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1.
Nat Commun ; 11(1): 4064, 2020 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-32792542

RESUMO

Regulation of the programming of tumour-associated macrophages (TAMs) controls tumour growth and anti-tumour immunity. We examined the role of FGF2 in that regulation. Tumours in mice genetically deficient in low-molecular weight FGF2 (FGF2LMW) regress dependent on T cells. Yet, TAMS not T cells express FGF receptors. Bone marrow derived-macrophages from Fgf2LMW-/- mice co-injected with cancer cells reduce tumour growth and express more inflammatory cytokines. FGF2 is induced in the tumour microenvironment following fractionated radiation in murine tumours consistent with clinical reports. Combination treatment of in vivo tumours with fractionated radiation and a blocking antibody to FGF2 prolongs tumour growth delay, increases long-term survival and leads to a higher iNOS+/CD206+ TAM ratio compared to irradiation alone. These studies show for the first time that FGF2 affects macrophage programming and is a critical regulator of immunity in the tumour microenvironment.


Assuntos
Fator 2 de Crescimento de Fibroblastos/metabolismo , Radioterapia/métodos , Animais , Linhagem Celular Tumoral , Fator 2 de Crescimento de Fibroblastos/genética , Células HT29 , Humanos , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Ativação de Macrófagos/efeitos da radiação , Lectinas de Ligação a Manose/genética , Lectinas de Ligação a Manose/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Nus , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/efeitos da radiação , Ensaios Antitumorais Modelo de Xenoenxerto
2.
Medicine (Baltimore) ; 99(34): e21821, 2020 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-32846824

RESUMO

BACKGROUND: Traditional Chinese medicine Tongxinluo (TXL) has been widely used to treat coronary artery disease in China, since it could reduce myocardial infarct size and ischemia/reperfusion injury in both non-diabetic and diabetic conditions. It has been shown that TXL could regulate peroxisome proliferator activated receptor-α (PPAR-α), a positive modulator of angiopoietin-like 4 (Angptl4), in diabetic rats. Endothelial junction substructure components, such as VE-cadherin, are involved in the protection of reperfusion injury. Thus, we hypothesized cell-intrinsic and endothelial-specific Angptl4 mediated the protection of TXL on endothelial barrier under high glucose condition against ischemia/reperfusion-injury via PPAR-α pathway. METHODS: Incubated with high glucose medium, the human cardiac microvascular endothelial cells (HCMECs) were then exposed to oxygen-glucose-serum deprivation (2 hours) and restoration (2 hours) stimulation, with or without TXL, insulin, or rhAngptl4 pretreatment. RESULTS: TXL, insulin, and rhAngptl4 had similar protective effects on the endothelial barrier. TXL treatment reversed the endothelial barrier breakdown in HCMECs significantly as identified by decreasing endothelial permeability, upregulating the expression of JAM-A, VE-cadherin, and integrin-α5 and increasing the membrane location of VE-cadherin and integrin-α5, and these effects of TXL were as effective as insulin and rhAngptl4. However, Angptl4 knock-down with small interfering RNA (siRNA) interference and PPAR-α inhibitor MK886 partially abrogated these beneficial effects of TXL. Western blotting also revealed that similar with insulin, TXL upregulated the expression of Angptl4 in HCMECs, which could be inhibited by Angptl4 siRNA or MK886 exposure. TXL treatment increased PPAR-α activity, which could be diminished by MK886 but not by Angptl4 siRNA. CONCLUSION: These data suggest cell-intrinsic and endothelial-specific Angptl4 mediates the protection of TXL against endothelial barrier breakdown during oxygen-glucose-serum deprivation and restoration under high glucose condition partly via the PPAR-α/Angptl4 pathway.


Assuntos
Proteína 4 Semelhante a Angiopoietina/metabolismo , Medicamentos de Ervas Chinesas/farmacologia , Células Endoteliais/efeitos dos fármacos , Endotélio/efeitos dos fármacos , Endotélio/fisiopatologia , PPAR alfa/metabolismo , Proteína 4 Semelhante a Angiopoietina/genética , Proteína 4 Semelhante a Angiopoietina/farmacologia , Caderinas/metabolismo , Permeabilidade Capilar , Moléculas de Adesão Celular/metabolismo , Células Cultivadas , Vasos Coronários/citologia , Técnicas de Silenciamento de Genes , Glucose/metabolismo , Glucose/farmacologia , Humanos , Indóis/farmacologia , Insulina/farmacologia , Integrina alfa5/metabolismo , Inibidores de Lipoxigenase/farmacologia , Microvasos/citologia , Oxigênio/metabolismo , Oxigênio/farmacologia , Receptores de Superfície Celular/metabolismo , Traumatismo por Reperfusão/metabolismo , Transdução de Sinais
3.
Cancer Treat Rev ; 88: 102064, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32622272

RESUMO

The overexpression of human epidermal growth factor receptor-2 (HER2) results in a biologically and clinically aggressive breast cancer (BC) subtype. Since the introduction of anti-HER2 targeted agents, survival rates of patients with HER2-positive metastatic BC have dramatically improved. Currently, although the treatment decision process in metastatic BC is primarily based on HER2 and hormone-receptor (HR) status, a rapidly growing body of data suggests that several other sources of biological heterogeneity may characterize HER2-positive metastatic BC. Moreover, pivotal clinical trials of new anti-HER2 antibody-drug conjugates showed encouraging results in HER2-low metastatic BC, thus leading to the possibility, in the near future, to expand the pool of patients suitable for HER2-targeted treatments. The present review summarizes and puts in perspective available evidence on biomarkers that hold the greatest promise to become potentially useful tools for optimizing HER2-positive metastatic BC patients' prognostic stratification and treatment in the next future. These biomarkers include HER2 levels and heterogeneity, HER3, intrinsic molecular subtypes by PAM50 analysis, DNA mutations, and immune-related factors. Molecular discordance between primary and metastatic tumors is also discussed.


Assuntos
Neoplasias da Mama/metabolismo , Receptor ErbB-2/metabolismo , Receptores de Superfície Celular/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Biomarcadores Tumorais/biossíntese , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/enzimologia , Neoplasias da Mama/genética , Feminino , Humanos , Metástase Neoplásica , Ensaios Clínicos Controlados Aleatórios como Assunto , Receptor ErbB-2/biossíntese , Receptor ErbB-2/genética , Receptor ErbB-3/biossíntese , Receptor ErbB-3/genética , Receptor ErbB-3/metabolismo
4.
Phys Rev Lett ; 125(1): 018102, 2020 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-32678664

RESUMO

Many types of cells require the ability to pinpoint the location of an external stimulus from the arrival of diffusing signaling molecules at cell-surface receptors. How does the organization (number and spatial configuration) of these receptors shape the limit of a cell's ability to infer the source location? In the idealized scenario of a spherical cell, we apply asymptotic analysis to compute splitting probabilities between individual receptors and formulate an information-theoretic framework to quantify the role of receptor organization. Clustered configurations of receptors provide an advantage in detecting sources aligned with the clusters, suggesting a possible multiscale mechanism for single-cell source inference.


Assuntos
Comunicação Celular/fisiologia , Modelos Biológicos , Receptores de Superfície Celular/fisiologia , Análise por Conglomerados , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Análise de Célula Única/métodos
5.
Virol Sin ; 35(3): 311-320, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32602046

RESUMO

The mechanism of how SARS-CoV-2 causes severe multi-organ failure is largely unknown. Acute kidney injury (AKI) is one of the frequent organ damage in severe COVID-19 patients. Previous studies have shown that human renal tubule cells could be the potential host cells targeted by SARS-CoV-2. Traditional cancer cell lines or immortalized cell lines are genetically and phenotypically different from host cells. Animal models are widely used, but often fail to reflect a physiological and pathogenic status because of species tropisms. There is an unmet need for normal human epithelial cells for disease modeling. In this study, we successfully established long term cultures of normal human kidney proximal tubule epithelial cells (KPTECs) in 2D and 3D culture systems using conditional reprogramming (CR) and organoids techniques. These cells had the ability to differentiate and repair DNA damage, and showed no transforming property. Importantly, the CR KPTECs maintained lineage function with expression of specific transporters (SLC34A3 and cubilin). They also expressed angiotensin-converting enzyme 2 (ACE2), a receptor for SARS-CoV and SARS-CoV-2. In contrast, cancer cell line did not express endogenous SLC34A3, cubilin and ACE2. Very interestingly, ACE2 expression was around twofold higher in 3D organoids culture compared to that in 2D CR culture condition. Pseudovirion assays demonstrated that SARS-CoV spike (S) protein was able to enter CR cells with luciferase reporter. This integrated 2D CR and 3D organoid cultures provide a physiological ex vivo model to study kidney functions, innate immune response of kidney cells to viruses, and a novel platform for drug discovery and safety evaluation.


Assuntos
Betacoronavirus/metabolismo , Técnicas de Cultura de Células/métodos , Infecções por Coronavirus/virologia , Coronavirus/metabolismo , Células Epiteliais/virologia , Rim/virologia , Pneumonia Viral/virologia , Animais , Betacoronavirus/patogenicidade , Linhagem Celular , Coronavirus/patogenicidade , Dano ao DNA , Modelos Animais de Doenças , Humanos , Organoides , Pandemias , Peptidil Dipeptidase A/metabolismo , Receptores de Superfície Celular/metabolismo , Vírus da SARS/metabolismo , Vírus da SARS/patogenicidade , Proteínas Cotransportadoras de Sódio-Fosfato Tipo IIc/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo
6.
Int J Nanomedicine ; 15: 4407-4415, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606685

RESUMO

Objective: Silica nanoparticles (SiO2 NPs) have been extensively employed in biomedical field. SiO2 NPs are primarily designed to enter the circulatory system; however, little information is available on potential adverse effects of SiO2 NPs on the nervous system. Methods: The neurotoxicity of SiO2 NPs at different concentrations (3, 6, 12 ng/nL) on zebrafish embryos was determined using immunofluorescence and microarray techniques, and subsequently confirmed by qRT-PCR. Results: SiO2 NPs disrupt the axonal integrity and decrease the length of axons in Tg (NBT: EGFP) transgenic lines. The number of apoptotic cells in the brain and central nervous system of zebrafish embryos was increased in the presence of 12 ng/nL of SiO2 NPs, but the difference did not reach statistical significance. Screening for changes in the expression of genes involved in the neuroactive ligand-receptor interaction pathway was performed by microarray and confirmed by qRT-PCR. These analyses demonstrated that SiO2 NPs markedly downregulated genes associated with neural function (grm6a, drd1b, chrnb3b, adrb2a, grin2ab, npffr2.1, npy8br, gabrd, chrma3, gabrg3, gria3a, grm1a, adra2b, and glra3). Conclusion: The obtained results documented that SiO2 NPs can induce developmental neurotoxicity by affecting the neuroactive ligand-receptor interaction signaling pathway. This new evidence may help to clarify the mechanism of SiO2 NPs-mediated neurotoxicity.


Assuntos
Embrião não Mamífero/metabolismo , Nanopartículas/toxicidade , Neurotoxinas/toxicidade , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Dióxido de Silício/toxicidade , Peixe-Zebra/embriologia , Animais , Apoptose/efeitos dos fármacos , Embrião não Mamífero/efeitos dos fármacos , Ligantes , Transdução de Sinais/efeitos dos fármacos
7.
Front Immunol ; 11: 1221, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32574271

RESUMO

Lactoferrin is a nutrient classically found in mammalian milk. It binds iron and is transferred via a variety of receptors into and between cells, serum, bile, and cerebrospinal fluid. It has important immunological properties, and is both antibacterial and antiviral. In particular, there is evidence that it can bind to at least some of the receptors used by coronaviruses and thereby block their entry. Of importance are Heparan Sulfate Proteoglycans (HSPGs) and the host receptor angiotensin-converting enzyme 2 (ACE2), as based on other activities lactoferrin might prevent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from attaching to the host cells. Lactoferrin (and more specifically enteric-coated LF because of increased bioavailability) may consequently be of preventive and therapeutic value during the present COVID-19 pandemic.


Assuntos
Proteoglicanas de Heparan Sulfato/metabolismo , Lactoferrina/fisiologia , Lactoferrina/uso terapêutico , Peptidil Dipeptidase A/metabolismo , Receptores Virais/metabolismo , Animais , Antibacterianos/uso terapêutico , Infecções Bacterianas/prevenção & controle , Suplementos Nutricionais , Humanos , Lactoferrina/metabolismo , Receptores de Superfície Celular/metabolismo , Viroses/prevenção & controle
8.
Front Immunol ; 11: 1229, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32574272

RESUMO

COVID-19 is caused by the Severe Acute Respiratory Syndrome (SARS) coronavirus (Cov)-2, an enveloped virus with a positive-polarity, single-stranded RNA genome. The initial outbreak of the pandemic began in December 2019, and it is affecting the human health of the global community. In common with previous pandemics (Influenza H1N1 and SARS-CoV) and the epidemics of Middle east respiratory syndrome (MERS)-CoV, CoVs target bronchial and alveolar epithelial cells. Virus protein ligands (e.g., haemagglutinin or trimeric spike glycoprotein for Influenza and CoV, respectively) interact with cellular receptors, such as (depending on the virus) either sialic acids, Dipeptidyl peptidase 4 (DPP4), or angiotensin-converting enzyme 2 (ACE2). Host proteases, e.g., cathepsins, furin, or members of the type II transmembrane serine proteases (TTSP) family, such as Transmembrane protease serine 2 (TMPRSS2), are involved in virus entry by proteolytically activating virus ligands. Also involved are Toll Like Receptor (TLR) family members, which upregulate anti-viral and pro-inflammatory mediators [interleukin (IL)-6 and IL-8 and type I and type III Interferons among others], through the activation of Nuclear Factor (NF)-kB. When these events (virus cellular entry and innate immune responses) are uncontrolled, a deleterious systemic response is sometimes encountered in infected patients, leading to the well-described "cytokine storm" and an ensuing multiple organ failure promoted by a downregulation of dendritic cell, macrophage, and T-cell function. We aim to describe how the lung and systemic host innate immune responses affect survival either positively, through downregulating initial viral load, or negatively, by triggering uncontrolled inflammation. An emphasis will be put on host cellular signaling pathways and proteases involved with a view on tackling these therapeutically.


Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Imunidade Inata , Pulmão/imunologia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/imunologia , Transdução de Sinais , Animais , Antivirais/uso terapêutico , Infecções por Coronavirus/metabolismo , Sistemas de Liberação de Medicamentos , Células Epiteliais/virologia , Humanos , Pulmão/virologia , Camundongos , Células Mieloides/virologia , Pandemias , Pneumonia Viral/metabolismo , Receptores de Superfície Celular/metabolismo , Receptores Virais/metabolismo , Serina Proteases/metabolismo , Internalização do Vírus
9.
Nature ; 583(7814): 127-132, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32555459

RESUMO

Cellular senescence is characterized by stable cell-cycle arrest and a secretory program that modulates the tissue microenvironment1,2. Physiologically, senescence serves as a tumour-suppressive mechanism that prevents the expansion of premalignant cells3,4 and has a beneficial role in wound-healing responses5,6. Pathologically, the aberrant accumulation of senescent cells generates an inflammatory milieu that leads to chronic tissue damage and contributes to diseases such as liver and lung fibrosis, atherosclerosis, diabetes and osteoarthritis1,7. Accordingly, eliminating senescent cells from damaged tissues in mice ameliorates the symptoms of these pathologies and even promotes longevity1,2,8-10. Here we test the therapeutic concept that chimeric antigen receptor (CAR) T cells that target senescent cells can be effective senolytic agents. We identify the urokinase-type plasminogen activator receptor (uPAR)11 as a cell-surface protein that is broadly induced during senescence and show that uPAR-specific CAR T cells efficiently ablate senescent cells in vitro and in vivo. CAR T cells that target uPAR extend the survival of mice with lung adenocarcinoma that are treated with a senescence-inducing combination of drugs, and restore tissue homeostasis in mice in which liver fibrosis is induced chemically or by diet. These results establish the therapeutic potential of senolytic CAR T cells for senescence-associated diseases.


Assuntos
Envelhecimento/patologia , Senescência Celular/imunologia , Cirrose Hepática/terapia , Longevidade/imunologia , Neoplasias Pulmonares/terapia , Receptores de Antígenos Quiméricos/imunologia , Rejuvenescimento , Linfócitos T/imunologia , Adenocarcinoma/imunologia , Adenocarcinoma/patologia , Adenocarcinoma/terapia , Animais , Tetracloreto de Carbono , Feminino , Xenoenxertos , Humanos , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/imunologia , Cirrose Hepática/patologia , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores de Antígenos Quiméricos/metabolismo , Receptores de Ativador de Plasminogênio Tipo Uroquinase/genética , Receptores de Ativador de Plasminogênio Tipo Uroquinase/metabolismo , Linfócitos T/metabolismo , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/metabolismo
10.
PLoS One ; 15(5): e0233383, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32428035

RESUMO

ERECTA gene family encodes leucine-rich repeat receptor-like kinases that control major aspects of plant development such as elongation of aboveground organs, leaf initiation, development of flowers, and epidermis differentiation. To clarify the importance of ERECTA signaling for the development of soybean (Glycine max), we expressed the dominant-negative ERECTA gene from Arabidopsis thaliana that is truncated in the kinase domain (AtΔKinase). Expression of AtΔKinase in soybean resulted in the short stature, reduced number of leaves, reduced leaf surface area and enhanced branching in the transgenic plants. The transgenic AtΔKinase soybean plants exhibited increased tolerance to water deficit stress due to the reduction of total leaf area and reduced transpiration compared to the wild-type plants. Production of seeds in AtΔKinase lines was higher compared to wild type at regular conditions of cultivation and after exposure to drought stress. Transgenic seedlings expressing AtΔKinase were also able to withstand salt stress better than the wild-type. Established results demonstrated the significance of native soybean genes (GmER and GmERL) in development and stress response of soybean, and suggested that the truncated ERECTA gene of Arabidopsis thaliana can be used to manipulate the growth and stress response of different crop species.


Assuntos
Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/farmacologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/farmacologia , Receptores de Superfície Celular/genética , Soja/crescimento & desenvolvimento , Estresse Fisiológico/efeitos dos fármacos , Arabidopsis , Proteínas de Arabidopsis/metabolismo , Folhas de Planta/metabolismo , Plantas Geneticamente Modificadas , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Superfície Celular/metabolismo , Soja/anatomia & histologia
11.
PLoS One ; 15(5): e0231813, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32442167

RESUMO

The interactions between membrane receptors and their endogenous ligands are key interactions in organisms. Recently, we have shown that a high number of genes encoding human receptors appeared at the same moment as their ligand in the animal tree of life. However, a set of receptors appeared before their present ligand. Different scenarios have been proposed to explain how a receptor can be conserved if its ligand is not yet appeared. However, these scenarios have been proposed individually and have never been studied in a global way. In this study, we investigated 30 mammalian pairs of ligand/receptor for which the first ligand appeared after its receptor in the tree of life, by using common indexes of selection, and proposed different scenarios explaining the earlier appearance of a receptor relative to its ligand. Based on 3D structural studies, our indexes allowed us to classify the evolution of these partners into different scenarios: 1) a scenario where the binding interface of the receptor is already present and under purifying selection before the appearance of the ligand; 2) a scenario where the binding interface seems to have appeared progressively, and 3) a scenario where the binding site seems to have been reshuffled since its appearance. As some scenarios were confirmed by the literature, we concluded that simple indexes can give a good highlight of the evolutive history of two partners that did not appear at the same time. Based on these scenarios, we also hypothesize that the replacement of a ligand by another is a frequent phenomenon during evolution.


Assuntos
Sítios de Ligação/genética , Evolução Molecular , Ligantes , Ligação Proteica/genética , Receptores de Superfície Celular , Sequência de Aminoácidos/genética , Animais , Simulação por Computador , Humanos , Modelos Moleculares , Conformação Molecular , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo
12.
PLoS Pathog ; 16(5): e1008501, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32369532

RESUMO

Plant-parasitic nematodes cause huge agricultural economic losses. Two major families of Bacillus thuringiensis crystal proteins, Cry5 and Cry6, show nematicidal activity. Previous work showed that binding to midgut receptors is a limiting step in Cry toxin mode of action. In the case of Cry5Ba, certain Caenorhabditis elegans glycolipids were identified as receptors of this toxin. However, the receptors for Cry6 toxin remain unknown. In this study, the C. elegans CUB-like-domain containing protein RBT-1, released by phosphatidylinositol-specific phospholipase C (PI-PLC), was identified as a Cry6Aa binding protein by affinity chromatography. RBT-1 contained a predicted glycosylphosphatidylinositol (GPI) anchor site and was shown to locate in lipid rafts in the surface of the midgut cells. Western ligand blot assays and ELISA binding analysis confirmed the binding interaction between Cry6Aa and RBT-1 showing high affinity and specificity. In addition, the mutation of rbt-1 gene decreased the susceptibility of C. elegans to Cry6Aa but not that of Cry5Ba. Furthermore, RBT-1 mediated the uptake of Cry6Aa into C. elegans gut cells, and was shown to be involved in triggering pore-formation activity, indicating that RBT-1 is required for the interaction of Cry6Aa with the nematode midgut cells. These results support that RBT-1 is a functional receptor for Cry6Aa.


Assuntos
Bacillus thuringiensis/metabolismo , Proteínas de Bactérias/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Endotoxinas/metabolismo , Proteínas Hemolisinas/metabolismo , Receptores de Superfície Celular/metabolismo , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/microbiologia , Proteínas de Caenorhabditis elegans/genética , Microdomínios da Membrana/genética , Microdomínios da Membrana/metabolismo , Mutação , Oligossacarídeos/genética , Oligossacarídeos/metabolismo , Receptores de Superfície Celular/genética
13.
Nat Immunol ; 21(6): 626-635, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32424362

RESUMO

The inflammasome NLRP6 plays a crucial role in regulating inflammation and host defense against microorganisms in the intestine. However, the molecular mechanisms by which NLRP6 function is inhibited to prevent excessive inflammation remain unclear. Here, we demonstrate that the deubiquitinase Cyld prevents excessive interleukin 18 (IL-18) production in the colonic mucosa by deubiquitinating NLRP6. We show that deubiquitination inhibited the NLRP6-ASC inflammasome complex and regulated the maturation of IL-18. Cyld deficiency in mice resulted in elevated levels of active IL-18 and severe colonic inflammation following Citrobacter rodentium infection. Further, in patients with ulcerative colitis, the concentration of active IL-18 was inversely correlated with CYLD expression. Thus, we have identified a novel regulatory mechanism that inhibits the NLRP6-IL-18 pathway in intestinal inflammation.


Assuntos
Enzima Desubiquitinante CYLD/metabolismo , Enterocolite/etiologia , Enterocolite/metabolismo , Inflamassomos/metabolismo , Interleucina-18/metabolismo , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Receptores de Superfície Celular/metabolismo , Animais , Citrobacter rodentium , Enzima Desubiquitinante CYLD/genética , Modelos Animais de Doenças , Suscetibilidade a Doenças , Infecções por Enterobacteriaceae/imunologia , Infecções por Enterobacteriaceae/metabolismo , Infecções por Enterobacteriaceae/microbiologia , Infecções por Enterobacteriaceae/patologia , Enterocolite/patologia , Expressão Gênica , Humanos , Interleucina-18/antagonistas & inibidores , Mucosa Intestinal/microbiologia , Mucosa Intestinal/patologia , Camundongos , Camundongos Knockout , Ligação Proteica/imunologia , Ubiquitinação
14.
PLoS One ; 15(5): e0225356, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32437440

RESUMO

High plasma LDL cholesterol (LDL-c) concentration is a major risk factor for atherosclerosis. Hepatic LDL receptor (LDLR) regulates LDL metabolism, and thereby plasma LDL-c concentration. Recently, we have identified the (pro)renin receptor [(P)RR] as a novel regulator of LDL metabolism, which regulates LDLR degradation and hence its protein abundance and activity. In silico analysis suggests that the (P)RR is a target of miR-148a. In this study we determined whether miR-148a could regulate LDL metabolism by regulating (P)RR expression in HepG2 and Huh7 cells. We found that miR-148a suppressed (P)RR expression by binding to the 3'-untranslated regions (3'-UTR) of the (P)RR mRNA. Mutating the binding sites for miR-148a in the 3'-UTR of (P)RR mRNA completely abolished the inhibitory effects of miR-148a on (P)RR expression. In line with our recent findings, reduced (P)RR expression resulted in decreased cellular LDL uptake, likely as a consequence of decreased LDLR protein abundance. Overexpressing the (P)RR prevented miR-148a-induced reduction in LDLR abundance and cellular LDL uptake. Our study supports a new concept that miR-148a is a regulator of (P)RR expression. By reducing (P)RR abundance, miR-148a decreases LDLR protein abundance and consequently cellular LDL uptake.


Assuntos
Lipoproteínas LDL/metabolismo , MicroRNAs/fisiologia , Receptores de Superfície Celular/metabolismo , Receptores de LDL/metabolismo , ATPases Vacuolares Próton-Translocadoras/metabolismo , Células HEK293 , Células Hep G2 , Humanos
15.
Nat Commun ; 11(1): 2285, 2020 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-32385277

RESUMO

Advanced metastatic cancer poses utmost clinical challenges and may present molecular and cellular features distinct from an early-stage cancer. Herein, we present single-cell transcriptome profiling of metastatic lung adenocarcinoma, the most prevalent histological lung cancer type diagnosed at stage IV in over 40% of all cases. From 208,506 cells populating the normal tissues or early to metastatic stage cancer in 44 patients, we identify a cancer cell subtype deviating from the normal differentiation trajectory and dominating the metastatic stage. In all stages, the stromal and immune cell dynamics reveal ontological and functional changes that create a pro-tumoral and immunosuppressive microenvironment. Normal resident myeloid cell populations are gradually replaced with monocyte-derived macrophages and dendritic cells, along with T-cell exhaustion. This extensive single-cell analysis enhances our understanding of molecular and cellular dynamics in metastatic lung cancer and reveals potential diagnostic and therapeutic targets in cancer-microenvironment interactions.


Assuntos
Adenocarcinoma de Pulmão/genética , Reprogramação Celular/genética , Neoplasias Pulmonares/genética , Análise de Sequência de RNA , Análise de Célula Única , Imunidade Adaptativa , Adenocarcinoma de Pulmão/irrigação sanguínea , Adenocarcinoma de Pulmão/imunologia , Adenocarcinoma de Pulmão/patologia , Linhagem da Célula , Progressão da Doença , Células Endoteliais/patologia , Humanos , Ligantes , Neoplasias Pulmonares/irrigação sanguínea , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Células Mieloides/patologia , Miofibroblastos/patologia , Metástase Neoplásica , Estadiamento de Neoplasias , Neovascularização Patológica/patologia , Fenótipo , Receptores de Superfície Celular/metabolismo , Células Estromais/metabolismo , Análise de Sobrevida
16.
Nat Commun ; 11(1): 2277, 2020 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-32385295

RESUMO

Cullin RING-type E3 ubiquitin ligases SCFTIR1/AFB1-5 and their AUX/IAA targets perceive the phytohormone auxin. The F-box protein TIR1 binds a surface-exposed degron in AUX/IAAs promoting their ubiquitylation and rapid auxin-regulated proteasomal degradation. Here, by adopting biochemical, structural proteomics and in vivo approaches we unveil how flexibility in AUX/IAAs and regions in TIR1 affect their conformational ensemble allowing surface accessibility of degrons. We resolve TIR1·auxin·IAA7 and TIR1·auxin·IAA12 complex topology, and show that flexible intrinsically disordered regions (IDRs) in the degron's vicinity, cooperatively position AUX/IAAs on TIR1. We identify essential residues at the TIR1 N- and C-termini, which provide non-native interaction interfaces with IDRs and the folded PB1 domain of AUX/IAAs. We thereby establish a role for IDRs in modulating auxin receptor assemblies. By securing AUX/IAAs on two opposite surfaces of TIR1, IDR diversity supports locally tailored positioning for targeted ubiquitylation, and might provide conformational flexibility for a multiplicity of functional states.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Ácidos Indolacéticos/metabolismo , Proteínas Intrinsicamente Desordenadas/metabolismo , Proteólise , Receptores de Superfície Celular/metabolismo , Proteínas de Arabidopsis/química , Sítios de Ligação , Proteínas Intrinsicamente Desordenadas/química , Simulação de Dinâmica Molecular , Mutação/genética , Filogenia , Domínios Proteicos , Ubiquitinação
17.
Nat Commun ; 11(1): 2674, 2020 05 29.
Artigo em Inglês | MEDLINE | ID: mdl-32471987

RESUMO

Increasing evidence indicates that guidance molecules used during development for cellular and axonal navigation also play roles in synapse maturation and homeostasis. In C. elegans the netrin receptor UNC-40/DCC controls the growth of dendritic-like muscle cell extensions towards motoneurons and is required to recruit type A GABA receptors (GABAARs) at inhibitory neuromuscular junctions. Here we show that activation of UNC-40 assembles an intracellular synaptic scaffold by physically interacting with FRM-3, a FERM protein orthologous to FARP1/2. FRM-3 then recruits LIN-2, the ortholog of CASK, that binds the synaptic adhesion molecule NLG-1/Neuroligin and physically connects GABAARs to prepositioned NLG-1 clusters. These processes are orchestrated by the synaptic organizer CePunctin/MADD-4, which controls the localization of GABAARs by positioning NLG-1/neuroligin at synapses and regulates the synaptic content of GABAARs through the UNC-40-dependent intracellular scaffold. Since DCC is detected at GABA synapses in mammals, DCC might also tune inhibitory neurotransmission in the mammalian brain.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Moléculas de Adesão Celular/metabolismo , Receptores de Superfície Celular/metabolismo , Receptores de GABA-A/metabolismo , Transmissão Sináptica/fisiologia , Animais , Orientação de Axônios/fisiologia , Moléculas de Adesão Celular Neuronais/metabolismo , Proteínas do Citoesqueleto/metabolismo , Proteínas de Helminto/metabolismo , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Junção Neuromuscular/metabolismo , Sinapses/fisiologia
18.
PLoS One ; 15(5): e0233854, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32470006

RESUMO

Alpha-toxin (Hla) is a major virulence factor of Staphylococcus aureus (S. aureus) and plays an important role in S. aureus-induced pneumonia. It binds as a monomer to the cell surface of eukaryotic host cells and forms heptameric transmembrane pores. Sensitivities toward the toxin of various types of potential host cells have been shown to vary substantially, and the reasons for these differences are unclear. We used three human model airway epithelial cell lines (16HBE14o-, S9, A549) to correlate cell sensitivity (measured as rate of paracellular gap formation in the cell layers) with Hla monomer binding, presence of the potential Hla receptors ADAM10 or α5ß1 integrin, presence of the toxin-stabilizing factor caveolin-1 as well as plasma membrane lipid composition (phosphatidylserine/choline, sphingomyelin). The abundance of ADAM10 correlated best with gap formation or cell sensitivities, respectively, when the three cell types were compared. Caveolin-1 or α5ß1 integrin did not correlate with toxin sensitivity. The relative abundance of sphingomyelin in plasma membranes may also be used as a proxi for cellular sensitivity against alpha-toxin as sphingomyelin abundances correlated well with the intensities of alpha-toxin mediated gap formation in the cell layers.


Assuntos
Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/toxicidade , Membrana Celular/metabolismo , Células Epiteliais/metabolismo , Proteínas Hemolisinas/metabolismo , Proteínas Hemolisinas/toxicidade , Interações Hospedeiro-Patógeno , Sistema Respiratório/patologia , Células A549 , Caveolina 1/metabolismo , Membrana Celular/efeitos dos fármacos , Tamanho Celular , Células Epiteliais/efeitos dos fármacos , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Humanos , Modelos Biológicos , Fosfolipídeos/metabolismo , Ligação Proteica , Receptores de Superfície Celular/metabolismo
19.
FASEB J ; 34(5): 6008-6016, 2020 05.
Artigo em Inglês | MEDLINE | ID: covidwho-46910

RESUMO

The sudden outbreak of COVID-19 has led to more than seven thousand deaths. Unfortunately, there are no specific drugs available to cure this disease. Type 2 taste receptors (TAS2Rs) may play an important role in host defense mechanisms. Based on the idea of host-directed therapy (HDT), we performed a negative co-expression analysis using big data of 60 000 Affymetrix expression arrays and 5000 TCGA data sets to determine the functions of TAS2R10, which can be activated by numerous bitter substances. Excitingly, we found that the main functions of TAS2R10 involved controlling infectious diseases caused by bacteria, viruses, and parasites, suggesting that TAS2R10 is a key trigger of host defense pathways. To quickly guide the clinical treatment of 2019-nCoV, we searched currently available drugs that are agonists of TAS2Rs. We identified many cheap, available, and safe medicines, such as diphenidol, quinine, chloroquine, artemisinin, chlorpheniramine, yohimbine, and dextromethorphan, which may target the most common symptoms caused by 2019-nCoV. We suggest that a cocktail-like recipe of existing bitter drugs may help doctors to fight this catastrophic disease and that the general public may drink or eat bitter substances, such as coffee, tea, or bitter vegetables, to reduce the risk of infection.


Assuntos
Antivirais/uso terapêutico , Biologia Computacional , Infecções por Coronavirus/tratamento farmacológico , Pneumonia Viral/tratamento farmacológico , Receptores Acoplados a Proteínas-G/agonistas , Antivirais/farmacologia , Betacoronavirus , Bases de Dados Genéticas , Bases de Dados de Produtos Farmacêuticos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Pandemias , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores Acoplados a Proteínas-G/genética
20.
Nat Cell Biol ; 22(4): 389-400, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32231305

RESUMO

In mouse embryonic stem cells (mESCs), chemical blockade of Gsk3α/ß and Mek1/2 (2i) instructs a self-renewing ground state whose endogenous inducers are unknown. Here we show that the axon guidance cue Netrin-1 promotes naive pluripotency by triggering profound signalling, transcriptomic and epigenetic changes in mESCs. Furthermore, we demonstrate that Netrin-1 can substitute for blockade of Gsk3α/ß and Mek1/2 to sustain self-renewal of mESCs in combination with leukaemia inhibitory factor and regulates the formation of the mouse pluripotent blastocyst. Mechanistically, we reveal how Netrin-1 and the balance of its receptors Neo1 and Unc5B co-regulate Wnt and MAPK pathways in both mouse and human ESCs. Netrin-1 induces Fak kinase to inactivate Gsk3α/ß and stabilize ß-catenin while increasing the phosphatase activity of a Ppp2r2c-containing Pp2a complex to reduce Erk1/2 activity. Collectively, this work identifies Netrin-1 as a regulator of pluripotency and reveals that it mediates different effects in mESCs depending on its receptor dosage, opening perspectives for balancing self-renewal and lineage commitment.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Proteínas do Tecido Nervoso/genética , Receptores de Netrina/genética , Netrina-1/genética , Receptores de Superfície Celular/genética , Via de Sinalização Wnt/genética , Animais , Linhagem Celular , Embrião de Mamíferos , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Quinase 1 de Adesão Focal/genética , Quinase 1 de Adesão Focal/metabolismo , Glicogênio Sintase Quinase 3 beta/antagonistas & inibidores , Glicogênio Sintase Quinase 3 beta/genética , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/genética , Isoenzimas/metabolismo , Fator Inibidor de Leucemia/genética , Fator Inibidor de Leucemia/metabolismo , MAP Quinase Quinase 1/antagonistas & inibidores , MAP Quinase Quinase 1/genética , MAP Quinase Quinase 1/metabolismo , MAP Quinase Quinase 2/antagonistas & inibidores , MAP Quinase Quinase 2/genética , MAP Quinase Quinase 2/metabolismo , Masculino , Camundongos , Camundongos Knockout , Camundongos SCID , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Receptores de Netrina/metabolismo , Netrina-1/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Proteína Fosfatase 2/genética , Proteína Fosfatase 2/metabolismo , Receptores de Superfície Celular/metabolismo , beta Catenina/genética , beta Catenina/metabolismo
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