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27111142

RESUMO

Group 2 innate lymphoid cells (ILC2 cells) are important for type 2 immune responses and are activated by the epithelial cytokines interleukin 33 (IL-33), IL-25 and thymic stromal lymphopoietin (TSLP). Here we demonstrated that IL-1ß was a critical activator of ILC2 cells, inducing proliferation and cytokine production and regulating the expression of epithelial cytokine receptors. IL-1ß also governed ILC2 plasticity by inducing low expression of the transcription factor T-bet and the cytokine receptor chain IL-12Rß2, which enabled the conversion of these cells into an ILC1 phenotype in response to IL-12. This transition was marked by an atypical chromatin landscape characterized by the simultaneous transcriptional accessibility of the locus encoding interferon-γ (IFN-γ) and the loci encoding IL-5 and IL-13. Finally, IL-1ß potentiated ILC2 activation and plasticity in vivo, and IL-12 acted as the switch that determined an ILC2-versus-ILC1 response. Thus, we have identified a previously unknown role for IL-1ß in facilitating ILC2 maturation and plasticity.


Assuntos
Plasticidade Celular , Imunidade Inata , Interleucina-12/metabolismo , Interleucina-1beta/metabolismo , Linfócitos/imunologia , Animais , Diferenciação Celular , Plasticidade Celular/imunologia , Células Cultivadas , Citocinas/metabolismo , Humanos , Interferon gama/metabolismo , Interleucina-17/metabolismo , Interleucina-33/metabolismo , Camundongos , Camundongos SCID , Receptores de Interleucina-12/genética , Receptores de Interleucina-12/metabolismo , Transdução de Sinais , Proteínas com Domínio T/genética , Proteínas com Domínio T/metabolismo , Células Th1/imunologia , Equilíbrio Th1-Th2 , Células Th2/imunologia , Linfopoietina do Estroma do Timo
4.
Nat Immunol ; 17(12): 1373-1380, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27695001

RESUMO

The respiratory tract is heavily populated with innate immune cells, but the mechanisms that control such cells are poorly defined. Here we found that the E3 ubiquitin ligase TRIM29 was a selective regulator of the activation of alveolar macrophages, the expression of type I interferons and the production of proinflammatory cytokines in the lungs. We found that deletion of TRIM29 enhanced macrophage production of type I interferons and protected mice from infection with influenza virus, while challenge of Trim29-/- mice with Haemophilus influenzae resulted in lethal lung inflammation due to massive production of proinflammatory cytokines by macrophages. Mechanistically, we demonstrated that TRIM29 inhibited interferon-regulatory factors and signaling via the transcription factor NF-κB by degrading the adaptor NEMO and that TRIM29 directly bound NEMO and subsequently induced its ubiquitination and proteolytic degradation. These data identify TRIM29 as a key negative regulator of alveolar macrophages and might have important clinical implications for local immunity and immunopathology.


Assuntos
Infecções por Haemophilus/imunologia , Haemophilus influenzae/imunologia , Vírus da Influenza A/imunologia , Macrófagos/imunologia , Infecções por Orthomyxoviridae/imunologia , Sistema Respiratório/imunologia , Fatores de Transcrição/metabolismo , Animais , Células Cultivadas , Imunidade Inata , Interferon Tipo I/genética , Interferon Tipo I/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Macrófagos/microbiologia , Macrófagos/virologia , Camundongos , Camundongos Knockout , NF-kappa B/metabolismo , Proteólise , Transdução de Sinais , Fatores de Transcrição/genética , Ubiquitinação
6.
Nat Immunol ; 16(9): 970-9, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26168081

RESUMO

Interleukin 17-producing helper T cells (T(H)17 cells) have a major role in protection against infections and in mediating autoimmune diseases, yet the mechanisms involved are incompletely understood. We found that interleukin 26 (IL-26), a human T(H)17 cell-derived cytokine, is a cationic amphipathic protein that kills extracellular bacteria via membrane-pore formation. Furthermore, T(H)17 cell-derived IL-26 formed complexes with bacterial DNA and self-DNA released by dying bacteria and host cells. The resulting IL-26-DNA complexes triggered the production of type I interferon by plasmacytoid dendritic cells via activation of Toll-like receptor 9, but independently of the IL-26 receptor. These findings provide insights into the potent antimicrobial and proinflammatory function of T(H)17 cells by showing that IL-26 is a natural human antimicrobial that promotes immune sensing of bacterial and host cell death.


Assuntos
DNA Bacteriano/imunologia , DNA/imunologia , Imunidade Inata/imunologia , Interleucinas/imunologia , Células Th17/imunologia , Receptor Toll-Like 9/imunologia , Animais , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Humanos , Interferon Tipo I/imunologia , Interferon Tipo I/metabolismo , Camundongos , Psoríase/imunologia , Receptores de Interleucina/imunologia , Receptores de Interleucina/metabolismo
7.
PLoS Pathog ; 20(7): e1012379, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-39037956

RESUMO

RNA helicases are involved in the innate immune response against pathogens, including bacteria and viruses; however, their mechanism in the human airway epithelial cells is still not fully understood. Here, we demonstrated that DEAH (Asp-Glu-Ala-His) box polypeptide 35 (DHX35), a member of the DExD/H (Asp-Glu-x-Asp/His)-box helicase family, boosts antiviral innate immunity in human airway epithelial cells. DHX35 knockdown attenuated the production of interferon-ß (IFN-ß), IL6, and CXCL10, whereas DHX35 overexpression increased their production. Upon stimulation, DHX35 was constitutively expressed, but it translocated from the nucleus into the cytosol, where it recognized cytosolic poly(I:C) and poly(dA:dT) via its HELICc domain. Mitochondrial antiviral signaling protein (MAVS) acted as an adaptor for DHX35 and interacted with the HELICc domain of DHX35 using amino acids 360-510. Interestingly, DHX35 interacted with retinoic acid-inducible gene 1 (RIG-I), enhanced the binding affinity of RIG-I with poly(I:C) and poly(dA:dT), and formed a signalsome with MAVS to activate interferon regulatory factor 3 (IRF3), NF-κB-p65, and MAPK signaling pathways. These results indicate that DHX35 not only acted as a cytosolic nucleic acid sensor but also synergized with RIG-I to enhance antiviral immunity in human airway epithelial cells. Our results demonstrate a novel molecular mechanism for DHX35 in RIG-I-mediated innate immunity and provide a novel candidate for drug and vaccine design to control viral infections in the human airway.


Assuntos
Proteína DEAD-box 58 , RNA Helicases DEAD-box , Imunidade Inata , Receptores Imunológicos , Humanos , Proteína DEAD-box 58/metabolismo , Proteína DEAD-box 58/imunologia , RNA Helicases DEAD-box/metabolismo , RNA Helicases DEAD-box/imunologia , Receptores Imunológicos/metabolismo , Poli I-C/imunologia , Poli I-C/farmacologia , RNA Helicases/metabolismo , RNA Helicases/imunologia , Transdução de Sinais/imunologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Células Epiteliais/virologia , Células HEK293
9.
J Biol Chem ; 300(7): 107463, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38876304

RESUMO

Chemotherapeutic agents for treating colorectal cancer (CRC) primarily induce apoptosis in tumor cells. The ubiquitin-proteasome system is critical for apoptosis regulation. Deubiquitinating enzymes (DUBs) remove ubiquitin from substrates to reverse ubiquitination. Although over 100 DUB members have been discovered, the biological functions of only a small proportion of DUBs have been characterized. Here, we aimed to systematically identify the DUBs that contribute to the development of CRC. Among the DUBs, ubiquitin-specific protease 36 (USP36) is upregulated in CRC. We showed that the knockdown of USP36 induces intrinsic and extrinsic apoptosis. Through gene silencing and coimmunoprecipitation techniques, we identified survivin and cIAP1 as USP36 targets. Mechanistically, USP36 binds and removes lysine-11-linked ubiquitin chains from cIAP1 and lysine-48-linked ubiquitin chains from survivin to abolish protein degradation. Overexpression of USP36 disrupts the formation of the XIAP-second mitochondria-derived activator of caspase complex and promotes receptor-interacting protein kinase 1 ubiquitination, validating USP36 as an inhibitor to intrinsic and extrinsic apoptosis through deubiquitinating survivin and cIAP1. Therefore, our results suggest that USP36 is involved in CRC progression and is a potential therapeutic target.


Assuntos
Apoptose , Neoplasias Colorretais , Proteínas Inibidoras de Apoptose , Survivina , Ubiquitina Tiolesterase , Ubiquitinação , Humanos , Linhagem Celular Tumoral , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica , Proteínas Inibidoras de Apoptose/metabolismo , Proteínas Inibidoras de Apoptose/genética , Survivina/metabolismo , Survivina/genética , Ubiquitina Tiolesterase/metabolismo , Ubiquitina Tiolesterase/genética
10.
Nat Immunol ; 14(2): 172-8, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23222971

RESUMO

DDX41 is a sensor of intracellular double-stranded DNA (dsDNA) in myeloid dendritic cells (mDCs) that triggers a type I interferon response via the signaling adaptor STING. We identified the E3 ligase TRIM21 as a DDX41-interacting protein and found that knockdown of or deficiency in TRIM21 resulted in enhanced type I interferon responses to intracellular dsDNA and DNA viruses. Overexpression of TRIM21 resulted in more degradation of DDX41 and less production of interferon-ß (IFN-ß) in response to intracellular dsDNA. The SPRY-PRY domain of TRIM21 interacted with the DEADc domain of DDX41. Lys9 and Lys115 of DDX41 were the targets of TRIM21-mediated ubiquitination. TRIM21 is therefore an interferon-inducible E3 ligase that induces the Lys48 (K48)-linked ubiquitination and degradation of DDX41 and negatively regulates the innate immune response to intracellular dsDNA.


Assuntos
DNA Viral/imunologia , DNA/imunologia , Células Dendríticas/imunologia , Imunidade Inata , Ribonucleoproteínas/imunologia , Animais , DNA/genética , DNA Viral/genética , Células Dendríticas/patologia , Células Dendríticas/virologia , Regulação da Expressão Gênica , Interferon beta/biossíntese , Interferon beta/imunologia , Lisina/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Camundongos Transgênicos , Orthoreovirus de Mamíferos/fisiologia , Estrutura Terciária de Proteína , Proteólise , Ribonucleoproteínas/deficiência , Ribonucleoproteínas/genética , Transdução de Sinais/imunologia , Ubiquitinação , Vesiculovirus/fisiologia
11.
Nat Immunol ; 13(12): 1155-61, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23142775

RESUMO

The induction of type I interferons by the bacterial secondary messengers cyclic di-GMP (c-di-GMP) or cyclic di-AMP (c-di-AMP) is dependent on a signaling axis that involves the adaptor STING, the kinase TBK1 and the transcription factor IRF3. Here we identified the heliase DDX41 as a pattern-recognition receptor (PRR) that sensed both c-di-GMP and c-di-AMP. DDX41 specifically and directly interacted with c-di-GMP. Knockdown of DDX41 via short hairpin RNA in mouse or human cells inhibited the induction of genes encoding molecules involved in the innate immune response and resulted in defective activation of STING, TBK1 and IRF3 in response to c-di-GMP or c-di-AMP. Our results suggest a mechanism whereby c-di-GMP and c-di-AMP are detected by DDX41, which forms a complex with STING to signal to TBK1-IRF3 and activate the interferon response.


Assuntos
GMP Cíclico/análogos & derivados , RNA Helicases DEAD-box/metabolismo , Fosfatos de Dinucleosídeos/metabolismo , Interferon Tipo I/imunologia , Listeria monocytogenes/imunologia , Listeria monocytogenes/metabolismo , Receptores de Reconhecimento de Padrão/metabolismo , Animais , Linhagem Celular , GMP Cíclico/metabolismo , RNA Helicases DEAD-box/genética , Humanos , Imunidade Inata , Fator Regulador 3 de Interferon/metabolismo , Macrófagos/imunologia , Proteínas de Membrana/metabolismo , Camundongos , Proteínas Serina-Treonina Quinases/metabolismo , Interferência de RNA , RNA Interferente Pequeno , Receptores de Reconhecimento de Padrão/genética , Sistemas do Segundo Mensageiro , Transdução de Sinais
12.
Mar Drugs ; 22(6)2024 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-38921594

RESUMO

Endothelial hyperpermeability is pivotal in sepsis-associated multi-organ dysfunction. Increased von Willebrand factor (vWF) plasma levels, stemming from activated platelets and endothelium injury during sepsis, can bind to integrin αvß3, exacerbating endothelial permeability. Hence, targeting this pathway presents a potential therapeutic avenue for sepsis. Recently, we identified isaridin E (ISE), a marine-derived fungal cyclohexadepsipeptide, as a promising antiplatelet and antithrombotic agent with a low bleeding risk. ISE's influence on septic mortality and sepsis-induced lung injury in a mouse model of sepsis, induced by caecal ligation and puncture, is investigated in this study. ISE dose-dependently improved survival rates, mitigating lung injury, thrombocytopenia, pulmonary endothelial permeability, and vascular inflammation in the mouse model. ISE markedly curtailed vWF release from activated platelets in septic mice by suppressing vesicle-associated membrane protein 8 and soluble N-ethylmaleide-sensitive factor attachment protein 23 overexpression. Moreover, ISE inhibited healthy human platelet adhesion to cultured lipopolysaccharide (LPS)-stimulated human umbilical vein endothelial cells (HUVECs), thereby significantly decreasing vWF secretion and endothelial hyperpermeability. Using cilengitide, a selective integrin αvß3 inhibitor, it was found that ISE can improve endothelial hyperpermeability by inhibiting vWF binding to αvß3. Activation of the integrin αvß3-FAK/Src pathway likely underlies vWF-induced endothelial dysfunction in sepsis. In conclusion, ISE protects against sepsis by inhibiting endothelial hyperpermeability and platelet-endothelium interactions.


Assuntos
Plaquetas , Células Endoteliais da Veia Umbilical Humana , Sepse , Fator de von Willebrand , Animais , Sepse/tratamento farmacológico , Fator de von Willebrand/metabolismo , Humanos , Camundongos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Masculino , Plaquetas/efeitos dos fármacos , Plaquetas/metabolismo , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Integrina alfaVbeta3/metabolismo , Integrina alfaVbeta3/antagonistas & inibidores , Permeabilidade Capilar/efeitos dos fármacos
13.
Nat Immunol ; 12(10): 959-65, 2011 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-21892174

RESUMO

The recognition of pathogenic DNA is important to the initiation of antiviral responses. Here we report the identification of DDX41, a member of the DEXDc family of helicases, as an intracellular DNA sensor in myeloid dendritic cells (mDCs). Knockdown of DDX41 expression by short hairpin RNA blocked the ability of mDCs to mount type I interferon and cytokine responses to DNA and DNA viruses. Overexpression of both DDX41 and the membrane-associated adaptor STING together had a synergistic effect in promoting Ifnb promoter activity. DDX41 bound both DNA and STING and localized together with STING in the cytosol. Knockdown of DDX41 expression blocked activation of the mitogen-activated protein kinase TBK1 and the transcription factors NF-κB and IRF3 by B-form DNA. Our results suggest that DDX41 is an additional DNA sensor that depends on STING to sense pathogenic DNA.


Assuntos
RNA Helicases DEAD-box/fisiologia , DNA Helicases/fisiologia , DNA/metabolismo , Células Dendríticas/fisiologia , Proteínas de Membrana/fisiologia , Animais , Células da Medula Óssea/metabolismo , Células Cultivadas , Humanos , Interferons/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/virologia , Estrutura Terciária de Proteína , Transdução de Sinais
16.
Immunity ; 41(4): 592-604, 2014 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-25308333

RESUMO

Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is a pattern-recognition receptor for a variety of endogenous and exogenous ligands. However, LOX-1 function in the host immune response is not fully understood. Here, we report that LOX-1 expressed on dendritic cells (DCs) and B cells promotes humoral responses. On B cells LOX-1 signaling upregulated CCR7, promoting cellular migration toward lymphoid tissues. LOX-1 signaling on DCs licensed the cells to promote B cell differentiation into class-switched plasmablasts and led to downregulation of chemokine receptor CXCR5 and upregulation of chemokine receptor CCR10 on plasmablasts, enabling their exit from germinal centers and migration toward local mucosa and skin. Finally, we found that targeting influenza hemagglutinin 1 (HA1) subunit to LOX-1 elicited HA1-specific protective antibody responses in rhesus macaques. Thus, LOX-1 expressed on B cells and DC cells has complementary functions to promote humoral immune responses.


Assuntos
Linfócitos B/imunologia , Células Dendríticas/imunologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Switching de Imunoglobulina/imunologia , Receptores Depuradores Classe E/imunologia , Animais , Formação de Anticorpos/imunologia , Diferenciação Celular/imunologia , Movimento Celular/imunologia , Centro Germinativo/citologia , Humanos , Imunoglobulina A/biossíntese , Imunoglobulina G/biossíntese , Imunoglobulina M/biossíntese , Ativação Linfocitária/imunologia , Macaca mulatta , Masculino , Mucosa/citologia , Receptores CCR10/biossíntese , Receptores CCR7/biossíntese , Receptores CXCR5/biossíntese , Receptores Depuradores Classe E/biossíntese , Transdução de Sinais/imunologia , Pele/citologia
17.
Environ Geochem Health ; 45(9): 6853-6867, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-36566469

RESUMO

This study systematically analyzed the contents, compositions, and sources of polycyclic aromatic hydrocarbons (PAHs) in river sediments near an important energy and chemical base in northwest China. In addition, their possible adverse effects on the ecology and human health were assessed. The PAH concentrations in this study area ranged from 2641.28 to 16783.72 (ng/g dw). PAHs of medium molecular weight (3-ring and 4-ring) showed the largest proportion, followed by PAHs of higher molecular weight (5-ring and 6-ring). The results of molecular diagnostic ratios and principal component analysis revealed that PAHs in the region have complex sources, with incomplete combustion of local fossil fuels and traffic exhaust factors being the main sources. The total toxic equivalent concentration of PAHs varied from 10.05 to 760.26 ng/g, and according to the sediment quality guidelines, PAHs have high potential ecological risk in the lower reaches of the river. The mean effect range-median quotient for the region was 0.46, and the combined ecological risk was at moderate to high levels (21% probability of toxicity). The lifetime carcinogenic risks for adults and children exposed to PAHs were 2.95 × 10-3 and 1.87 × 10-2, respectively, which are much higher than the limit of 10-4, indicating moderate to high potential cancer risks. Therefore, the local government should consider taking some environmental remediation measures. This study can provide theoretical support for pollution prevention measures and ecological restoration strategies for rivers in resource-rich areas.


Assuntos
Hidrocarbonetos Policíclicos Aromáticos , Poluentes Químicos da Água , Criança , Humanos , Carvão Mineral/análise , Rios/química , Hidrocarbonetos Policíclicos Aromáticos/toxicidade , Hidrocarbonetos Policíclicos Aromáticos/análise , Monitoramento Ambiental , Poluentes Químicos da Água/toxicidade , Poluentes Químicos da Água/análise , Sedimentos Geológicos/química , Medição de Risco , China
18.
J Neurosci ; 41(6): 1274-1287, 2021 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-33380470

RESUMO

Microglia have crucial roles in sculpting synapses and maintaining neural circuits during development. To test the hypothesis that microglia continue to regulate neural circuit connectivity in adult brain, we have investigated the effects of chronic microglial depletion, via CSF1R inhibition, on synaptic connectivity in the visual cortex in adult mice of both sexes. We find that the absence of microglia dramatically increases both excitatory and inhibitory synaptic connections to excitatory cortical neurons assessed with functional circuit mapping experiments in acutely prepared adult brain slices. Microglia depletion leads to increased densities and intensities of perineuronal nets. Furthermore, in vivo calcium imaging across large populations of visual cortical neurons reveals enhanced neural activities of both excitatory neurons and parvalbumin-expressing interneurons in the visual cortex following microglia depletion. These changes recover following adult microglia repopulation. In summary, our new results demonstrate a prominent role of microglia in sculpting neuronal circuit connectivity and regulating subsequent functional activity in adult cortex.SIGNIFICANCE STATEMENT Microglia are the primary immune cell of the brain, but recent evidence supports that microglia play an important role in synaptic sculpting during development. However, it remains unknown whether and how microglia regulate synaptic connectivity in adult brain. Our present work shows chronic microglia depletion in adult visual cortex induces robust increases in perineuronal nets, and enhances local excitatory and inhibitory circuit connectivity to excitatory neurons. Microglia depletion increases in vivo neural activities of both excitatory neurons and parvalbumin inhibitory neurons. Our new results reveal new potential avenues to modulate adult neural plasticity by microglia manipulation to better treat brain disorders, such as Alzheimer's disease.


Assuntos
Microglia/metabolismo , Rede Nervosa/metabolismo , Estimulação Luminosa/métodos , Córtex Visual/metabolismo , Aminopiridinas/farmacologia , Animais , Feminino , Masculino , Camundongos , Microglia/química , Microglia/efeitos dos fármacos , Rede Nervosa/química , Rede Nervosa/efeitos dos fármacos , Pirróis/farmacologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Córtex Visual/química , Córtex Visual/efeitos dos fármacos
19.
Immunity ; 39(1): 123-35, 2013 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-23871209

RESUMO

The NLRP3 inflammasome plays a major role in innate immune responses by activating caspase-1, resulting in secretion of interleukin-18 (IL-18) and IL-1ß. Although cytosolic double-stranded RNA (dsRNA) and bacterial RNA are known to activate the NLRP3 inflammasome, the upstream sensor is unknown. We investigated the potential function of DExD/H-box RNA helicase family members (previously shown to sense cytosolic DNA and RNA to induce type 1 interferon responses) in RNA-induced NLRP3 inflammasome activation. Among the helicase family members tested, we found that targeting of DHX33 expression by short hairpin RNA efficiently blocked the activation of caspase-1 and secretion of IL-18 and IL-1ß in human macrophages that were activated by cytosolic poly I:C, reoviral RNA, or bacterial RNA. DHX33 bound dsRNA via the helicase C domain. DHX33 interacted with NLRP3 and formed the inflammasome complex following stimulation with RNA. We therefore identified DHX33 as a cytosolic RNA sensor that activates the NLRP3 inflammasome.


Assuntos
Proteínas de Transporte/imunologia , RNA Helicases DEAD-box/imunologia , Inflamassomos/imunologia , Macrófagos/imunologia , RNA/imunologia , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Caspase 1/imunologia , Caspase 1/metabolismo , Linhagem Celular , Citosol/imunologia , Citosol/metabolismo , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Expressão Gênica/imunologia , Células HEK293 , Humanos , Immunoblotting , Inflamassomos/genética , Inflamassomos/metabolismo , Interleucina-18/imunologia , Interleucina-18/metabolismo , Interleucina-1beta/imunologia , Interleucina-1beta/metabolismo , Ativação de Macrófagos/imunologia , Macrófagos/metabolismo , Microscopia Confocal , Proteína 3 que Contém Domínio de Pirina da Família NLR , Poli I-C/imunologia , Ligação Proteica/imunologia , RNA/genética , RNA/metabolismo , Interferência de RNA , RNA Bacteriano/imunologia , RNA de Cadeia Dupla/genética , RNA de Cadeia Dupla/imunologia , RNA de Cadeia Dupla/metabolismo , RNA Viral/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa
20.
Microb Cell Fact ; 21(1): 59, 2022 Apr 09.
Artigo em Inglês | MEDLINE | ID: mdl-35397581

RESUMO

BACKGROUND: 7ß-hydroxylated steroids (7ß-OHSt) possess significant activities in anti-inflammatory and neuroprotection, and some of them have been widely used in clinics. However, the production of 7ß-OHSt is still a challenge due to the lack of cheap 7ß-hydroxy precursor and the difficulty in regio- and stereo-selectively hydroxylation at the inert C7 site of steroids in industry. The conversion of phytosterols by Mycolicibacterium species to the commercial precursor, androst-4-ene-3,17-dione (AD), is one of the basic ways to produce different steroids. This study presents a way to produce a basic 7ß-hydroxy precursor, 7ß-hydroxyandrost-4-ene-3,17-dione (7ß-OH-AD) in Mycolicibacterium, for 7ß-OHSt synthesis. RESULTS: A mutant of P450-BM3, mP450-BM3, was mutated and engineered into an AD producing strain for the efficient production of 7ß-OH-AD. The enzyme activity of mP450-BM3 was then increased by 1.38 times through protein engineering and the yield of 7ß-OH-AD was increased from 34.24 mg L- 1 to 66.25 mg L- 1. To further enhance the performance of 7ß-OH-AD producing strain, the regeneration of nicotinamide adenine dinucleotide phosphate (NADPH) for the activity of mP450-BM3-0 was optimized by introducing an NAD kinase (NADK) and a glucose-6-phosphate dehydrogenase (G6PDH). Finally, the engineered strain could produce 164.52 mg L- 1 7ß-OH-AD in the cofactor recycling and regeneration system. CONCLUSIONS: This was the first report on the one-pot biosynthesis of 7ß-OH-AD from the conversion of cheap phytosterols by an engineered microorganism, and the yield was significantly increased through the mutation of mP450-BM3 combined with overexpression of NADK and G6PDH. The present strategy may be developed as a basic industrial pathway for the commercial production of high value products from cheap raw materials.


Assuntos
Fitosteróis , Biotransformação , Mycobacteriaceae , Fitosteróis/metabolismo , Regeneração , Esteroides
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