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2.
Nat Commun ; 11(1): 4510, 2020 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-32908143

RESUMO

With human median lifespan extending into the 80s in many developed countries, the societal burden of age-related muscle loss (sarcopenia) is increasing. mTORC1 promotes skeletal muscle hypertrophy, but also drives organismal aging. Here, we address the question of whether mTORC1 activation or suppression is beneficial for skeletal muscle aging. We demonstrate that chronic mTORC1 inhibition with rapamycin is overwhelmingly, but not entirely, positive for aging mouse skeletal muscle, while genetic, muscle fiber-specific activation of mTORC1 is sufficient to induce molecular signatures of sarcopenia. Through integration of comprehensive physiological and extensive gene expression profiling in young and old mice, and following genetic activation or pharmacological inhibition of mTORC1, we establish the phenotypically-backed, mTORC1-focused, multi-muscle gene expression atlas, SarcoAtlas (https://sarcoatlas.scicore.unibas.ch/), as a user-friendly gene discovery tool. We uncover inter-muscle divergence in the primary drivers of sarcopenia and identify the neuromuscular junction as a focal point of mTORC1-driven muscle aging.


Assuntos
Envelhecimento/fisiologia , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Fibras Musculares Esqueléticas/patologia , Junção Neuromuscular/patologia , Sarcopenia/patologia , Envelhecimento/efeitos dos fármacos , Animais , Linhagem Celular , Modelos Animais de Doenças , Eletromiografia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Humanos , Microdissecção e Captura a Laser , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina/antagonistas & inibidores , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Camundongos , Mioblastos , Junção Neuromuscular/efeitos dos fármacos , Técnicas de Patch-Clamp , RNA-Seq , Sarcopenia/genética , Sarcopenia/fisiopatologia , Sarcopenia/prevenção & controle , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Sirolimo/administração & dosagem
3.
Vet Microbiol ; 247: 108793, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32768236

RESUMO

Porcine epidemic diarrhea virus (PEDV) belongs to the Alphacoronavirus genus in the Coronaviridae family. Similar to other coronaviruses, PEDV encodes two papain-like proteases. Papain-like protease (PLP)2 has been proposed to play a key role in antagonizing host innate immunity. However, the function of PLP1 remains unclear. In this study, we found that overexpression of PLP1 significantly promoted PEDV replication and inhibited production of interferon-ß. Immunoprecipitation and mass spectrometry were used to identify cellular interaction partners of PLP1. Host cell poly(C) binding protein 2 (PCBP2) was determined to bind and interact with PLP1. Both endogenous and overexpressed PCBP2 co-localized with PLP1 in the cytoplasm. Overexpression of PLP1 upregulated expression of PCBP2. Furthermore, overexpression of PCBP2 promoted PEDV replication. Silencing of endogenous PCBP2 using small interfering RNAs attenuated PEDV replication. Taken together, these data demonstrated that PLP1 negatively regulated the production of type 1 interferon by interacting with PCBP2 and promoted PEDV replication.


Assuntos
Papaína/metabolismo , Vírus da Diarreia Epidêmica Suína/metabolismo , Proteínas não Estruturais Virais/metabolismo , Replicação Viral/fisiologia , Animais , Chlorocebus aethiops , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Células HEK293 , Humanos , Interferon beta/genética , Interferon beta/metabolismo , Proteína Proteolipídica de Mielina/metabolismo , Papaína/genética , Vírus da Diarreia Epidêmica Suína/genética , Vírus da Diarreia Epidêmica Suína/fisiologia , Interferência de RNA , Proteínas de Ligação a RNA , Fator de Necrose Tumoral alfa/farmacologia , Células Vero , Proteínas não Estruturais Virais/genética
4.
Nat Commun ; 11(1): 4313, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32855402

RESUMO

It has been suggested that beige fat thermogenesis is tightly controlled by epigenetic regulators that sense environmental cues such as temperature. Here, we report that subcutaneous adipose expression of the DNA demethylase TET1 is suppressed by cold and other stimulators of beige adipocyte thermogenesis. TET1 acts as an autonomous repressor of key thermogenic genes, including Ucp1 and Ppargc1a, in beige adipocytes. Adipose-selective Tet1 knockout mice generated by using Fabp4-Cre improves cold tolerance and increases energy expenditure and protects against diet-induced obesity and insulin resistance. Moreover, the suppressive role of TET1 in the thermogenic gene regulation of beige adipocytes is largely DNA demethylase-independent. Rather, TET1 coordinates with HDAC1 to mediate the epigenetic changes to suppress thermogenic gene transcription. Taken together, TET1 is a potent beige-selective epigenetic breaker of the thermogenic gene program. Our findings may lead to a therapeutic strategy to increase energy expenditure in obesity and related metabolic disorders.


Assuntos
Adipócitos Bege/metabolismo , Proteínas de Ligação a DNA/metabolismo , Epigênese Genética , Obesidade/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Termogênese/genética , Animais , Calorimetria Indireta , Linhagem Celular , Temperatura Baixa/efeitos adversos , Proteínas de Ligação a DNA/genética , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Metabolismo Energético/genética , Regulação da Expressão Gênica/fisiologia , Células HEK293 , Humanos , Resistência à Insulina/genética , Masculino , Camundongos , Camundongos Knockout , Obesidade/etiologia , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Proteínas Proto-Oncogênicas/genética , RNA-Seq , Gordura Subcutânea/citologia , Gordura Subcutânea/metabolismo , Proteína Desacopladora 1/metabolismo
5.
PLoS Negl Trop Dis ; 14(8): e0008509, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32804927

RESUMO

Leishmania species are responsible for a broad spectrum of diseases, denominated Leishmaniasis, affecting over 12 million people worldwide. During the last decade, there have been impressive efforts for sequencing the genome of most of the pathogenic Leishmania spp. as well as hundreds of strains, but large-scale proteomics analyses did not follow these achievements and the Leishmania proteome remained mostly uncharacterized. Here, we report a comprehensive comparative study of the proteomes of strains representing L. braziliensis, L. panamensis and L. guyanensis species. Proteins extracted by SDS-mediated lysis were processed following the multi-enzyme digestion-filter aided sample preparation (FASP) procedure and analysed by high accuracy mass spectrometry. "Total Protein Approach" and "Proteomic Ruler" were applied for absolute quantification of proteins. Principal component analysis demonstrated very high reproducibility among biological replicates and a very clear differentiation of the three species. Our dataset comprises near 7000 proteins, representing the most complete Leishmania proteome yet known, and provides a comprehensive quantitative picture of the proteomes of the three species in terms of protein concentration and copy numbers. Analysis of the abundance of proteins from the major energy metabolic processes allow us to highlight remarkably differences among the species and suggest that these parasites depend on distinct energy substrates to obtain ATP. Whereas L. braziliensis relies the more on glycolysis, L. panamensis and L. guyanensis seem to depend mainly on mitochondrial respiration. These results were confirmed by biochemical assays showing opposite profiles for glucose uptake and O2 consumption in these species. In addition, we provide quantitative data about different membrane proteins, transporters, and lipids, all of which contribute for significant species-specific differences and provide rich substrate for explore new molecules for diagnosing purposes. Data are available via ProteomeXchange with identifier PXD017696.


Assuntos
Leishmania/metabolismo , Proteínas de Protozoários/metabolismo , Regulação da Expressão Gênica/fisiologia , Glucose/metabolismo , Leishmania/genética , Consumo de Oxigênio , Proteômica , Proteínas de Protozoários/genética , Especificidade da Espécie
6.
Proc Natl Acad Sci U S A ; 117(33): 20149-20158, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32747560

RESUMO

The C2 domain containing protein extended synaptotagmin (E-Syt) plays important roles in both lipid homeostasis and the intracellular signaling; however, its role in physiology remains largely unknown. Here, we show that hypothalamic E-Syt3 plays a critical role in diet-induced obesity (DIO). E-Syt3 is characteristically expressed in the hypothalamic nuclei. Whole-body or proopiomelanocortin (POMC) neuron-specific ablation of E-Syt3 ameliorated DIO and related comorbidities, including glucose intolerance and dyslipidemia. Conversely, overexpression of E-Syt3 in the arcuate nucleus moderately promoted food intake and impaired energy expenditure, leading to increased weight gain. Mechanistically, E-Syt3 ablation led to increased processing of POMC to α-melanocyte-stimulating hormone (α-MSH), increased activities of protein kinase C and activator protein-1, and enhanced expression of prohormone convertases. These findings reveal a previously unappreciated role for hypothalamic E-Syt3 in DIO and related metabolic disorders.


Assuntos
Regulação da Expressão Gênica/fisiologia , Obesidade/induzido quimicamente , Obesidade/genética , Sinaptotagminas/metabolismo , Animais , Dieta Hiperlipídica/efeitos adversos , Predisposição Genética para Doença , Hipotálamo/metabolismo , Masculino , Camundongos , Camundongos Knockout , Neurônios/metabolismo , Pró-Proteína Convertase 1/genética , Pró-Proteína Convertase 1/metabolismo , Pró-Proteína Convertase 2/genética , Pró-Proteína Convertase 2/metabolismo , Sinaptotagminas/genética
7.
Nat Commun ; 11(1): 3520, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32665551

RESUMO

PRDM (PRDI-BF1 and RIZ homology domain containing) family members are sequence-specific transcriptional regulators involved in cell identity and fate determination, often dysregulated in cancer. The PRDM15 gene is of particular interest, given its low expression in adult tissues and its overexpression in B-cell lymphomas. Despite its well characterized role in stem cell biology and during early development, the role of PRDM15 in cancer remains obscure. Herein, we demonstrate that while PRDM15 is largely dispensable for mouse adult somatic cell homeostasis in vivo, it plays a critical role in B-cell lymphomagenesis. Mechanistically, PRDM15 regulates a transcriptional program that sustains the activity of the PI3K/AKT/mTOR pathway and glycolysis in B-cell lymphomas. Abrogation of PRDM15 induces a metabolic crisis and selective death of lymphoma cells. Collectively, our data demonstrate that PRDM15 fuels the metabolic requirement of B-cell lymphomas and validate it as an attractive and previously unrecognized target in oncology.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição/metabolismo , Animais , Apoptose/genética , Apoptose/fisiologia , Western Blotting , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Imunoprecipitação da Cromatina , Biologia Computacional , Proteínas de Ligação a DNA/genética , Feminino , Citometria de Fluxo , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/fisiologia , Humanos , Linfoma/genética , Linfoma/metabolismo , Camundongos , Camundongos SCID , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Distribuição Aleatória , Fatores de Transcrição/genética , Transcriptoma/genética
8.
PLoS Negl Trop Dis ; 14(7): e0008439, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32628683

RESUMO

Leishmaniasis constitutes the 9th largest disease burden among all infectious diseases. Control of this disease is based on a short list of chemotherapeutic agents headed by pentavalent antimonials, followed by miltefosine and amphotericin B; drugs that are far from ideal due to host toxicity, elevated cost, limited access, and high rates of drug resistance. Knowing that the composition of extracellular vesicles (EVs) can vary according to the state of their parental cell, we hypothesized that EVs released by drug-resistant Leishmania infantum parasites could contain unique and differently enriched proteins depending on the drug-resistance mechanisms involved in the survival of their parental cell line. To assess this possibility, we studied EV production, size, morphology, and protein content of three well-characterized drug-resistant L. infantum cell lines and a wild-type strain. Our results are the first to demonstrate that drug-resistance mechanisms can induce changes in the morphology, size, and distribution of L. infantum EVs. In addition, we identified L. infantum's core EV proteome. This proteome is highly conserved among strains, with the exception of a handful of proteins that are enriched differently depending on the drug responsible for induction of antimicrobial resistance. Furthermore, we obtained the first snapshot of proteins enriched in EVs released by antimony-, miltefosine- and amphotericin-resistant parasites. These include several virulence factors, transcription factors, as well as proteins encoded by drug-resistance genes. This detailed study of L. infantum EVs sheds new light on the potential roles of EVs in Leishmania biology, particularly with respect to the parasite's survival in stressful conditions. This work outlines a crucial first step towards the discovery of EV-based profiles capable of predicting response to antileishmanial agents.


Assuntos
Antiprotozoários/farmacologia , Resistência a Medicamentos , Regulação da Expressão Gênica/efeitos dos fármacos , Leishmania infantum/efeitos dos fármacos , Leishmania infantum/metabolismo , Biologia Computacional , Vesículas Extracelulares , Regulação da Expressão Gênica/fisiologia , Proteoma , Proteômica , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo
9.
Mol Immunol ; 125: 9-14, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32619931

RESUMO

MiR-483-3p is involved in the pathogenesis of acute myocardial infarctions, but its association with myocardial ischemia reperfusion (IR) remains mostly unknown. In this study, an in vitro model of myocardial IR injury was established by putting H9c2 cells into hypoxia reoxygenation (HR) treatment to explore the effects and possible mechanisms of miR-483-3p in myocardial IR injury. HR exposure resulted in increased miR-483-3p levels in H9c2 cells. MiR-483-3p was overexpressed or downregulated in H9c2 cells by transfection of miR-483-3p mimic or miR-483-3p inhibitor. In HR-treated H9c2 cells, MiR-483-3p mimics inhibited cell viability, promoted lactate dehydrogenase release, and increased apoptosis, but miR-483-3p inhibitors caused the opposite effects. MDM4 was verified to be the target mRNA of miR-483-3p and negatively modulated by miR-483-3p. MiR-483-3p inhibitor upregulated MDM4 and Bcl-2, but downregulated p53 and Bax in HR-treated H9c2 cells, whereas miR-483-3p overexpression produced the opposite effects.. Moreover, MDM4 siRNA transfection partially reversed the role of miR-483-3p inhibition in HR injury and p53 pathway inactivation of H9c2 cells. In summary, by targeting the MDM4/p53 pathway, miR-483-3p inhibition may alleviate myocardial HR injury. MiR-483-3p may be a potential therapeutic target of myocardial IR injury.


Assuntos
Regulação da Expressão Gênica/fisiologia , MicroRNAs/metabolismo , Traumatismo por Reperfusão Miocárdica/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Linhagem Celular , Modelos Animais de Doenças , Técnicas In Vitro , Ratos
10.
Proc Natl Acad Sci U S A ; 117(31): 18172-18174, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32690689

RESUMO

The assembly of T cell receptor (TCR) and immunoglobulin (Ig) genes by V(D)J recombination generates the antigen receptor (AgR) diversity that is vital for adaptive immunity. At most AgR loci, V(D)J recombination is regulated so that only one allele assembles a functional gene, ensuring that nearly every T and B cell expresses a single type, or specificity, of AgR. The genomic organizations of some AgR loci permit the assembly and expression of two distinct genes on each allele; however, this is prevented by undetermined mechanisms. We show that the poor qualities of recombination signal sequences (RSSs) flanking Vß gene segments suppress the assembly and expression of two distinct TCRß genes from a single allele. Our data demonstrate that an intrinsic genetic mechanism that stochastically limits Vß recombination efficiency governs monogenic TCRß expression, thereby restraining the expression of multiple AgRs on αß T cells.


Assuntos
Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/fisiologia , Recombinação V(D)J , Animais , Feminino , Regulação da Expressão Gênica/fisiologia , Heterozigoto , Masculino , Camundongos , Linfócitos T
11.
Ocul Immunol Inflamm ; 28(5): 735-738, 2020 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-32589459

RESUMO

PURPOSE: The spike proteins of SARS-CoV-2 interact with ACE2 or basigin/CD147 receptors, regulating human-to-human transmissions of COVID-19 together with serine protease TMPRSS2. The expression of these receptors on the ocular surface is unknown. MATERIAL AND METHODS: Gene expression of SARS-CoV-2 receptors was investigated in conjunctival epithelial cell samples and in ex-vivo cornea samples using microarray or transcriptome sequencing. RESULTS: ACE2 is expressed in conjunctival samples at a low level, while BSG and TMPRSS2 are expressed at intermediate levels in both conjunctiva and cornea. Other receptors such as ANPEP, AGTR2 are expressed at low level in the conjunctiva. Two RNA editing enzymes involved in antiviral responses, APOBEC3A, and ADAR-1 were also highly expressed. CONCLUSIONS: The ocular surface may represent an entry point for the SARS-CoV-2 in the human body. The conjunctiva and the cornea can adopt antiviral countermeasures which may explain the low prevalence of eye involvement.


Assuntos
Betacoronavirus/fisiologia , Túnica Conjuntiva/metabolismo , Córnea/metabolismo , Infecções por Coronavirus/metabolismo , Regulação da Expressão Gênica/fisiologia , Pneumonia Viral/metabolismo , Receptores Virais/genética , Adenosina Desaminase/genética , Adolescente , Adulto , Idoso , Basigina/genética , Criança , Citidina Desaminase/genética , Perfilação da Expressão Gênica , Humanos , Pessoa de Meia-Idade , Pandemias , Peptidil Dipeptidase A/genética , Proteínas/genética , Proteínas de Ligação a RNA/genética , Serina Endopeptidases/genética , Adulto Jovem
12.
Life Sci ; 256: 117921, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32526288

RESUMO

Estrogen is a hormone responsible for modulating several physiological processes such as immune response and bone homeostasis. Physiological fluctuations of estrogen concentration are one of the defining principles behind its mechanism. In cases of estrogen deficiency, such as in menopausal women, a more intense bone resorption may occur due to an increase in osteoclast activity. One of the main factors that influence osteoclast formation and response is the immune system, mainly through cytokines secreted by B and T cells. The purpose of this review is to highlight how estrogen can modulate the secretion of cytokines that can alter bone physiology, thereby establishing an axis between estrogen, immune cells, and osteoclastogenesis.


Assuntos
Linfócitos B/metabolismo , Citocinas/metabolismo , Estrogênios/metabolismo , Osteogênese/fisiologia , Linfócitos T/metabolismo , Animais , Reabsorção Óssea/metabolismo , Osso e Ossos/metabolismo , Diferenciação Celular , Citocinas/genética , Moduladores de Receptor Estrogênico/metabolismo , Feminino , Regulação da Expressão Gênica/fisiologia , Humanos , Menopausa/metabolismo , Osteoclastos/metabolismo
13.
Proc Natl Acad Sci U S A ; 117(26): 15293-15304, 2020 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-32541062

RESUMO

Organisms possess photoperiodic timing mechanisms to detect variations in day length and temperature as the seasons progress. The nature of the molecular mechanisms interpreting and signaling these environmental changes to elicit downstream neuroendocrine and physiological responses are just starting to emerge. Here, we demonstrate that, in Drosophila melanogaster, EYES ABSENT (EYA) acts as a seasonal sensor by interpreting photoperiodic and temperature changes to trigger appropriate physiological responses. We observed that tissue-specific genetic manipulation of eya expression is sufficient to disrupt the ability of flies to sense seasonal cues, thereby altering the extent of female reproductive dormancy. Specifically, we observed that EYA proteins, which peak at night in short photoperiod and accumulate at higher levels in the cold, promote reproductive dormancy in female D. melanogaster Furthermore, we provide evidence indicating that the role of EYA in photoperiodism and temperature sensing is aided by the stabilizing action of the light-sensitive circadian clock protein TIMELESS (TIM). We postulate that increased stability and level of TIM at night under short photoperiod together with the production of cold-induced and light-insensitive TIM isoforms facilitate EYA accumulation in winter conditions. This is supported by our observations that tim null mutants exhibit reduced incidence of reproductive dormancy in simulated winter conditions, while flies overexpressing tim show an increased incidence of reproductive dormancy even in long photoperiod.


Assuntos
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/fisiologia , Proteínas do Olho/metabolismo , Fotoperíodo , Estações do Ano , Temperatura , Animais , Proteínas de Drosophila/genética , Proteínas do Olho/genética , Regulação da Expressão Gênica/fisiologia , Reprodução
14.
PLoS Genet ; 16(6): e1008831, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32555673

RESUMO

Conspecific male animals fight for resources such as food and mating opportunities but typically stop fighting after assessing their relative fighting abilities to avoid serious injuries. Physiologically, how the fighting behavior is controlled remains unknown. Using the fighting fish Betta splendens, we studied behavioral and brain-transcriptomic changes during the fight between the two opponents. At the behavioral level, surface-breathing, and biting/striking occurred only during intervals between mouth-locking. Eventually, the behaviors of the two opponents became synchronized, with each pair showing a unique behavioral pattern. At the physiological level, we examined the expression patterns of 23,306 brain transcripts using RNA-sequencing data from brains of fighting pairs after a 20-min (D20) and a 60-min (D60) fight. The two opponents in each D60 fighting pair showed a strong gene expression correlation, whereas those in D20 fighting pairs showed a weak correlation. Moreover, each fighting pair in the D60 group showed pair-specific gene expression patterns in a grade of membership analysis (GoM) and were grouped as a pair in the heatmap clustering. The observed pair-specific individualization in brain-transcriptomic synchronization (PIBS) suggested that this synchronization provides a physiological basis for the behavioral synchronization. An analysis using the synchronized genes in fighting pairs of the D60 group found genes enriched for ion transport, synaptic function, and learning and memory. Brain-transcriptomic synchronization could be a general phenomenon and may provide a new cornerstone with which to investigate coordinating and sustaining social interactions between two interacting partners of vertebrates.


Assuntos
Comportamento Animal/fisiologia , Encéfalo/fisiologia , Peixes/fisiologia , Regulação da Expressão Gênica/fisiologia , Transcriptoma/fisiologia , Agressão , Animais , Técnicas de Observação do Comportamento , Comportamento Cooperativo , Relações Interpessoais , Transporte de Íons/fisiologia , Aprendizagem/fisiologia , Masculino , Memória/fisiologia , RNA-Seq , Gravação em Vídeo
15.
Life Sci ; 256: 118008, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32593709

RESUMO

AIMS: We investigate the effects of RT on the mechanical function, gene, and protein expression of key factors involved in bone remodeling during aging. MAIN METHODS: Male rats of 3 and 21 months of age were randomly allocated into four groups (8 per group): young sedentary (YS), young trained (YT), old sedentary (OS), and old trained (OT). RT was performed three times per week (12 weeks). Bone tenacity and stiffness were measured by biomechanical tests and mRNA levels of COL1A1, MEPE, SOST, OPG, BMP-2, PPAR-y, MMP-2-9-13, and TIMP-1 were evaluated by quantitative PCR. COL1A1 protein and MMP-2 activity were detected by western blotting and zymography assays. KEY FINDINGS: Aging increased stiffness, while BMP-2, OPG, COL1A1 and MMP-2 mRNA levels reduced (OS vs YS; p ≤ 0.05). RT increased the tenacity of the femur and reduced PPAR-γ regardless of age (YT vs. YS; OT vs. OS; p ≤ 0.05). RT downregulated SOST mRNA levels only in the OT group (vs. OS group, p ≤ 0.05). RT mitigated the age-associated increase in MMP-9 mRNA levels (p ≤ 0.05). In young animals, upregulation in MEPE, MMP-13, TIMP-1 were observed after RT, as well an increase in COL1A1 protein and MMP-2 activity (p ≤ 0.05). SIGNIFICANCE: RT improved bone tenacity independent of aging, which is relevant for mechanical function, while, at protein levels, RT upregulated MMP-2 activity and collagen 1 only in young rats. This study highlights the importance of exercise on bone health and identifies specific molecular changes in response to RT. Our findings provide insights into the mechanisms involved in age-related changes.


Assuntos
Envelhecimento/fisiologia , Remodelação Óssea/fisiologia , Condicionamento Físico Animal/fisiologia , Treinamento de Resistência/métodos , Fatores Etários , Animais , Remodelação Óssea/genética , Regulação da Expressão Gênica/fisiologia , Masculino , RNA Mensageiro/genética , Distribuição Aleatória , Ratos , Ratos Wistar
16.
Nat Commun ; 11(1): 3140, 2020 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-32561780

RESUMO

MeCP2 plays a multifaceted role in gene expression regulation and chromatin organization. Interaction between MeCP2 and methylated DNA in the regulation of gene expression is well established. However, the widespread distribution of MeCP2 suggests it has additional interactions with chromatin. Here we demonstrate, by both biochemical and genomic analyses, that MeCP2 directly interacts with nucleosomes and its genomic distribution correlates with that of H3K27me3. In particular, the methyl-CpG-binding domain of MeCP2 shows preferential interactions with H3K27me3. We further observe that the impact of MeCP2 on transcriptional changes correlates with histone post-translational modification patterns. Our findings indicate that MeCP2 interacts with genomic loci via binding to DNA as well as histones, and that interaction between MeCP2 and histone proteins plays a key role in gene expression regulation.


Assuntos
Regulação da Expressão Gênica/fisiologia , Histonas/metabolismo , Proteína 2 de Ligação a Metil-CpG/metabolismo , Transcrição Genética/fisiologia , Animais , Sequenciamento de Cromatina por Imunoprecipitação , DNA/genética , DNA/metabolismo , DNA (Citosina-5-)-Metiltransferase 1/genética , DNA (Citosina-5-)-Metiltransferases/genética , Metilação de DNA/fisiologia , Técnicas de Inativação de Genes , Loci Gênicos , Células HCT116 , Células HEK293 , Histonas/genética , Humanos , Proteína 2 de Ligação a Metil-CpG/genética , Camundongos , Camundongos Knockout , Nucleossomos/genética , Nucleossomos/metabolismo , Processamento de Proteína Pós-Traducional/fisiologia , Sítio de Iniciação de Transcrição/fisiologia
17.
Mol Immunol ; 124: 172-179, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32585511

RESUMO

The family of TRIM proteins with E3 ubiquitin ligase activity plays important roles in virus infection in vertebrates and invertebrates. In this study, a novel Trim gene shows high similarity to Trim23 (designated as MnTrim23) was identified from Macrobrachium nipponense. The MnTrim23 protein contains three conserved domains (one RING finger domain, two B-box, and one Coiled-coil region) at its N-terminal and one ARF domain at its C-terminal. The ARF domain characterizes the members of the Trim23 family. MnTrim23 belongs to C-IX family. Phylogenetic analysis shows that MnTrim23 has a closer genetic distance with other Trim23 proteins from invertebrates than that from vertebrates. MnTrim23 has higher expression level in the intestine and hepatopancreas than in the other immune tissues. The expression levels of MnTrim23 in the gills, stomach, and intestines are significantly up-regulated after white spot syndrome virus (WSSV) infection. Moreover, knockdown of MnTrim23 inhibits WSSV replication and VP28 expression, suggesting that MnTrim23 plays a positive role in WSSV infection. Further studies revealed that MnTrim23 negatively regulates the Relish transcription factor-mediated expression of antimicrobial peptides (AMPs). Synthetic AMPs inhibit VP28 expression and WSSV replication. These findings indicate that Trim23 promotes WSSV replication by inhibiting the expression of AMPs that are positively regulated by the host NF-κB signal pathway.


Assuntos
Peptídeos Catiônicos Antimicrobianos/biossíntese , Interações entre Hospedeiro e Microrganismos/fisiologia , Penaeidae/virologia , Proteínas com Motivo Tripartido/metabolismo , Vírus da Síndrome da Mancha Branca 1/fisiologia , Animais , Peptídeos Catiônicos Antimicrobianos/imunologia , Regulação da Expressão Gênica/fisiologia , Penaeidae/imunologia , Penaeidae/metabolismo , Proteínas com Motivo Tripartido/imunologia , Replicação Viral/fisiologia
18.
Proc Natl Acad Sci U S A ; 117(27): 15581-15590, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32576685

RESUMO

Protein synthesis represents a major metabolic activity of the cell. However, how it is affected by aging and how this in turn impacts cell function remains largely unexplored. To address this question, herein we characterized age-related changes in both the transcriptome and translatome of mouse tissues over the entire life span. We showed that the transcriptome changes govern those in the translatome and are associated with altered expression of genes involved in inflammation, extracellular matrix, and lipid metabolism. We also identified genes that may serve as candidate biomarkers of aging. At the translational level, we uncovered sustained down-regulation of a set of 5'-terminal oligopyrimidine (5'-TOP) transcripts encoding protein synthesis and ribosome biogenesis machinery and regulated by the mTOR pathway. For many of them, ribosome occupancy dropped twofold or even more. Moreover, with age, ribosome coverage gradually decreased in the vicinity of start codons and increased near stop codons, revealing complex age-related changes in the translation process. Taken together, our results reveal systematic and multidimensional deregulation of protein synthesis, showing how this major cellular process declines with age.


Assuntos
Envelhecimento/fisiologia , Regulação da Expressão Gênica/fisiologia , Biossíntese de Proteínas/fisiologia , Ribossomos/metabolismo , Animais , Códon de Iniciação/metabolismo , Biologia Computacional , Masculino , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA-Seq , Ribossomos/genética , Transdução de Sinais/fisiologia , Serina-Treonina Quinases TOR/metabolismo , Transcriptoma/fisiologia
19.
Invest Ophthalmol Vis Sci ; 61(5): 26, 2020 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-32416603

RESUMO

Purpose: IFN-stimulated gene (ISG) 15 is a type 1 IFN-induced protein and known to modify target proteins in a manner similar to ubiquitylation (protein conjugation by ISG15 is termed ISGylation). We sought to determine the role of ISG15 and its underlying mechanisms in corneal innate immune defense against Pseudomonas aeruginosa keratitis. Methods: ISG15 expression in cultured human corneal epithelial cells (HCECs) and mouse corneas was determined by PCR and Western blot analysis. Gene knockout mice were used to define the role of ISG15 signaling in controlling the severity of P. aeruginosa keratitis, which was assessed with photographing, clinical scoring, bacterial counting, myeloperoxidase assay, and quantitative PCR determination of cytokine expression. Integrin LFA-1 inhibitor was used to assess its involvement of ISG15 signaling in P. aeruginosa-infected corneas. Results: Heat-killed P. aeruginosa induced ISG15 expression in cultured HCECs and accumulation in the conditioned media. Isg15 deficiency accelerated keratitis progress, suppressed IFNγ and CXCL10, and promoted IL-1ß while exhibiting no effects on IFNα expression. Moreover, exogenous ISG15 protected the corneas of wild-type mice from P. aeruginosa infection while markedly reducing the severity of P. aeruginosa keratitis in type 1 IFN-receptor knockout mice. Exogenous ISG15 increased bacteriostatic activity of B6 mouse corneal homogenates, and inhibition of LFA-1 exacerbated the severity of and abolished protective effects of ISG15 on P. aeruginosa keratitis. Conclusions: Type 1 INF-induced ISG15 regulates the innate immune response and greatly reduces the susceptibility of B6 mouse corneas to P. aeruginosa infection in an LFA-1-dependent manner.


Assuntos
Úlcera da Córnea/imunologia , Citocinas/fisiologia , Infecções Oculares Bacterianas/imunologia , Imunidade Inata/fisiologia , Infecções por Pseudomonas/imunologia , Ubiquitinas/fisiologia , Animais , Carga Bacteriana , Western Blotting , Células Cultivadas , Úlcera da Córnea/metabolismo , Úlcera da Córnea/fisiopatologia , Citocinas/metabolismo , Epitélio Anterior/metabolismo , Infecções Oculares Bacterianas/metabolismo , Infecções Oculares Bacterianas/fisiopatologia , Regulação da Expressão Gênica/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Peroxidase/metabolismo , Infecções por Pseudomonas/metabolismo , Infecções por Pseudomonas/fisiopatologia , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/fisiologia
20.
Invest Ophthalmol Vis Sci ; 61(5): 28, 2020 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-32421148

RESUMO

Purpose: Bestrophinopathies are a group of untreatable inherited retinal dystrophies caused by mutations in the retinal pigment epithelium (RPE) Cl- channel bestrophin 1. We tested whether sodium phenylbutyrate (4PBA) could rescue the function of mutant bestrophin 1 associated with autosomal dominant and recessive disease. We then sought analogues of 4PBA with increased potency and determined the mode of action for 4PBA and a lead compound 2-naphthoxyacetic acid (2-NOAA). Lastly, we tested if 4PBA and 2-NOAA could functionally rescue bestrophin 1 function in RPE generated from induced pluripotent stem cells (iPSC-RPEs) derived from patients with a dominant or recessive bestrophinopathy. Methods: Global and plasma membrane expression was determined by Western blot and immunofluorescent microscopy, respectively. The effect of 4PBA and 2-NOAA on transcription was measured by quantitative RT-PCR and the rate of protein turnover by cycloheximide chase and Western blot. Channel function was measured by whole-cell patch clamp. Results: 4PBA and 2-NOAA can rescue the global and membrane expression of mutant bestrophin 1 associated with autosomal dominant disease (Best vitelliform macular dystrophy [BVMD]) and autosome recessive bestrophinopathy (ARB), and these small molecules have different modes of action. Both 4PBA and 2-NOAA significantly increased the channel function of mutant BVMD and ARB bestrophin 1 in HEK293T and iPSC-RPE cells derived from patients with BVMD and ARB. For 4PBA, the increased mutant channel function in BVMD and ARB iPSC-RPE was equal to that of wild-type iPSC-RPE bestrophin 1. Conclusions: The restoration of bestrophin 1 function in patient-derived RPE confirms the US Food and Drug Administration-approved drug 4PBA as a promising therapeutic treatment for bestrophinopathies.


Assuntos
Antineoplásicos/farmacologia , Bestrofinas/genética , Oftalmopatias Hereditárias/tratamento farmacológico , Regulação da Expressão Gênica/fisiologia , Glicolatos/farmacologia , Fenilbutiratos/farmacologia , Doenças Retinianas/tratamento farmacológico , Epitélio Pigmentado da Retina/efeitos dos fármacos , Western Blotting , Membrana Celular/metabolismo , Canais de Cloreto/metabolismo , Cicloeximida/farmacologia , Eletroforese em Gel de Poliacrilamida , Oftalmopatias Hereditárias/genética , Oftalmopatias Hereditárias/metabolismo , Genes Recessivos , Células HEK293/efeitos dos fármacos , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Microscopia de Fluorescência , Técnicas de Patch-Clamp , Reação em Cadeia da Polimerase em Tempo Real , Doenças Retinianas/genética , Doenças Retinianas/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Transfecção
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