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1.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 28(5): 1710-1717, 2020 Oct.
Artigo em Chinês | MEDLINE | ID: mdl-33067979

RESUMO

AbstractObjective: To investigate the effect of ALAS2 downregulation on the expression of BNIP3L and erythroid differentiation in K562 cells. METHODS: The expression of ALAS2 was down-regulated by transfection of lentivirus, then quantitative real-time PCR was performed to detect the transfection efficiency. Flow cytometry analysis was applied to evaluate apoptosis of cells, erythroid differentiation, mitochondrial membrane potential and reactive oxygen species (ROS) level. Western blot was used to detect the BNIP3L expression, Co-immunoprecipitation was performed to analyze the relationship between ALAS2 and BNIP3L. RESULTS: Compared with sh-NC group, knockdown of ALAS2 induced downregulation of BNIP3L mRNA and protein expression(P<0.01) and erythroid related transcription factors GATA1, Nrf2 expression, as well as reduction of ROS level(P<0.05). Mitochondrial membrane potential of control (sh-NC) group was lower than that of shALAS2 group(P<0.05), but there was no significant change of cell apoptotic rate in two groups. CD71highCD235ahigh + CD71lowCD235ahigh cells of sh-NC and shALAS2 groups were 53.5%, 92.9% at 96 h after hemin induction, respectively. No direct action between ALAS2 and BNIP3L was observed. CONCLUSION: The intracellular heme level can affect the expression of BNIP3L which may be related with the regulation of ROS and transcription factors GATA1 and Nrf2. Higher BNIP3L facilitates cell differentiation but lower BNIP3L is favorable for cells survival.


Assuntos
Proteínas de Transporte , Mitofagia , 5-Aminolevulinato Sintetase/metabolismo , Apoptose , Diferenciação Celular , Humanos , Células K562 , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor
2.
Nat Commun ; 11(1): 4913, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-33004813

RESUMO

Reprograming of proline metabolism is critical for tumor growth. Here we show that PINCH-1 is highly expressed in lung adenocarcinoma and promotes proline synthesis through regulation of mitochondrial dynamics. Knockout (KO) of PINCH-1 increases dynamin-related protein 1 (DRP1) expression and mitochondrial fragmentation, which suppresses kindlin-2 mitochondrial translocation and interaction with pyrroline-5-carboxylate reductase 1 (PYCR1), resulting in inhibition of proline synthesis and cell proliferation. Depletion of DRP1 reverses PINCH-1 deficiency-induced defects on mitochondrial dynamics, proline synthesis and cell proliferation. Furthermore, overexpression of PYCR1 in PINCH-1 KO cells restores proline synthesis and cell proliferation, and suppresses DRP1 expression and mitochondrial fragmentation. Finally, ablation of PINCH-1 from lung adenocarcinoma in mouse increases DRP1 expression and inhibits PYCR1 expression, proline synthesis, fibrosis and tumor growth. Our results identify a signaling axis consisting of PINCH-1, DRP1 and PYCR1 that regulates mitochondrial dynamics and proline synthesis, and suggest an attractive strategy for alleviation of tumor growth.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adenocarcinoma de Pulmão/patologia , Proteínas com Domínio LIM/metabolismo , Neoplasias Pulmonares/patologia , Proteínas de Membrana/metabolismo , Células A549 , Proteínas Adaptadoras de Transdução de Sinal/genética , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/mortalidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Proliferação de Células/genética , Proteínas do Citoesqueleto/metabolismo , Modelos Animais de Doenças , Dinaminas/metabolismo , Feminino , Técnicas de Inativação de Genes , Humanos , Proteínas com Domínio LIM/genética , Pulmão/citologia , Pulmão/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidade , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Mitocôndrias/metabolismo , Dinâmica Mitocondrial , Proteínas Musculares/metabolismo , Proteínas de Neoplasias/metabolismo , Prolina/biossíntese , Proteínas Proto-Oncogênicas p21(ras)/genética , Pirrolina Carboxilato Redutases/metabolismo , Análise de Sobrevida
3.
Nat Commun ; 11(1): 4866, 2020 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-32978391

RESUMO

Mitochondria house evolutionarily conserved pathways of carbon and nitrogen metabolism that drive cellular energy production. Mitochondrial bioenergetics is regulated by calcium uptake through the mitochondrial calcium uniporter (MCU), a multi-protein complex whose assembly in the inner mitochondrial membrane is facilitated by the scaffold factor MCUR1. Intriguingly, many fungi that lack MCU contain MCUR1 homologs, suggesting alternate functions. Herein, we characterize Saccharomyces cerevisiae homologs Put6 and Put7 of MCUR1 as regulators of mitochondrial proline metabolism. Put6 and Put7 are tethered to the inner mitochondrial membrane in a large hetero-oligomeric complex, whose abundance is regulated by proline. Loss of this complex perturbs mitochondrial proline homeostasis and cellular redox balance. Yeast cells lacking either Put6 or Put7 exhibit a pronounced defect in proline utilization, which can be corrected by the heterologous expression of human MCUR1. Our work uncovers an unexpected role of MCUR1 homologs in mitochondrial proline metabolism.


Assuntos
Proteínas de Membrana/metabolismo , Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Prolina/metabolismo , Saccharomyces cerevisiae/metabolismo , Canais de Cálcio , Regulação Fúngica da Expressão Gênica , Genes Fúngicos/genética , Homeostase , Humanos , Proteínas de Membrana/genética , Redes e Vias Metabólicas/genética , Membranas Mitocondriais/metabolismo , Proteínas Mitocondriais/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crescimento & desenvolvimento , Transcriptoma
4.
Int Heart J ; 61(5): 1079-1083, 2020 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-32879264

RESUMO

A Japanese girl with polycystic kidney disease (PKD) developed normally, but at 8 months of age, she was hospitalized for acute onset dyspnea. On the day after admission to hospital, her general condition suddenly became worse. An echocardiogram showed left ventricular dilatation with thin walls, severe mitral valve regurgitation, and a reduced ejection fraction. She died of acute cardiac failure 3 hours after the sudden change. Postmortem analysis with light microscopy showed disarray of cardiomyocytes without obvious infiltration of lymphocytes, and we diagnosed her heart failure as idiopathic dilated cardiomyopathy (DCM). Clinical exome sequencing showed compound heterozygous variants in JPH2 (p.T237A/p.I414L) and a heterozygous nonsense mutation in PKD1 (p.Q4193*). To date, several variants in the JPH2 gene have been reported to be pathogenic for adult-onset hypertrophic cardiomyopathy or DCM in an autosomal dominant manner and infantile-onset DCM in an autosomal recessive manner. Additionally, autosomal dominant polycystic kidney disease is a systemic disease associated with several extrarenal manifestations, such as cardiomyopathy. Here we report a sudden infant death case of DCM and discuss the genetic variants of DCM and PKD.


Assuntos
Cardiomiopatia Dilatada/genética , Proteínas de Membrana/genética , Proteínas Musculares/genética , Rim Policístico Autossômico Dominante/genética , Canais de Cátion TRPP/genética , Cardiomiopatia Dilatada/complicações , Cardiomiopatia Dilatada/patologia , Morte Súbita Cardíaca/etiologia , Evolução Fatal , Feminino , Insuficiência Cardíaca/sangue , Insuficiência Cardíaca/etiologia , Heterozigoto , Humanos , Lactente , Insuficiência da Valva Mitral/etiologia , Miocárdio/patologia , Peptídeo Natriurético Encefálico/sangue , Fragmentos de Peptídeos/sangue
5.
Am J Physiol Lung Cell Mol Physiol ; 319(4): L670-L674, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32878480

RESUMO

The severity of coronavirus disease 2019 (COVID-19) is linked to an increasing number of risk factors, including exogenous (environmental) stimuli such as air pollution, nicotine, and cigarette smoke. These three factors increase the expression of angiotensin I converting enzyme 2 (ACE2), a key receptor involved in the entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-the etiological agent of COVID-19-into respiratory tract epithelial cells. Patients with severe COVID-19 are managed with oxygen support, as are at-risk individuals with chronic lung disease. To date, no study has examined whether an increased fraction of inspired oxygen (FiO2) may affect the expression of SARS-CoV-2 entry receptors and co-receptors, including ACE2 and the transmembrane serine proteases TMPRSS1, TMPRSS2, and TMPRSS11D. To address this, steady-state mRNA levels for genes encoding these SARS-CoV-2 receptors were assessed in the lungs of mouse pups chronically exposed to elevated FiO2, and in the lungs of preterm-born human infants chronically managed with an elevated FiO2. These two scenarios served as models of chronic elevated FiO2 exposure. Additionally, SARS-CoV-2 receptor expression was assessed in primary human nasal, tracheal, esophageal, bronchial, and alveolar epithelial cells, as well as primary mouse alveolar type II cells exposed to elevated oxygen concentrations. While gene expression of ACE2 was unaffected, gene and protein expression of TMPRSS11D was consistently upregulated by exposure to an elevated FiO2. These data highlight the need for further studies that examine the relative contribution of the various viral co-receptors on the infection cycle, and point to oxygen supplementation as a potential risk factor for COVID-19.


Assuntos
Infecções por Coronavirus/patologia , Proteínas de Membrana/metabolismo , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/patologia , Mucosa Respiratória/metabolismo , Serina Endopeptidases/metabolismo , Serina Proteases/metabolismo , Células Epiteliais Alveolares/metabolismo , Animais , Betacoronavirus , Células Cultivadas , Feminino , Humanos , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Oxigênio/administração & dosagem , Oxigênio/análise , Pandemias , Receptores Virais/metabolismo , Fatores de Risco , Serina Endopeptidases/genética , Serina Proteases/genética , Índice de Gravidade de Doença
6.
PLoS Genet ; 16(8): e1009003, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32866139

RESUMO

Sensory systems rely on neuromodulators, such as serotonin, to provide flexibility for information processing as stimuli vary, such as light intensity throughout the day. Serotonergic neurons broadly innervate the optic ganglia of Drosophila melanogaster, a widely used model for studying vision. It remains unclear whether serotonin modulates the physiology of interneurons in the optic ganglia. To address this question, we first mapped the expression patterns of serotonin receptors in the visual system, focusing on a subset of cells with processes in the first optic ganglion, the lamina. Serotonin receptor expression was found in several types of columnar cells in the lamina including 5-HT2B in lamina monopolar cell L2, required for spatiotemporal luminance contrast, and both 5-HT1A and 5-HT1B in T1 cells, whose function is unknown. Subcellular mapping with GFP-tagged 5-HT2B and 5-HT1A constructs indicated that these receptors localize to layer M2 of the medulla, proximal to serotonergic boutons, suggesting that the medulla neuropil is the primary site of serotonergic regulation for these neurons. Exogenous serotonin increased basal intracellular calcium in L2 terminals in layer M2 and modestly decreased the duration of visually induced calcium transients in L2 neurons following repeated dark flashes, but otherwise did not alter the calcium transients. Flies without functional 5-HT2B failed to show an increase in basal calcium in response to serotonin. 5-HT2B mutants also failed to show a change in amplitude in their response to repeated light flashes but other calcium transient parameters were relatively unaffected. While we did not detect serotonin receptor expression in L1 neurons, they, like L2, underwent serotonin-induced changes in basal calcium, presumably via interactions with other cells. These data demonstrate that serotonin modulates the physiology of interneurons involved in early visual processing in Drosophila.


Assuntos
Receptor 5-HT1B de Serotonina/genética , Receptores 5-HT1 de Serotonina/genética , Receptores 5-HT2 de Serotonina/genética , Neurônios Serotoninérgicos/metabolismo , Serotonina/metabolismo , Animais , Ritmo Circadiano/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Regulação da Expressão Gênica/genética , Interneurônios/metabolismo , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , Neurotransmissores/genética , Receptores de Serotonina/genética , Serotonina/genética , Percepção Visual/genética
7.
Zhongguo Dang Dai Er Ke Za Zhi ; 22(9): 980-983, 2020 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-32933630

RESUMO

OBJECTIVE: To study the expression level of cAMP response element-binding protein (CREB) in children with recurrent wheezing under three years of age and its effect on the expression of the serum orosomucoid 1-like protein 3 (ORMDL3) gene. METHODS: Thirty-six children with recurrent wheezing under three years of age who visited the hospital from June 2017 to June 2019 were selected as the recurrent wheezing group. Twenty-four healthy children from physical examination were selected as the control group. The CREB expression level in peripheral blood was measured by quantitative real-time PCR. Human bronchial epithelial cells (BEAS-2B) were cultured, and dual-luciferase reporter assay and quantitative real-time PCR were used to investigate the effects of overexpression and siRNA interference of CREB on the promoter activity and mRNA expression of the ORMDL3 gene in the BEAS-2B cells. RESULTS: The expression level of CREB in the recurrent wheezing group was significantly higher than that in the control group (P<0.001). In BEAS-2B cells, overexpression of CREB significantly up-regulated the promoter activity and mRNA expression of the ORMDL3 gene (P<0.05), while siRNA interference of CREB significantly reduced the promoter activity and mRNA expression of the ORMDL3 gene (P<0.05). CONCLUSIONS: The expression of CREB is increased in children with recurrent wheezing, and CREB may be involved in the pathogenesis of recurrent wheezing by regulating expression of the ORMDL3 gene.


Assuntos
Proteínas de Membrana/genética , Sons Respiratórios , Pré-Escolar , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico , Células Epiteliais , Humanos , Regiões Promotoras Genéticas
8.
PLoS One ; 15(9): e0236234, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32970676

RESUMO

BACKGROUND: Humans routinely attempt to manage pest rodent populations with anticoagulant rodenticides (ARs). We require information on resistance to ARs within rodent populations to have effective eradication programs that minimise exposure in non-target species. Mutations to the VKORC1 gene have been shown to confer resistance in rodents with high proportions of resistance in mice found in all European populations tested. We screened mutations in Mus musculus within Western Australia, by sampling populations from the capital city (Perth) and a remote island (Browse Island). These are the first Australian mouse populations screened for resistance using this method. Additionally, the mitochondrial D-loop of house mice was sequenced to explore population genetic structure, identify the origin of Western Australian mice, and to elucidate whether resistance was linked to certain haplotypes. RESULTS: No resistance-related VKORC1 mutations were detected in either house mouse population. A genetic introgression in the intronic sequence of the VKORC1 gene of Browse Island house mouse was detected which is thought to have originated through hybridisation with the Algerian mouse (Mus spretus). Analysis of the mitochondrial D-loop reported two haplotypes in the house mouse population of Perth, and two haplotypes in the population of Browse Island. CONCLUSIONS: Both house mouse populations exhibited no genetic resistance to ARs, in spite of free use of ARs in Western Australia. Therefore weaker anticoagulant rodenticides can be employed in pest control and eradication attempts, which will result in reduced negative impacts on non-target species. Biosecurity measures must be in place to avoid introduction of resistant house mice, and new house mouse subspecies to Western Australia.


Assuntos
Anticoagulantes/toxicidade , Proteínas de Membrana/genética , Camundongos/genética , Controle de Pragas , Rodenticidas/toxicidade , Vitamina K Epóxido Redutases/genética , Animais , Resistência a Medicamentos , Feminino , Variação Genética , Masculino , Mutação , Controle de Pragas/métodos , Austrália Ocidental
9.
Mol Cell ; 79(6): 1051-1065.e10, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32877643

RESUMO

Mitochondria contain their own gene expression systems, including membrane-bound ribosomes dedicated to synthesizing a few hydrophobic subunits of the oxidative phosphorylation (OXPHOS) complexes. We used a proximity-dependent biotinylation technique, BioID, coupled with mass spectrometry to delineate in baker's yeast a comprehensive network of factors involved in biogenesis of mitochondrial encoded proteins. This mitochondrial gene expression network (MiGENet) encompasses proteins involved in transcription, RNA processing, translation, or protein biogenesis. Our analyses indicate the spatial organization of these processes, thereby revealing basic mechanistic principles and the proteins populating strategically important sites. For example, newly synthesized proteins are directly handed over to ribosomal tunnel exit-bound factors that mediate membrane insertion, co-factor acquisition, or their mounting into OXPHOS complexes in a special early assembly hub. Collectively, the data reveal the connectivity of mitochondrial gene expression, reflecting a unique tailoring of the mitochondrial gene expression system.


Assuntos
Mitocôndrias/genética , Proteínas Mitocondriais/genética , Proteínas Ribossômicas/genética , Proteínas de Saccharomyces cerevisiae/genética , Regulação Fúngica da Expressão Gênica , Proteínas de Membrana/genética , Fosforilação Oxidativa , Biossíntese de Proteínas/genética , Saccharomyces cerevisiae/genética
10.
DNA Cell Biol ; 39(10): 1895-1906, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32882141

RESUMO

Acute aortic dissection (AD) is one of the most severe and highly mortality vascular disease. Its actual prevalence may be seriously underestimated. We studied different expression genes to understand gene profile change between acute AD and nondiseased individuals, and then discover potential biomarkers and therapeutic targets of acute AD. In our study, acute AD differentially expressed mRNAs and miRNAs were identified through bioinformatics analysis on Gene Expression Omnibus data sets GSE52093, GSE98770, and GSE92427. Then, comprehensive target prediction and network analysis methods were used to evaluate protein-protein interaction networks and to identify Gene Ontology terms for differentially expressed mRNAs. Differentially expressed mRNAs-miRNAs involved in acute AD were assessed as well. Finally, the quantitative real-time PCR and in vitro experiment was used to validate the results. We found Integral Membrane Protein 2C (ITM2C) was low expressed and miR-107-5p was highly expressed in acute AD tissues. Meanwhile, overexpression miR-107-5p promoted the cell proliferation and inhibited the cell apoptosis in RASMC cells. miR-107-5p inhibited the progression of acute AD through targeted ITM2C.


Assuntos
Aneurisma Dissecante/genética , MicroRNAs/genética , Aneurisma Dissecante/metabolismo , Aneurisma Dissecante/patologia , Animais , Apoptose , Biomarcadores/metabolismo , Proliferação de Células , Células Cultivadas , Redes Reguladoras de Genes , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , MicroRNAs/metabolismo , Miócitos de Músculo Liso/metabolismo , Mapas de Interação de Proteínas , Ratos , Transcriptoma
11.
Nat Commun ; 11(1): 4589, 2020 09 11.
Artigo em Inglês | MEDLINE | ID: mdl-32917887

RESUMO

Mandibuloacral dysplasia syndromes are mainly due to recessive LMNA or ZMPSTE24 mutations, with cardinal nuclear morphological abnormalities and dysfunction. We report five homozygous null mutations in MTX2, encoding Metaxin-2 (MTX2), an outer mitochondrial membrane protein, in patients presenting with a severe laminopathy-like mandibuloacral dysplasia characterized by growth retardation, bone resorption, arterial calcification, renal glomerulosclerosis and severe hypertension. Loss of MTX2 in patients' primary fibroblasts leads to loss of Metaxin-1 (MTX1) and mitochondrial dysfunction, including network fragmentation and oxidative phosphorylation impairment. Furthermore, patients' fibroblasts are resistant to induced apoptosis, leading to increased cell senescence and mitophagy and reduced proliferation. Interestingly, secondary nuclear morphological defects are observed in both MTX2-mutant fibroblasts and mtx-2-depleted C. elegans. We thus report the identification of a severe premature aging syndrome revealing an unsuspected link between mitochondrial composition and function and nuclear morphology, establishing a pathophysiological link with premature aging laminopathies and likely explaining common clinical features.


Assuntos
Acro-Osteólise/metabolismo , Predisposição Genética para Doença/genética , Lipodistrofia/metabolismo , Mandíbula/anormalidades , Proteínas de Membrana/metabolismo , Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Acro-Osteólise/diagnóstico por imagem , Acro-Osteólise/genética , Acro-Osteólise/patologia , Senilidade Prematura/genética , Senilidade Prematura/metabolismo , Animais , Apoptose , Caenorhabditis elegans , Proliferação de Células , Criança , Regulação para Baixo , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Regulação da Expressão Gênica , Genótipo , Homozigoto , Humanos , Lipodistrofia/diagnóstico por imagem , Lipodistrofia/genética , Lipodistrofia/patologia , Masculino , Mandíbula/diagnóstico por imagem , Proteínas de Membrana/genética , Metaloendopeptidases , Proteínas de Transporte da Membrana Mitocondrial/genética , Proteínas Mitocondriais/genética , Mutação , Fenótipo , Pele , Sequenciamento Completo do Genoma
12.
PLoS One ; 15(8): e0237514, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32790781

RESUMO

Golgi phosphoprotein 3 (GOLPH3) is a peripheral membrane protein localized at the trans-Golgi network that is also distributed in a large cytosolic pool. GOLPH3 has been involved in several post-Golgi protein trafficking events, but its precise function at the molecular level is not well understood. GOLPH3 is also considered the first oncoprotein of the Golgi apparatus, with important roles in several types of cancer. Yet, it is unknown how GOLPH3 is regulated to achieve its contribution in the mechanisms that lead to tumorigenesis. Binding of GOLPH3 to Golgi membranes depends on its interaction to phosphatidylinositol-4-phosphate. However, an early finding showed that GTP promotes the binding of GOLPH3 to Golgi membranes and vesicles. Nevertheless, it remains largely unknown whether this response is consequence of the function of GTP-dependent regulatory factors, such as proteins of the RAB family of small GTPases. Interestingly, in Drosophila melanogaster the ortholog of GOLPH3 interacts with- and behaves as effector of the ortholog of RAB1. However, there is no experimental evidence implicating GOLPH3 as a possible RAB1 effector in mammalian cells. Here, we show that human GOLPH3 interacted directly with either RAB1A or RAB1B, the two isoforms of RAB1 in humans. The interaction was nucleotide dependent and it was favored with GTP-locked active state variants of these GTPases, indicating that human GOLPH3 is a bona fide effector of RAB1A and RAB1B. Moreover, the expression in cultured cells of the GTP-locked variants resulted in less distribution of GOLPH3 in the Golgi apparatus, suggesting an intriguing model of GOLPH3 regulation.


Assuntos
Complexo de Golgi/metabolismo , Proteínas de Membrana/metabolismo , Proteínas rab1 de Ligação ao GTP/metabolismo , Células HeLa , Humanos , Proteínas de Membrana/genética , Transporte Proteico , Proteínas rab1 de Ligação ao GTP/genética , Rede trans-Golgi
13.
Genes (Basel) ; 11(8)2020 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-32785186

RESUMO

The coronaviruses are a large family of enveloped RNA viruses that commonly cause gastrointestinal or respiratory illnesses in the infected host. Avian coronavirus infectious bronchitis virus (IBV) is a highly contagious respiratory pathogen of chickens that can affect the kidneys and reproductive systems resulting in bird mortality and decreased reproductivity. The interferon-inducible transmembrane (IFITM) proteins are activated in response to viral infections and represent a class of cellular restriction factors that restrict the replication of many viral pathogens. Here, we characterize the relative mRNA expression of the chicken IFITM genes in response to IBV infection, in vivo, ex vivo and in vitro using the pathogenic M41-CK strain, the nephropathogenic QX strain and the nonpathogenic Beaudette strain. In vivo we demonstrate a significant upregulation of chIFITM1, 2, 3 and 5 in M41-CK- and QX-infected trachea two days post-infection. In vitro infection with Beaudette, M41-CK and QX results in a significant upregulation of chIFITM1, 2 and 3 at 24 h post-infection. We confirmed a differential innate response following infection with distinct IBV strains and believe that our data provide new insights into the possible role of chIFITMs in early IBV infection.


Assuntos
Galinhas/genética , Galinhas/virologia , Infecções por Coronavirus/veterinária , Interações Hospedeiro-Patógeno/genética , Proteínas de Membrana/genética , Animais , Infecções por Coronavirus/genética , Regulação Viral da Expressão Gênica , Interações Hospedeiro-Patógeno/fisiologia , Vírus da Bronquite Infecciosa/patogenicidade , Vírus da Bronquite Infecciosa/fisiologia , Técnicas de Cultura de Órgãos , Doenças das Aves Domésticas/etiologia , Doenças das Aves Domésticas/genética , Doenças das Aves Domésticas/virologia , Carga Viral , Tropismo Viral
14.
Virology ; 548: 73-81, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32838948

RESUMO

The host protein SERINC5 inhibits the infectivity of HIV-1 virions in an Env-dependent manner and is counteracted by Nef. The conformation of the Env trimer reportedly correlates with sensitivity to SERINC5. Here, we tested the hypothesis that the "open" conformation of the Env trimer revealed by sensitivity to the V3-loop specific antibody 447-52D directly correlates with sensitivity to SERINC5. Of five Envs tested, SF162 was the most sensitive to neutralization by 447-52D, but it was not the most sensitive to SERINC5; instead the Env of LAI was substantially more sensitive to SERINC5 than all the other Envs. Mutational opening of the trimer by substitution of two tyrosines that mediate interaction between the V2 and V3 loops sensitized the Envs of JRFL and LAI to 447-52D as previously reported, but only BaL was sensitized to SERINC5. These data suggest that trimer "openness" is not sufficient for sensitivity to SERINC5.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Proteínas de Membrana/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/química , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Anticorpos Neutralizantes/imunologia , Infecções por HIV/genética , Infecções por HIV/virologia , HIV-1/química , HIV-1/genética , HIV-1/fisiologia , Humanos , Proteínas de Membrana/química , Proteínas de Membrana/genética , Mutação , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética
15.
Virology ; 548: 82-92, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32838949

RESUMO

Japanese encephalitis virus (JEV) is an infectious pathogen spreading in a wide range of vertebrate species. Pigs are amplifying hosts of JEV and thought to be maintained in nature predominantly by avian-mosquito cycles. In the innate immune system, interferon-inducible transmembrane protein (IFITM) is a small transmembrane protein family and has been identified as the first line of defense against a broad range of RNA virus invasion. In this paper, we found that swine IFITM (sIFITM) could restrict the replication of both JEV vaccine strain and wild strain NJ-2008. The cysteine S-palmitoylation modification of sIFITM plays important roles in their anti-JEV effects and intracellular distributions. Our findings show the anti-JEV activities of swine interferon-inducible transmembrane proteins and broaden the antiviral spectrum of IFITM protein family. The preliminary exploration of S-palmitoylation modification of sIFITM may contribute to understanding of the antiviral molecular mechanism of sIFITM.


Assuntos
Vírus da Encefalite Japonesa (Espécie)/fisiologia , Encefalite Japonesa/virologia , Proteínas de Membrana/imunologia , Animais , Vírus da Encefalite Japonesa (Espécie)/genética , Lipoilação , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Família Multigênica , Suínos , Replicação Viral
16.
Ecotoxicol Environ Saf ; 204: 111060, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32768747

RESUMO

OBJECTIVE: Serratia sp. S2 is a wild strain with chromium resistance and reduction ability. Chromium(VI) metabolic-protein-coding gene ChrA and ChrT were cloned from Serratia sp. S2, and ligated with prokaryotic expression vectors pET-28a (+) and transformed into E. coli BL21 to construct ChrA, ChrT and ChrAT engineered bacteria. By studying the characteristics of Cr(VI) metabolism in engineered bacteria, the function and mechanism of the sole expression and coexpression of ChrA and ChrT genes were studied. METHODS: Using Serratia sp. S2 genome as template, ChrA and ChrT genes were amplified by PCR, and prokaryotic expression vectors was ligated to form the recombinant plasmid pET-28a (+)-ChrA, pET-28a (+)-ChrT and pET-28a (+)-ChrAT, and transformed into E. coli BL21 to construct ChrA, ChrT, ChrAT engineered bacteria. The growth curve, tolerance, and reduction of Cr(VI), the distribution of intracellular and extracellular Cr, activity of chromium reductase and intracellular oxidative stress in engineered bacteria were measured to explore the metabolic characteristics of Cr(VI) in ChrA, ChrT, ChrAT engineered bacteria. RESULTS: ChrA, ChrT and ChrAT engineered bacteria were successfully constructed by gene recombination technology. The tolerance to Cr(VI) was Serratia sp. S2 > ChrAT ≈ ChrA > ChrT > Control (P < 0.05), and the reduction ability to Cr(VI) was Serratia sp. S2 > ChrAT ≈ ChrT > ChrA (P < 0.05). The chromium distribution experiments confirmed that Cr(VI) and Cr(III) were the main valence states. Effect of electron donors on chromium reductase activity was NADPH > NADH > non-NAD(P)H (P < 0.05). The activity of chromium reductase increased significantly with NAD(P)H (P < 0.05). The Glutathione and NPSH (Non-protein Sulfhydryl) levels of ChrA, ChrAT engineered bacteria increased significantly (P < 0.05) under the condition of Cr(VI), but there was no significant difference in the indexes of ChrT engineered bacteria (P > 0.05). CONCLUSION: ChrAT engineered bacteria possesses resistance and reduction abilities of Cr(VI). ChrA protein endows the strain with the ability to resist Cr(VI). ChrT protein reduces Cr(VI) to Cr(III) by using NAD(P)H as electronic donor. The reduction process promotes the production of GSH, GSSG and NPSH to maintain the intracellular reduction state, which further improves the Cr(VI) tolerance and reduction ability of ChrAT engineered bacteria.


Assuntos
Proteínas de Bactérias/genética , Compostos de Cromo/metabolismo , Poluentes Ambientais/metabolismo , Genes Bacterianos , Proteínas de Membrana/genética , Microrganismos Geneticamente Modificados/genética , Serratia/genética , Biodegradação Ambiental , Escherichia coli/genética , Escherichia coli/metabolismo , Microrganismos Geneticamente Modificados/metabolismo , Modelos Teóricos , Oxirredução , Oxirredutases/metabolismo , Serratia/metabolismo
17.
Nat Commun ; 11(1): 4012, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32782388

RESUMO

Transmembrane B cell lymphoma 2-associated X protein inhibitor motif-containing (TMBIM) 6, a Ca2+ channel-like protein, is highly up-regulated in several cancer types. Here, we show that TMBIM6 is closely associated with survival in patients with cervical, breast, lung, and prostate cancer. TMBIM6 deletion or knockdown suppresses primary tumor growth. Further, mTORC2 activation is up-regulated by TMBIM6 and stimulates glycolysis, protein synthesis, and the expression of lipid synthesis genes and glycosylated proteins. Moreover, ER-leaky Ca2+ from TMBIM6, a unique characteristic, is shown to affect mTORC2 assembly and its association with ribosomes. In addition, we identify that the BIA compound, a potentialTMBIM6 antagonist, prevents TMBIM6 binding to mTORC2, decreases mTORC2 activity, and also regulates TMBIM6-leaky Ca2+, further suppressing tumor formation and progression in cancer xenograft models. This previously unknown signaling cascade in which mTORC2 activity is enhanced via the interaction with TMBIM6 provides potential therapeutic targets for various malignancies.


Assuntos
Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Indenos/farmacologia , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Proteínas de Membrana/antagonistas & inibidores , Neoplasias/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Cálcio/metabolismo , Linhagem Celular Tumoral , Transformação Celular Neoplásica/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Neoplasias/genética , Neoplasias/patologia , Ligação Proteica , Ribossomos/metabolismo , Transdução de Sinais , Análise de Sobrevida , Ensaios Antitumorais Modelo de Xenoenxerto , Peixe-Zebra
18.
Nature ; 584(7820): 252-256, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32760004

RESUMO

A fundamental challenge in developing treatments for autism spectrum disorders is the heterogeneity of the condition. More than one hundred genetic mutations confer high risk for autism, with each individual mutation accounting for only a small fraction of cases1-3. Subsets of risk genes can be grouped into functionally related pathways, most prominently those involving synaptic proteins, translational regulation, and chromatin modifications. To attempt to minimize this genetic complexity, recent therapeutic strategies have focused on the neuropeptides oxytocin and vasopressin4-6, which regulate aspects of social behaviour in mammals7. However, it is unclear whether genetic risk factors predispose individuals to autism as a result of modifications to oxytocinergic signalling. Here we report that an autism-associated mutation in the synaptic adhesion molecule Nlgn3 results in impaired oxytocin signalling in dopaminergic neurons and in altered behavioural responses to social novelty tests in mice. Notably, loss of Nlgn3 is accompanied by a disruption of translation homeostasis in the ventral tegmental area. Treatment of Nlgn3-knockout mice with a new, highly specific, brain-penetrant inhibitor of MAP kinase-interacting kinases resets the translation of mRNA and restores oxytocin signalling and social novelty responses. Thus, this work identifies a convergence between the genetic autism risk factor Nlgn3, regulation of translation, and oxytocinergic signalling. Focusing on such common core plasticity elements might provide a pragmatic approach to overcoming the heterogeneity of autism. Ultimately, this would enable mechanism-based stratification of patient populations to increase the success of therapeutic interventions.


Assuntos
Transtorno Autístico/metabolismo , Transtorno Autístico/psicologia , Modelos Animais de Doenças , Ocitocina/metabolismo , Comportamento Social , Animais , Moléculas de Adesão Celular Neuronais/deficiência , Moléculas de Adesão Celular Neuronais/genética , Fator de Iniciação 4E em Eucariotos/metabolismo , Masculino , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/genética , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fosforilação/efeitos dos fármacos , Biossíntese de Proteínas/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais/efeitos dos fármacos , Área Tegmentar Ventral/citologia , Área Tegmentar Ventral/efeitos dos fármacos
19.
Gene ; 761: 145043, 2020 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-32777530

RESUMO

Tonoplast Intrinsic Proteins (TIPs) constitute a significant class of the aquaporins. The TIPs control water trade among cytosolic and vacuolar compartments and can also transport glycerol, ammonia, urea, hydrogen peroxide, metals/metalloids, and so forth. Additionally, TIPs are engaged with different abiotic stress responses and developmental processes like leaf expansion, root elongation and seed germination. In this study, ten TIP genes in the rice genome were identified from Oryza sativa ssp indica. Among these, representative groups of TIP genes were cloned and sequenced whilst some TIP sequences showed stop codons in the coding region. The secondary structure analysis represented six conserved transmembrane helices along with the inter-helical regions having conserved motifs. The representative three-dimensional tetrameric design of protein sequence of TIP1;1 displayed key features like NPA motifs, aromatic/arginine (ar/R) selectivity filters, and Froger's residues. The vacuolar localization, transmembrane topological properties, and conserved motif analysis of the cloned genes altogether supported their identity as TIPs. An unrooted phylogenetic tree delineated the relatedness of TIPs from Oryza with different species and bunched them into five clades. The promoter analysis uncovered key regulons associated with administering abiotic stress responses. Gene expression studies showed thatTIPsare differentially regulated under salt and drought stress at various time points in shoots and roots of rice. Also, the pattern of expression was found to be significantly variable in five different rice tissues. The heat-map based tissue and stress- specific expression analysis supported the experimental findings. In conclusion, the identification and transcript-level expression studies of TIPs significantly contribute towards the comprehension of their utilitarian significance in the abiotic stress response.


Assuntos
Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Oryza/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Aquaporinas/genética , Aquaporinas/metabolismo , Clonagem Molecular/métodos , Secas , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas/genética , Genes de Plantas/genética , Oryza/metabolismo , Filogenia , Folhas de Planta/metabolismo , Cloreto de Sódio/metabolismo , Estresse Fisiológico/genética , Vacúolos/genética , Água/metabolismo
20.
PLoS Genet ; 16(8): e1008975, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32750056

RESUMO

The C. elegans proteins PAQR-2 (a homolog of the human seven-transmembrane domain AdipoR1 and AdipoR2 proteins) and IGLR-2 (a homolog of the mammalian LRIG proteins characterized by a single transmembrane domain and the presence of immunoglobulin domains and leucine-rich repeats in their extracellular portion) form a complex that protects against plasma membrane rigidification by promoting the expression of fatty acid desaturases and the incorporation of polyunsaturated fatty acids into phospholipids, hence increasing membrane fluidity. In the present study, we leveraged a novel gain-of-function allele of PAQR-1, a PAQR-2 paralog, to carry out structure-function studies. We found that the transmembrane domains of PAQR-2 are responsible for its functional requirement for IGLR-2, that PAQR-1 does not require IGLR-2 but acts via the same pathway as PAQR-2, and that the divergent N-terminal cytoplasmic domains of the PAQR-1 and PAQR-2 proteins serve a regulatory function and may regulate access to the catalytic site of these proteins. We also show that overexpression of human AdipoR1 or AdipoR2 alone is sufficient to confer increased palmitic acid resistance in HEK293 cells, and thus act in a manner analogous to the PAQR-1 gain-of-function allele.


Assuntos
Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Proteínas de Membrana/genética , Receptores de Adiponectina/genética , Alelos , Animais , Caenorhabditis elegans/metabolismo , Membrana Celular/genética , Membrana Celular/metabolismo , Mutação com Ganho de Função/genética , Células HEK293 , Humanos , Fluidez de Membrana/genética , Fenótipo , Fosfolipídeos/genética , Fosfolipídeos/metabolismo
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