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1.
Cell ; 187(9): 2209-2223.e16, 2024 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-38670073

RESUMO

Nuclear factor κB (NF-κB) plays roles in various diseases. Many inflammatory signals, such as circulating lipopolysaccharides (LPSs), activate NF-κB via specific receptors. Using whole-genome CRISPR-Cas9 screens of LPS-treated cells that express an NF-κB-driven suicide gene, we discovered that the LPS receptor Toll-like receptor 4 (TLR4) is specifically dependent on the oligosaccharyltransferase complex OST-A for N-glycosylation and cell-surface localization. The tool compound NGI-1 inhibits OST complexes in vivo, but the underlying molecular mechanism remained unknown. We did a CRISPR base-editor screen for NGI-1-resistant variants of STT3A, the catalytic subunit of OST-A. These variants, in conjunction with cryoelectron microscopy studies, revealed that NGI-1 binds the catalytic site of STT3A, where it traps a molecule of the donor substrate dolichyl-PP-GlcNAc2-Man9-Glc3, suggesting an uncompetitive inhibition mechanism. Our results provide a rationale for and an initial step toward the development of STT3A-specific inhibitors and illustrate the power of contemporaneous base-editor and structural studies to define drug mechanism of action.


Assuntos
Sistemas CRISPR-Cas , Hexosiltransferases , Lipopolissacarídeos , Proteínas de Membrana , NF-kappa B , Transdução de Sinais , Receptor 4 Toll-Like , Hexosiltransferases/metabolismo , Hexosiltransferases/genética , NF-kappa B/metabolismo , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Humanos , Receptor 4 Toll-Like/metabolismo , Animais , Sistemas CRISPR-Cas/genética , Lipopolissacarídeos/metabolismo , Lipopolissacarídeos/farmacologia , Camundongos , Células HEK293 , Inflamação/metabolismo , Inflamação/genética , Glicosilação , Microscopia Crioeletrônica , Domínio Catalítico , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética
2.
Cell ; 183(5): 1312-1324.e10, 2020 11 25.
Artigo em Inglês | MEDLINE | ID: mdl-33212011

RESUMO

Interferon (IFN)-Is are crucial mediators of antiviral immunity and homeostatic immune system regulation. However, the source of IFN-I signaling under homeostatic conditions is unclear. We discovered that commensal microbes regulate the IFN-I response through induction of IFN-ß by colonic DCs. Moreover, the mechanism by which a specific commensal microbe induces IFN-ß was identified. Outer membrane (OM)-associated glycolipids of gut commensal microbes belonging to the Bacteroidetes phylum induce expression of IFN-ß. Using Bacteroides fragilis and its OM-associated polysaccharide A, we determined that IFN-ß expression was induced via TLR4-TRIF signaling. Antiviral activity of this purified microbial molecule against infection with either vesicular stomatitis virus (VSV) or influenza was demonstrated to be dependent on the induction of IFN-ß. In a murine VSV infection model, commensal-induced IFN-ß regulated natural resistance to virus infection. Due to the physiological importance of IFN-Is, discovery of an IFN-ß-inducing microbial molecule represents a potential approach for the treatment of some human diseases.


Assuntos
Imunidade Inata , Microbiota , Viroses/microbiologia , Animais , Bacteroides fragilis/fisiologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Colo/patologia , Colo/virologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/metabolismo , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Glicolipídeos/metabolismo , Imunidade Inata/efeitos dos fármacos , Interferon beta/sangue , Interferon beta/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Microbiota/efeitos dos fármacos , Polissacarídeos Bacterianos/farmacologia , Receptor 4 Toll-Like/metabolismo , Vesiculovirus/fisiologia , Viroses/genética
3.
Cell ; 178(3): 536-551.e14, 2019 07 25.
Artigo em Inglês | MEDLINE | ID: mdl-31257024

RESUMO

The expression of some proteins in the autophagy pathway declines with age, which may impact neurodegeneration in diseases, including Alzheimer's Disease. We have identified a novel non-canonical function of several autophagy proteins in the conjugation of LC3 to Rab5+, clathrin+ endosomes containing ß-amyloid in a process of LC3-associated endocytosis (LANDO). We found that LANDO in microglia is a critical regulator of immune-mediated aggregate removal and microglial activation in a murine model of AD. Mice lacking LANDO but not canonical autophagy in the myeloid compartment or specifically in microglia have a robust increase in pro-inflammatory cytokine production in the hippocampus and increased levels of neurotoxic ß-amyloid. This inflammation and ß-amyloid deposition were associated with reactive microgliosis and tau hyperphosphorylation. LANDO-deficient AD mice displayed accelerated neurodegeneration, impaired neuronal signaling, and memory deficits. Our data support a protective role for LANDO in microglia in neurodegenerative pathologies resulting from ß-amyloid deposition.


Assuntos
Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Endocitose , Proteínas Associadas aos Microtúbulos/metabolismo , Doença de Alzheimer/metabolismo , Animais , Proteína 5 Relacionada à Autofagia/deficiência , Proteína 5 Relacionada à Autofagia/genética , Proteínas Relacionadas à Autofagia/deficiência , Proteínas Relacionadas à Autofagia/genética , Antígenos CD36/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Hipocampo/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Peptídeos e Proteínas de Sinalização Intracelular/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Transgênicos , Microglia/citologia , Microglia/metabolismo , Células RAW 264.7 , Receptores Imunológicos/metabolismo , Receptor 4 Toll-Like/metabolismo
4.
Nat Immunol ; 22(3): 279-286, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33495652

RESUMO

The constituents of the gut microbiome are determined by the local habitat, which itself is shaped by immunological pressures, such as mucosal IgA. Using a mouse model of restricted antibody repertoire, we identified a role for antibody-microbe interactions in shaping a community of bacteria with an enhanced capacity to metabolize L-tyrosine. This model led to increased concentrations of p-cresol sulfate (PCS), which protected the host against allergic airway inflammation. PCS selectively reduced CCL20 production by airway epithelial cells due to an uncoupling of epidermal growth factor receptor (EGFR) and Toll-like receptor 4 (TLR4) signaling. Together, these data reveal a gut microbe-derived metabolite pathway that acts distally on the airway epithelium to reduce allergic airway responses, such as those underpinning asthma.


Assuntos
Anticorpos/metabolismo , Bactérias/metabolismo , Cresóis/metabolismo , Microbioma Gastrointestinal , Intestinos/microbiologia , Pulmão/metabolismo , Pneumonia/prevenção & controle , Hipersensibilidade Respiratória/prevenção & controle , Ésteres do Ácido Sulfúrico/metabolismo , Tirosina/metabolismo , Administração Oral , Alérgenos , Animais , Anticorpos/imunologia , Diversidade de Anticorpos , Bactérias/imunologia , Células Cultivadas , Quimiocina CCL20/metabolismo , Técnicas de Cocultura , Cresóis/administração & dosagem , Modelos Animais de Doenças , Receptores ErbB/metabolismo , Feminino , Interações Hospedeiro-Patógeno , Injeções Intravenosas , Pulmão/imunologia , Pulmão/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Pneumonia/imunologia , Pneumonia/metabolismo , Pneumonia/microbiologia , Hipersensibilidade Respiratória/imunologia , Hipersensibilidade Respiratória/metabolismo , Hipersensibilidade Respiratória/microbiologia , Transdução de Sinais , Ésteres do Ácido Sulfúrico/administração & dosagem , Receptor 4 Toll-Like/metabolismo , Tirosina/administração & dosagem
5.
Annu Rev Immunol ; 29: 139-62, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21219181

RESUMO

A key question in immunology concerns how sterile injury activates innate immunity to mediate damaging inflammation in the absence of foreign invaders. The discovery that HMGB1, a ubiquitous nuclear protein, mediates the activation of innate immune responses led directly to the understanding that HMGB1 plays a critical role at the intersection of the host inflammatory response to sterile and infectious threat. HMGB1 is actively released by stimulation of the innate immune system with exogenous pathogen-derived molecules and is passively released by ischemia or cell injury in the absence of invasion. Established molecular mechanisms of HMGB1 binding and signaling through TLR4 reveal signaling pathways that mediate cytokine release and tissue damage. Experimental strategies that selectively target HMGB1 and TLR4 effectively reverse and prevent activation of innate immunity and significantly attenuate damage in diverse models of sterile and infection-induced threat.


Assuntos
Proteína HMGB1/antagonistas & inibidores , Infecções/tratamento farmacológico , Inflamação/tratamento farmacológico , Animais , Sistemas de Liberação de Medicamentos , Proteína HMGB1/metabolismo , Humanos , Imunidade Inata , Infecções/metabolismo , Inflamação/metabolismo , Receptor 4 Toll-Like/metabolismo
6.
Immunity ; 57(3): 462-477.e9, 2024 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-38430908

RESUMO

Inducible nucleosome remodeling at hundreds of latent enhancers and several promoters shapes the transcriptional response to Toll-like receptor 4 (TLR4) signaling in macrophages. We aimed to define the identities of the transcription factors that promote TLR-induced remodeling. An analysis strategy based on ATAC-seq and single-cell ATAC-seq that enriched for genomic regions most likely to undergo remodeling revealed that the transcription factor nuclear factor κB (NF-κB) bound to all high-confidence peaks marking remodeling during the primary response to the TLR4 ligand, lipid A. Deletion of NF-κB subunits RelA and c-Rel resulted in the loss of remodeling at high-confidence ATAC-seq peaks, and CRISPR-Cas9 mutagenesis of NF-κB-binding motifs impaired remodeling. Remodeling selectivity at defined regions was conferred by collaboration with other inducible factors, including IRF3- and MAP-kinase-induced factors. Thus, NF-κB is unique among TLR4-activated transcription factors in its broad contribution to inducible nucleosome remodeling, alongside its ability to activate poised enhancers and promoters assembled into open chromatin.


Assuntos
NF-kappa B , Receptor 4 Toll-Like , NF-kappa B/metabolismo , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo , Nucleossomos , Transdução de Sinais , Regulação da Expressão Gênica , Fator de Transcrição RelA/metabolismo
7.
Nat Immunol ; 21(1): 30-41, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31819254

RESUMO

NLRP3-inflammasome-driven inflammation is involved in the pathogenesis of a variety of diseases. Identification of endogenous inflammasome activators is essential for the development of new anti-inflammatory treatment strategies. Here, we identified that apolipoprotein C3 (ApoC3) activates the NLRP3 inflammasome in human monocytes by inducing an alternative NLRP3 inflammasome via caspase-8 and dimerization of Toll-like receptors 2 and 4. Alternative inflammasome activation in human monocytes is mediated by the Toll-like receptor adapter protein SCIMP. This triggers Lyn/Syk-dependent calcium entry and the production of reactive oxygen species, leading to activation of caspase-8. In humanized mouse models, ApoC3 activated human monocytes in vivo to impede endothelial regeneration and promote kidney injury in an NLRP3- and caspase-8-dependent manner. These data provide new insights into the regulation of the NLRP3 inflammasome and the pathophysiological role of triglyceride-rich lipoproteins containing ApoC3. Targeting ApoC3 might prevent organ damage and provide an anti-inflammatory treatment for vascular and kidney diseases.


Assuntos
Injúria Renal Aguda/imunologia , Apolipoproteína C-III/imunologia , Caspase 8/metabolismo , Nefropatias/imunologia , Monócitos/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Injúria Renal Aguda/patologia , Proteínas Adaptadoras de Transdução de Sinal , Animais , Apolipoproteína C-III/genética , Linhagem Celular , Modelos Animais de Doenças , Células HEK293 , Humanos , Inflamassomos/imunologia , Inflamação/genética , Inflamação/imunologia , Nefropatias/patologia , Proteínas de Membrana , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Espécies Reativas de Oxigênio/metabolismo , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo
8.
Genes Dev ; 38(11-12): 536-553, 2024 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-38918046

RESUMO

The five NF-κB family members and three nuclear IκB proteins play important biological roles, but the mechanisms by which distinct members of these protein families contribute to selective gene transcription remain poorly understood, especially at a genome-wide scale. Using nascent transcript RNA-seq, we observed considerable overlap between p50-dependent and IκBζ-dependent genes in Toll-like receptor 4 (TLR4)-activated macrophages. Key immunoregulatory genes, including Il6, Il1b, Nos2, Lcn2, and Batf, are among the p50-IκBζ-codependent genes. IκBζ-bound genomic sites are occupied at earlier time points by NF-κB dimers. However, p50-IκBζ codependence does not coincide with preferential binding of either p50 or IκBζ, as RelA co-occupies hundreds of genomic sites with the two proteins. A common feature of p50-IκBζ-codependent genes is a nearby p50/RelA/IκBζ-cobound site exhibiting p50-dependent binding of both RelA and IκBζ. This and other results suggest that IκBζ acts in concert with RelA:p50 heterodimers. Notably, p50-IκBζ-codependent genes comprise a high percentage of genes exhibiting the greatest differential expression between TLR4-stimulated and tumor necrosis factor receptor (TNFR)-stimulated macrophages. Thus, our genome-centric analysis reveals a defined p50-IκBζ pathway that selectively activates a set of key immunoregulatory genes and serves as an important contributor to differential TNFR and TLR4 responses.


Assuntos
Regulação da Expressão Gênica , Macrófagos , Subunidade p50 de NF-kappa B , Transdução de Sinais , Receptor 4 Toll-Like , Animais , Camundongos , Proteínas Adaptadoras de Transdução de Sinal , Proteínas I-kappa B/genética , Proteínas I-kappa B/metabolismo , Inflamação/genética , Inflamação/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Subunidade p50 de NF-kappa B/genética , Subunidade p50 de NF-kappa B/metabolismo , Ligação Proteica , Transdução de Sinais/genética , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo , Fator de Transcrição RelA/metabolismo , Fator de Transcrição RelA/genética , Masculino
9.
Nat Immunol ; 20(6): 677-686, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31110312

RESUMO

Consumption of a high-energy Western diet triggers mild adaptive ß cell proliferation to compensate for peripheral insulin resistance; however, the underlying molecular mechanism remains unclear. In the present study we show that the toll-like receptors TLR2 and TLR4 inhibited the diet-induced replication of ß cells in mice and humans. The combined, but not the individual, loss of TLR2 and TLR4 increased the replication of ß cells, but not that of α cells, leading to enlarged ß cell area and hyperinsulinemia in diet-induced obesity. Loss of TLR2 and TLR4 increased the nuclear abundance of the cell cycle regulators cyclin D2 and Cdk4 in a manner dependent on the signaling mediator Erk. These data reveal a regulatory mechanism controlling the proliferation of ß cells in diet-induced obesity and suggest that selective targeting of the TLR2/TLR4 pathways may reverse ß cell failure in patients with diabetes.


Assuntos
Células Secretoras de Insulina/metabolismo , Obesidade/etiologia , Obesidade/metabolismo , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética , Animais , Proliferação de Células , Ciclina D2/metabolismo , Quinase 4 Dependente de Ciclina/metabolismo , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Feminino , Humanos , Insulina/sangue , Insulina/metabolismo , Células Secretoras de Insulina/ultraestrutura , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/metabolismo , Sistema de Sinalização das MAP Quinases , Masculino , Camundongos , Camundongos Knockout , Complexos Multiproteicos/metabolismo , Obesidade/tratamento farmacológico , Parabiose , Ligação Proteica , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo
10.
Immunity ; 55(9): 1591-1593, 2022 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-36103858

RESUMO

The invasive fungal pathogen Cryptococcus neoformans promotes type 2 immunity to escape host defenses by unknown mechanisms. In a recent issue of Nature, Dang and colleagues identify a secreted fungal protein that triggers TLR4 signaling and supports a type 2 permissive environment and C. neoformans growth.


Assuntos
Criptococose , Cryptococcus neoformans , Criptococose/metabolismo , Criptococose/microbiologia , Cryptococcus neoformans/metabolismo , Proteínas Fúngicas/metabolismo , Receptor 4 Toll-Like/metabolismo , Virulência
11.
Genes Dev ; 37(13-14): 605-620, 2023 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-37536952

RESUMO

The transcription factor RUNX1 is mutated in familial platelet disorder with associated myeloid malignancy (FPDMM) and in sporadic myelodysplastic syndrome and leukemia. RUNX1 was shown to regulate inflammation in multiple cell types. Here we show that RUNX1 is required in granulocyte-monocyte progenitors (GMPs) to epigenetically repress two inflammatory signaling pathways in neutrophils: Toll-like receptor 4 (TLR4) and type I interferon (IFN) signaling. RUNX1 loss in GMPs augments neutrophils' inflammatory response to the TLR4 ligand lipopolysaccharide through increased expression of the TLR4 coreceptor CD14. RUNX1 binds Cd14 and other genes encoding proteins in the TLR4 and type I IFN signaling pathways whose chromatin accessibility increases when RUNX1 is deleted. Transcription factor footprints for the effectors of type I IFN signaling-the signal transducer and activator of transcription (STAT1::STAT2) and interferon regulatory factors (IRFs)-were enriched in chromatin that gained accessibility in both GMPs and neutrophils when RUNX1 was lost. STAT1::STAT2 and IRF motifs were also enriched in the chromatin of retrotransposons that were derepressed in RUNX1-deficient GMPs and neutrophils. We conclude that a major direct effect of RUNX1 loss in GMPs is the derepression of type I IFN and TLR4 signaling, resulting in a state of fixed maladaptive innate immunity.


Assuntos
Neutrófilos , Receptor 4 Toll-Like , Receptor 4 Toll-Like/metabolismo , Monócitos/metabolismo , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Citocinas/metabolismo , Cromatina/metabolismo , Fator de Transcrição STAT1/metabolismo
12.
Nat Immunol ; 18(10): 1104-1116, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28825701

RESUMO

Cross-regulation of Toll-like receptor (TLR) responses by cytokines is essential for effective host defense, avoidance of toxicity and homeostasis, but the underlying mechanisms are not well understood. Our comprehensive epigenomics approach to the analysis of human macrophages showed that the proinflammatory cytokines TNF and type I interferons induced transcriptional cascades that altered chromatin states to broadly reprogram responses induced by TLR4. TNF tolerized genes encoding inflammatory molecules to prevent toxicity while preserving the induction of genes encoding antiviral and metabolic molecules. Type I interferons potentiated the inflammatory function of TNF by priming chromatin to prevent the silencing of target genes of the transcription factor NF-κB that encode inflammatory molecules. The priming of chromatin enabled robust transcriptional responses to weak upstream signals. Similar chromatin regulation occurred in human diseases. Our findings reveal that signaling crosstalk between interferons and TNF is integrated at the level of chromatin to reprogram inflammatory responses, and identify previously unknown functions and mechanisms of action of these cytokines.


Assuntos
Epigênese Genética , Inflamação/etiologia , Inflamação/metabolismo , Interferon Tipo I/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Sítios de Ligação , Montagem e Desmontagem da Cromatina , Imunoprecipitação da Cromatina , Análise por Conglomerados , Biologia Computacional/métodos , Citocinas/genética , Citocinas/metabolismo , Epigenômica/métodos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Lipopolissacarídeos/imunologia , Lúpus Eritematoso Sistêmico/genética , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/metabolismo , Monócitos/imunologia , Monócitos/metabolismo , Motivos de Nucleotídeos , Regiões Promotoras Genéticas , Ligação Proteica , Transporte Proteico , Transdução de Sinais , Receptor 4 Toll-Like/metabolismo , Fatores de Transcrição/metabolismo
13.
Immunity ; 51(6): 997-1011.e7, 2019 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-31851905

RESUMO

Toll-like receptor (TLR) activation induces inflammatory responses in macrophages by activating temporally defined transcriptional cascades. Whether concurrent changes in the cellular metabolism that occur upon TLR activation influence the quality of the transcriptional responses remains unknown. Here, we investigated how macrophages adopt their metabolism early after activation to regulate TLR-inducible gene induction. Shortly after TLR4 activation, macrophages increased glycolysis and tricarboxylic acid (TCA) cycle volume. Metabolic tracing studies revealed that TLR signaling redirected metabolic fluxes to generate acetyl-Coenzyme A (CoA) from glucose resulting in augmented histone acetylation. Signaling through the adaptor proteins MyD88 and TRIF resulted in activation of ATP-citrate lyase, which in turn facilitated the induction of distinct LPS-inducible gene sets. We postulate that metabolic licensing of histone acetylation provides another layer of control that serves to fine-tune transcriptional responses downstream of TLR activation. Our work highlights the potential of targeting the metabolic-epigenetic axis in inflammatory settings.


Assuntos
ATP Citrato (pro-S)-Liase/metabolismo , Acetilcoenzima A/metabolismo , Histonas/metabolismo , Macrófagos/metabolismo , Receptor 4 Toll-Like/metabolismo , Acetilação , Proteínas Adaptadoras de Transporte Vesicular/genética , Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Animais , Ciclo do Ácido Cítrico/fisiologia , Glicólise/fisiologia , Humanos , Lipopolissacarídeos/metabolismo , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Transdução de Sinais , Transcrição Gênica/genética
14.
Immunity ; 50(2): 418-431.e6, 2019 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-30770245

RESUMO

Sepsis is a bi-phasic inflammatory disease that threatens approximately 30 million lives and claims over 14 million annually, yet little is known regarding the molecular switches and pathways that regulate this disease. Here, we have described ABCF1, an ATP-Binding Cassette (ABC) family member protein, which possesses an E2 ubiquitin enzyme activity, through which it controls the Lipopolysaccharide (LPS)- Toll-like Receptor-4 (TLR4) mediated gram-negative insult by targeting key proteins for K63-polyubiquitination. Ubiquitination by ABCF1 shifts the inflammatory profile from an early phase MyD88-dependent to a late phase TRIF-dependent signaling pathway, thereby regulating TLR4 endocytosis and modulating macrophage polarization from M1 to M2 phase. Physiologically, ABCF1 regulates the shift from the inflammatory phase of sepsis to the endotoxin tolerance phase, and modulates cytokine storm and interferon-ß (IFN-ß)-dependent production by the immunotherapeutic mediator, SIRT1. Consequently, ABCF1 controls sepsis induced mortality by repressing hypotension-induced renal circulatory dysfunction.


Assuntos
Transportadores de Cassetes de Ligação de ATP/imunologia , Macrófagos/imunologia , Sepse/imunologia , Choque Séptico/imunologia , Enzimas de Conjugação de Ubiquitina/imunologia , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/metabolismo , Trifosfato de Adenosina/imunologia , Trifosfato de Adenosina/metabolismo , Animais , Citocinas/imunologia , Citocinas/metabolismo , Feminino , Interferon beta/imunologia , Interferon beta/metabolismo , Lipopolissacarídeos/administração & dosagem , Lipopolissacarídeos/imunologia , Ativação de Macrófagos/efeitos dos fármacos , Ativação de Macrófagos/genética , Ativação de Macrófagos/imunologia , Macrófagos/classificação , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Interferência de RNA , Sepse/genética , Sepse/metabolismo , Choque Séptico/genética , Choque Séptico/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Receptor 4 Toll-Like/imunologia , Receptor 4 Toll-Like/metabolismo , Enzimas de Conjugação de Ubiquitina/genética , Enzimas de Conjugação de Ubiquitina/metabolismo , Ubiquitinação/imunologia
15.
Nature ; 608(7921): 161-167, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35896747

RESUMO

Invasive fungal pathogens are major causes of human mortality and morbidity1,2. Although numerous secreted effector proteins that reprogram innate immunity to promote virulence have been identified in pathogenic bacteria, so far, there are no examples of analogous secreted effector proteins produced by human fungal pathogens. Cryptococcus neoformans, the most common cause of fungal meningitis and a major pathogen in AIDS, induces a pathogenic type 2 response characterized by pulmonary eosinophilia and alternatively activated macrophages3-8. Here, we identify CPL1 as an effector protein secreted by C. neoformans that drives alternative activation (also known as M2 polarization) of macrophages to enable pulmonary infection in mice. We observed that CPL1-enhanced macrophage polarization requires Toll-like receptor 4, which is best known as a receptor for bacterial endotoxin but is also a poorly understood mediator of allergen-induced type 2 responses9-12. We show that this effect is caused by CPL1 itself and not by contaminating lipopolysaccharide. CPL1 is essential for virulence, drives polarization of interstitial macrophages in vivo, and requires type 2 cytokine signalling for its effect on infectivity. Notably, C. neoformans associates selectively with polarized interstitial macrophages during infection, suggesting a mechanism by which C. neoformans generates its own intracellular replication niche within the host. This work identifies a circuit whereby a secreted effector protein produced by a human fungal pathogen reprograms innate immunity, revealing an unexpected role for Toll-like receptor 4 in promoting the pathogenesis of infectious disease.


Assuntos
Criptococose , Cryptococcus neoformans , Proteínas Fúngicas , Hipersensibilidade , Inflamação , Receptor 4 Toll-Like , Fatores de Virulência , Animais , Criptococose/imunologia , Criptococose/microbiologia , Criptococose/patologia , Cryptococcus neoformans/imunologia , Cryptococcus neoformans/patogenicidade , Citocinas/imunologia , Proteínas Fúngicas/imunologia , Proteínas Fúngicas/metabolismo , Hipersensibilidade/imunologia , Hipersensibilidade/microbiologia , Imunidade Inata , Inflamação/imunologia , Inflamação/microbiologia , Lipopolissacarídeos/imunologia , Pulmão/imunologia , Pulmão/microbiologia , Macrófagos/citologia , Macrófagos/imunologia , Macrófagos/microbiologia , Camundongos , Receptor 4 Toll-Like/imunologia , Receptor 4 Toll-Like/metabolismo , Virulência , Fatores de Virulência/imunologia
16.
Immunity ; 48(1): 59-74.e5, 2018 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-29343440

RESUMO

Toll-like receptors (TLRs) sense pathogen-associated molecular patterns to activate the production of inflammatory mediators. TLR4 recognizes lipopolysaccharide (LPS) and drives the secretion of inflammatory cytokines, often contributing to sepsis. We report that transient receptor potential melastatin-like 7 (TRPM7), a non-selective but Ca2+-conducting ion channel, mediates the cytosolic Ca2+ elevations essential for LPS-induced macrophage activation. LPS triggered TRPM7-dependent Ca2+ elevations essential for TLR4 endocytosis and the subsequent activation of the transcription factor IRF3. In a parallel pathway, the Ca2+ signaling initiated by TRPM7 was also essential for the nuclear translocation of NFκB. Consequently, TRPM7-deficient macrophages exhibited major deficits in the LPS-induced transcriptional programs in that they failed to produce IL-1ß and other key pro-inflammatory cytokines. In accord with these defects, mice with myeloid-specific deletion of Trpm7 are protected from LPS-induced peritonitis. Our study highlights the importance of Ca2+ signaling in macrophage activation and identifies the ion channel TRPM7 as a central component of TLR4 signaling.


Assuntos
Cálcio/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Canais de Cátion TRPM/metabolismo , Receptor 4 Toll-Like/metabolismo , Animais , Técnicas de Cultura de Células , Endocitose/efeitos dos fármacos , Feminino , Citometria de Fluxo , Imunofluorescência , Regulação da Expressão Gênica , Técnicas de Genotipagem , Immunoblotting , Fator Regulador 3 de Interferon/metabolismo , Lipopolissacarídeos/farmacologia , Macrófagos/metabolismo , Masculino , Camundongos , NF-kappa B/metabolismo , Técnicas de Patch-Clamp , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Canais de Cátion TRPM/genética
17.
Proc Natl Acad Sci U S A ; 121(2): e2316104121, 2024 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-38165941

RESUMO

The nuclear receptor corepressor (NCoR) forms a complex with histone deacetylase 3 (HDAC3) that mediates repressive functions of unliganded nuclear receptors and other transcriptional repressors by deacetylation of histone substrates. Recent studies provide evidence that NCoR/HDAC3 complexes can also exert coactivator functions in brown adipocytes by deacetylating and activating PPARγ coactivator 1α (PGC1α) and that signaling via receptor activator of nuclear factor kappa-B (RANK) promotes the formation of a stable NCoR/HDAC3/PGC1ß complex that coactivates nuclear factor kappa-B (NFκB)- and activator protein 1 (AP-1)-dependent genes required for osteoclast differentiation. Here, we demonstrate that activation of Toll-like receptor (TLR) 4, but not TLR3, the interleukin 4 (IL4) receptor nor the Type I interferon receptor, also promotes assembly of an NCoR/HDAC3/PGC1ß coactivator complex. Receptor-specific utilization of TNF receptor-associated factor 6 (TRAF6) and downstream activation of extracellular signal-regulated kinase 1 (ERK1) and TANK-binding kinase 1 (TBK1) accounts for the common ability of RANK and TLR4 to drive assembly of an NCoR/HDAC3/PGC1ß complex in macrophages. ERK1, the p65 component of NFκB, and the p300 histone acetyltransferase (HAT) are also components of the induced complex and are associated with local histone acetylation and transcriptional activation of TLR4-dependent enhancers and promoters. These observations identify a TLR4/TRAF6-dependent signaling pathway that converts NCoR from a corepressor of nuclear receptors to a coactivator of NFκB and AP-1 that may be relevant to functions of NCoR in other developmental and homeostatic processes.


Assuntos
Histonas , Fator 6 Associado a Receptor de TNF , Ativação Transcricional , Proteínas Correpressoras/genética , Histonas/genética , Histonas/metabolismo , Fator 6 Associado a Receptor de TNF/genética , Fator 6 Associado a Receptor de TNF/metabolismo , Fator de Transcrição AP-1/metabolismo , Receptor 4 Toll-Like/metabolismo , Transdução de Sinais , NF-kappa B/genética , NF-kappa B/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo
18.
Proc Natl Acad Sci U S A ; 121(33): e2406492121, 2024 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-39361877

RESUMO

Toll-like receptors (TLRs) on macrophages sense microbial components and trigger the production of numerous cytokines and chemokines that mediate the inflammatory response to infection. Although many of the components required for the activation of the TLR pathway have been identified, the mechanisms that appropriately regulate the magnitude and duration of the response and ultimately restore homeostasis are less well understood. Furthermore, a growing body of work indicates that TLR signaling reciprocally interacts with other fundamental cellular processes, including lipid metabolism but only a few specific molecular links between immune signaling and the macrophage lipidome have been studied in detail. Oxysterol-binding protein (Osbp) is the founding member of a family of lipid-binding proteins with diverse functions in lipid sensing, lipid transport, and cell signaling but its role in TLR responses is not well defined. Here, we demonstrate that altering the state of Osbp with its natural ligand, 25-hydroxycholesterol (25HC), or pharmacologically, sustains and thereby amplifies Tlr4-induced cytokine production in vitro and in vivo. CRISPR-induced knockdown of Osbp abrogates the ability of these ligands to sustain TLR responses. Lipidomic analysis suggested that the effect of Osbp on TLR signaling may be mediated by alterations in triglyceride production and treating cells with a Dgat1 inhibitor, which blocks triglyceride production and completely abrogates the effect of Osbp on TLR signaling. Thus, Osbp is a sterol sensor that transduces perturbations of the lipidome to modulate the resolution of macrophage inflammatory responses.


Assuntos
Citocinas , Hidroxicolesteróis , Macrófagos , Receptores de Esteroides , Transdução de Sinais , Animais , Macrófagos/metabolismo , Macrófagos/imunologia , Camundongos , Citocinas/metabolismo , Receptores de Esteroides/metabolismo , Receptores de Esteroides/genética , Hidroxicolesteróis/metabolismo , Receptores Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo , Camundongos Endogâmicos C57BL , Metabolismo dos Lipídeos , Células RAW 264.7
19.
Proc Natl Acad Sci U S A ; 121(22): e2322524121, 2024 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-38781216

RESUMO

Long noncoding RNAs (lncRNAs) account for the largest portion of RNA from the transcriptome, yet most of their functions remain unknown. Here, we performed two independent high-throughput CRISPRi screens to understand the role of lncRNAs in monocyte function and differentiation. The first was a reporter-based screen to identify lncRNAs that regulate TLR4-NFkB signaling in human monocytes and the second screen identified lncRNAs involved in monocyte to macrophage differentiation. We successfully identified numerous noncoding and protein-coding genes that can positively or negatively regulate inflammation and differentiation. To understand the functional roles of lncRNAs in both processes, we chose to further study the lncRNA LOUP [lncRNA originating from upstream regulatory element of SPI1 (also known as PU.1)], as it emerged as a top hit in both screens. Not only does LOUP regulate its neighboring gene, the myeloid fate-determining factor SPI1, thereby affecting monocyte to macrophage differentiation, but knockdown of LOUP leads to a broad upregulation of NFkB-targeted genes at baseline and upon TLR4-NFkB activation. LOUP also harbors three small open reading frames capable of being translated and are responsible for LOUP's ability to negatively regulate TLR4/NFkB signaling. This work emphasizes the value of high-throughput screening to rapidly identify functional lncRNAs in the innate immune system.


Assuntos
Diferenciação Celular , Inflamação , Macrófagos , Monócitos , RNA Longo não Codificante , Transdução de Sinais , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Humanos , Macrófagos/metabolismo , Macrófagos/citologia , Diferenciação Celular/genética , Monócitos/metabolismo , Monócitos/citologia , Inflamação/genética , Inflamação/metabolismo , Receptor 4 Toll-Like/metabolismo , Receptor 4 Toll-Like/genética , NF-kappa B/metabolismo , Transativadores/metabolismo , Transativadores/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/genética , Sistemas CRISPR-Cas , Regulação da Expressão Gênica
20.
Cell ; 147(4): 868-80, 2011 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-22078883

RESUMO

The transport of Toll-like Receptors (TLRs) to various organelles has emerged as an essential means by which innate immunity is regulated. While most of our knowledge is restricted to regulators that promote the transport of newly synthesized receptors, the regulators that control TLR transport after microbial detection remain unknown. Here, we report that the plasma membrane localized Pattern Recognition Receptor (PRR) CD14 is required for the microbe-induced endocytosis of TLR4. In dendritic cells, this CD14-dependent endocytosis pathway is upregulated upon exposure to inflammatory mediators. We identify the tyrosine kinase Syk and its downstream effector PLCγ2 as important regulators of TLR4 endocytosis and signaling. These data establish that upon microbial detection, an upstream PRR (CD14) controls the trafficking and signaling functions of a downstream PRR (TLR4). This innate immune trafficking cascade illustrates how pathogen detection systems operate to induce both membrane transport and signal transduction.


Assuntos
Endocitose , Receptores de Lipopolissacarídeos/metabolismo , Receptor 4 Toll-Like/metabolismo , Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Animais , Células Dendríticas/imunologia , Endossomos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lipopolissacarídeos , Camundongos , Camundongos Endogâmicos C57BL , Fosfolipase C gama/metabolismo , Proteínas Tirosina Quinases/metabolismo , Transdução de Sinais , Quinase Syk
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