RESUMO
Extracellular vesicles (EVs) have emerged as promising biomaterials for the treatment of different disease. However, only handful types of EVs with clinical transformation potential have been reported to date, and their preparation on a large scale under biosafety-controlled conditions is limited. In this study, we characterize a novel type of EV with promising clinical application potential: dehydration-induced extracellular vesicles (DIMVs). DIMV is a type of micron-diameter cell vesicle that contains more bioactive molecules, such as proteins and RNA, but not DNA, than previously reported cell vesicles. The preparation of DIMV is extraordinarily straightforward, which possesses a high level of biosafety, and the protein utilization ratio is roughly 600 times greater than that of naturally secreted EVs. Additional experiments demonstrate the viability of pre- or post-isolation DIMV modification, including gene editing, nucleic acid encapsulation or surface anchoring, size adjustment. Finally, on animal models, we directly show the biosafety and immunogenicity of DIMV, and investigate its potential application as tumour vaccine or drug carrier in cancer treatment.
Assuntos
Vesículas Extracelulares , Vesículas Extracelulares/metabolismo , Animais , Humanos , Camundongos , Desidratação/metabolismo , Vacinas AnticâncerRESUMO
Defective hydration of airway surface mucosa is associated with lung infection in cystic fibrosis (CF), partly caused by disruption of the epithelial barrier integrity. Although rehydration of the CF airway surface liquid (ASL) alleviates epithelium vulnerability to infection by junctional protein expression, the mechanisms linking ASL to barrier integrity are unknown. We show here the strong degradation of YAP1 and TAZ proteins in well-polarized CF human airway epithelial cells (HAECs), a process that was prevented by ASL rehydration. Conditional silencing of YAP1 in rehydrated CF HAECs indicated that YAP1 expression was necessary for the maintenance of junctional complexes. A higher plasma membrane tension in CF HAECs reduced endocytosis, concurrent with the maintenance of active ß1-integrin ectopically located at the apical membrane. Pharmacological inhibition of ß1-integrin accumulation restored YAP1 expression in CF HAECs. These results indicate that dehydration of the CF ASL affects epithelial plasma membrane tension, resulting in ectopic activation of a ß1-integrin/YAP1 signaling pathway associated with degradation of junctional proteins.
Assuntos
Fibrose Cística , Epitélio , Transdução de Sinais , Humanos , Fibrose Cística/metabolismo , Fibrose Cística/patologia , Desidratação/metabolismo , Epitélio/metabolismo , Epitélio/patologia , Integrina beta1/metabolismo , Mucosa Respiratória/metabolismoRESUMO
BACKGROUND: During postharvest dehydration, grapes are subject to metabolic changes including ethanol anabolism and catabolism. These changes affect the quality of the final product and ethanol production is a key step. Ethanol dissipation has never been measured during postharvest wine grape dehydration. Thus, the present study aimed to: (i) monitor ethanol dissipation and (ii) investigate chemical-biochemical changes in berries during dehydration. RESULTS: Ethanol dissipation from Raboso grapes, under controlled postharvest dehydration, was found to comprise up to 36% of weight loss (w.l.). Moreover, the activity of enzymes involved in the anaerobic metabolism of grapes was investigated. Ethanol dissipation was highly correlated with grape weight loss (r2 = 0.989). Alcohol dehydrogenase activity, responsible for the reduction of ethanol to acetaldehyde, declined significantly with w.l. Similarly, pyruvate decarboxylase and lactate dehydrogenase reduced their activity. High lipoxygenase activity was measured at 27% w.l., whereas polyphenol oxidation was constant and declined in the last sampling. CONCLUSION: Ethanol dissipation during postharvest dehydration allows for reducing anaerobic metabolism and promotes oxidative metabolism. The sensor used can be a useful commercial tool for monitoring berry metabolism. © 2023 Society of Chemical Industry.
Assuntos
Vitis , Vinho , Vitis/metabolismo , Vinho/análise , Etanol/análise , Desidratação/metabolismo , Redução de Peso , Frutas/químicaRESUMO
The water deficit in particular, reduces the productivity of vegetable crops. To minimize these harmful effects on agriculture, several agronomic and physiological practices are being studied, such as the use of bacteria and water stress attenuators, such as brassinosteroids. Considering the socioeconomic relevance of corn culture and its sensitivity when exposed to water deficit, the objective of the present study was to evaluate the action of brassinosteroids and azospirillum on nitrogen metabolism in corn plants subjected to water stress conditions. The experiment was carried out in a greenhouse, in a period of 47 days, with corn plants, using the hybrid K9606 VIP3. The design was completely randomized, in a 2x2x3 factorial scheme, with six replications. The first factor corresponds to two water regimes (presence and absence of water deficit). The second corresponds to inoculation via seed of Azospirillum brasiliense and absence of inoculation. And the third corresponds to the application of three concentrations of brassinosteroids (0, 0.3 and 0.6 µM). Were determined Nitrate; nitrate reductase; free ammonium; total soluble aminoacids; soluble proteins; proline; glycine betaine and glutamine synthetase. The lack of water in plants provided a reduction in the protein and nitrate reductase contents, in leaves and roots. For ammonium, plants with water deficit inoculated at a concentration of 0.3 µM, obtained an increase of 7.16 (70.26%) and 13.89 (77.04%) mmol NH4 + .Kg-1. DM (Dry mass) on the leaf and root respectively. The water deficit in the soil provided significant increases in the concentrations of glycine betaine, nitrate, proline and aminoacids, both in the leaves and in the roots of the corn plants. On the other hand, the contents of glutamine synthetase had a reduction in both leaves and roots.
Assuntos
Compostos de Amônio , Azospirillum brasilense , Zea mays , Brassinosteroides/metabolismo , Nitratos , Raízes de Plantas/metabolismo , Secas , Desidratação/metabolismo , Betaína/metabolismo , Glutamato-Amônia Ligase , Aminoácidos/metabolismo , Prolina/metabolismo , Nitrato Redutases/metabolismo , Nitrogênio/metabolismoRESUMO
Proteases such as neutrophil elastase cleave and activate the epithelial sodium channel (ENaC), causing airway dehydration. Our current study explores the impact of increased protease levels in vapers' airways on ENaC activity and airway dehydration. Human bronchial epithelial cultures (HBECs) were exposed to bronchoalveolar lavage fluid (BALF) from non-smokers, smokers and vapers. Airway surface liquid (ASL) height was measured by confocal microscopy as a marker of hydration. ENaC cleavage was measured by Western blotting. Human peripheral blood neutrophils were treated with a menthol-flavored e-liquid (Juul), and the resulting secretions were added to HBECs. BALF from smokers and vapers significantly and equally increased ENaC activity and decreased ASL height. The ASL height decrease was attenuated by protease inhibitors. Non-smokers' BALF had no effect on ENaC or ASL height. BALF from smokers and vapers, but not non-smokers, induced ENaC cleavage. E-liquid-treated neutrophil secretions cleaved ENaC and decreased ASL height. Our study demonstrated that elevated protease levels in vapers' airways have functional significance since they can activate ENaC, resulting in airway dehydration. Lung dehydration contributes to diseases like cystic fibrosis (CF), chronic obstructive pulmonary disease (COPD) and asthma. Thus, our data predict that vaping, like smoking, will cause airway surface dehydration that likely leads to lung disease.
Assuntos
Vaping , Humanos , Vaping/efeitos adversos , Proteólise , Desidratação/metabolismo , Mucosa Respiratória/metabolismo , Pulmão/metabolismo , Canais Epiteliais de Sódio/metabolismoRESUMO
Modeling plants for biomass production and metal uptake from surrounding environment is strongly dependent on the moisture content of soil. Therefore, experiments were conducted to find out how soil moisture affects the phenotypic traits, photosynthetic efficiency, metabolic profile, and metal accumulation in the hyperaccumulating ecotype of Sedum alfredii (S. alfredii). A total of six water potential gradients were set: 0 ~ -15 kPa (T1), -15 ~ -30 kPa (T2), -30 ~ -45 kPa (T3), -45 ~ -60 kPa (T4), -60 ~ -75 kPa (T5), and -75 ~ -90 kPa (T6). Different water potential treatments had a significant effect on plant growth and metal uptake efficiency. Compared to T3, T2 was more effective in promoting plant growth and development, with an increase in biomass of 23% and 17% in both fresh weight (FW) and dry weight (DW), respectively. T2 and T3 had the highest cadmium (Cd) content in the shoot (280.2 mg/kg) and (283.3 mg/kg), respectively, whereas T1 had the lowest values (204.7 mg/kg). Cd availability for plants in the soil was affected by moving soil moisture cycles. Changes in soil moisture that were either too high or too low compared to the ideal soil water content for S. alfredii growth resulted in a significant reduction in Cd accumulation in shoots. Tryptophan, phenylalanine, and other amino acids were accumulated in T5, whereas only tryptophan and phenylalanine slightly increased in T1. Sugars and alcohols such as sucrose, trehalose, mannitol, galactinol, and mannobiose increased in T5, while they decreased significantly in T1. Interestingly, in contrast to T1, the two impaired metabolic pathways in T5 (galactose and starch metabolism) were identified to be glucose metabolic pathways. These findings provide scientific information (based on experiments) to improve biomass production and metal uptake efficiency in hyperaccumulating ecotype of S. alfredii for phytoremediation-contaminated agricultural fields.
Assuntos
Sedum , Poluentes do Solo , Cádmio/análise , Sedum/metabolismo , Ecótipo , Solo/química , Desidratação/metabolismo , Triptofano , Poluentes do Solo/análise , Raízes de Plantas/metabolismo , Fotossíntese , Biodegradação AmbientalRESUMO
Adult autosomal dominant polycystic kidney disease (ADPKD) has been shown to be related as a "third hit" to the occurrence of acute or chronic kidney injury. Here, we examined whether dehydration, as a common kidney risk factor, could cause cystogenesis in chronic-onset Pkd1-/- mice by regulating macrophage activation. First, we confirmed that dehydration accelerated cytogenesis in Pkd1-/- mice and that macrophages infiltrated the kidney tissues even earlier than macroscopic cyst formation. Then, microarray analysis suggested that glycolysis pathway may be involved in macrophage activation in Pkd1-/- kidneys under conditions of dehydration. Further, we confirmed glycolysis pathway was activated and lactic acid (L-LA) was overproduced in the Pkd1-/- kidney under conditions of dehydration. We have already proved that L-LA strongly stimulated M2 macrophage polarization and overproduction of polyamine in macrophage in vitro, and in the present study, we further discovered that M2 polarization-induced polyamine production shortened the primary cilia length by disrupting the PC1/PC2 complex. Finally, the activation of L-LA-arginase 1-polyamine pathway contributed to cystogenesis and progressive cyst growth in Pkd1-/- mice recurrently exposed to dehydration.
Assuntos
Cistos , Ativação de Macrófagos , Doenças Renais Policísticas , Animais , Camundongos , Cistos/metabolismo , Desidratação/metabolismo , Modelos Animais de Doenças , Rim/patologia , Macrófagos , Doenças Renais Policísticas/patologiaRESUMO
Acidity is a determinant of the organoleptic quality of apple, whereas its regulatory mechanism under water stress remains obscure. Fruit from apple 'Yanfu 3' of Fuji trees grown under normal water irrigation (CK), excessive water deficit treatment (DRT) and excessive water irrigation treatment (WAT) were sampled at 85, 100, 115, 130, 145, 160 and 175 days after full bloom designated stages S1, S2, S3, S4, S5, S6 and S7, respectively. DRT treatment reduced the individual fruit weight and fruit moisture content, and increased fruit firmness. The malate content of DRT treatment was higher than that of CK and WAT from stages S1 to S7. RNA sequencing (RNA-seq) analysis of the transcriptome at stages S4, S6 and S7 indicated that malate anabolism was associated with cysteine and methionine, auxin signaling, glyoxylate and dicarboxylate and pyruvate metabolism. Overexpression of MdPEPC4 increased the malate content in apple calli induced by 4% PEG. Our study provides novel insights into the effects of water stress on the molecular mechanism underlying apple fruit acidity.
Assuntos
Malus , Malus/genética , Malus/metabolismo , Malatos/metabolismo , Desidratação/metabolismo , Frutas/genética , Frutas/metabolismo , Perfilação da Expressão Gênica , Transcriptoma , Regulação da Expressão Gênica de PlantasRESUMO
Freeze-tolerant species, such as wood frogs (Rana sylvatica), are susceptible to multiple co-occurring stresses that they must overcome to survive. Freezing is accompanied by mechanical stress and dehydration due to ice crystal formation in the extracellular space, ischemia/anoxia due to interruption in blood flood, and hyperglycemia due to cryoprotective measures. Wood frogs can survive dehydration, anoxia, and high glucose stress independently of freezing, thereby creating a multifactorial model for studying freeze-tolerance. Oxidative stress and high glucose levels favors the production of pro-oxidant molecules and advanced glycation end product (AGE) adducts that could cause substantial cellular damage. In this study, the involvement of the high mobility group box 1 (HMGB1)-AGE/RAGE (receptor for AGE) axis and the regulation of ETS1 and EGR1-mediated angiogenic responses were investigated in liver of wood frogs expose to freeze/thaw, anoxia/reoxygenation and dehydration/rehydration treatments. HMGB1 and not AGE-adducts are likely to induce the activation of ETS1 and EGR1 via the RAGE pathway. The increase in nuclear localization of both ETS1 and EGR1, but not DNA binding activity in response to stress hints to a potential spatial and temporal regulation in inducing angiogenic factors. Freeze/thaw and dehydration/rehydration treatments increase the levels of both pro- and anti-angiogenic factors, perhaps to prepare for the distribution of cryoprotectants or enable the repair of damaged capillaries and wounds when needed. Overall, wood frogs appear to anticipate the need for angiogenesis in response to freezing and dehydration but not anoxic treatments, probably due to mechanical stress associated with the two former conditions.
Assuntos
Proteína HMGB1 , Indutores da Angiogênese/metabolismo , Animais , Desidratação/metabolismo , Redes Reguladoras de Genes , Glucose/metabolismo , Produtos Finais de Glicação Avançada/genética , Proteína HMGB1/metabolismo , Hipóxia/metabolismo , Ranidae/genética , Ranidae/metabolismoRESUMO
The biochemical properties and microstructural changes of freeze-dried Japanese scallop (Patinopecten yessoensis) striated muscle during room temperature storage and rehydration were investigated. The results showed that the content of ATP in freeze-dried scallop muscle remained stable with no significant difference (p > 0.05). However, ATP was rapidly decomposed and AMP accumulated within 1.5 min of rehydration, and HxR and Hx were gradually produced from AMP decomposition with the extension of rehydration time. Besides, the results of chymotryptic digestion patterns demonstrated that the rod of myosin was unstable after dehydration, reflecting lower salt solubility than that of frozen-thawed scallop. In contrast, the myosin subfragment-1 (S-1) was stable, as indicated by the constant of Ca2+-ATPase activity of freeze-dried scallops throughout the storage and rehydration (p > 0.05). Furthermore, the microstructural analysis revealed that the Z line of the freeze-dried scallop was broken after dehydration process. This study might be useful for developing high-quality dehydrated scallops in the future.
Assuntos
Músculo Estriado , Pectinidae , Monofosfato de Adenosina/análise , Monofosfato de Adenosina/metabolismo , Trifosfato de Adenosina/análise , Trifosfato de Adenosina/metabolismo , Animais , Desidratação/metabolismo , Hidratação , Músculo Esquelético/química , Nucleotídeos/análise , Pectinidae/química , Proteínas/análiseRESUMO
Thiamin is a crucial vitamin with a vast variety of anti-oxidative and physiological roles in plants subjected to abiotic stresses. We examined the efficiency of foliar-applied thiamin (50 and 100 mM) on growth, yield quality and key-biochemical characteristics of two cultivars (FD1 and FD3) of cauliflower (Brassica oleracea L.) under water-deficit stress. Water stress at the rate of 50% field capacity (F.C.) markedly decreased the plant biomass, leaf total phenolics and ascorbic acid (AsA) contents. In contrast, drought-induced increase was noted in the leaf [hydrogen peroxide (H2O2), AsA, proline, malondialdehyde (MDA), glycinebetaine (GB), total soluble proteins and oxidative defense system in terms of high activities of peroxidase (POD), and catalase (CAT) enzymes] and the inflorescence (total phenolics, proline, GB, MDA, H2O2, and activities of SOD and CAT enzymes) characteristics of cauliflower. However, foliar-applied thiamin significantly improved growth and physio-biochemical attributes except leaf and inflorescence MDA and H2O2 contents of both cauliflower cultivars under water stress. Overall, application of thiamin enhanced the plant growth may be associated with suppressed reactive oxygen species (ROS) and upregulated antioxidants defense system of cauliflower.
Assuntos
Fenômenos Bioquímicos , Brassica , Antioxidantes/metabolismo , Ácido Ascórbico/metabolismo , Betaína/metabolismo , Botrytis/metabolismo , Brassica/metabolismo , Desidratação/metabolismo , Peróxido de Hidrogênio/metabolismo , Prolina/metabolismo , Tiamina/metabolismoRESUMO
Now-a-days, plant-based extracts, as a cheap source of growth activators, are being widely used to treat plants grown under extreme climatic conditions. So, a trial was conducted to assess the response of two maize (Zea mays L.) varieties, Sadaf (drought tolerant) and Sultan (drought sensitive) to foliar-applied sugar beet extract (SBE) under varying water-deficit conditions. Different SBE (control, 1%, 2%, 3% & 4%) levels were used in this study, and plants were exposed to water-deficit [(75% and 60% of field capacity (FC)] and control (100% FC) conditions. It was observed that root and shoot dry weights (growth), total soluble proteins, RWC-relative water contents, total phenolics, chlorophyll pigments and leaf area per plant decreased under different water stress regimes. While, proline, malondialdehyde (MDA), RMP-relative membrane permeability, H2O2-hydrogen peroxide and the activities of antioxidant enzymes [CAT-catalase, POD-peroxidase and SOD-superoxide dismutase] were found to be improved in water stress affected maize plants. Exogenous application of varying levels of SBE ameliorated the negative effects of water-deficit stress by enhancing the growth attributes, photosynthetic pigments, RWC, proline, glycinebetaine (GB), activities of POD and CAT enzymes and levels of total phenolics, whereas it reduced the lipid peroxidation in both maize varieties under varying water stress levels. It was noted that 3% and 4% levels of SBE were more effective than the other levels used in enhancing the growth as well as other characteristics of the maize varieties. Overall, the sugar beet extract proved to be beneficial for improving growth and metabolism of maize plants exposed to water stress.
Assuntos
Beta vulgaris , Betaína , Desidratação/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Estresse Fisiológico/efeitos dos fármacos , Zea mays/efeitos dos fármacos , Secas , Malondialdeído/metabolismo , Superóxido Dismutase/metabolismo , Zea mays/metabolismoRESUMO
The Arabian camel (Camelus dromedarius) is the most important livestock animal in arid and semi-arid regions and provides basic necessities to millions of people. In the current context of climate change, there is renewed interest in the mechanisms that enable camelids to survive in arid conditions. Recent investigations described genomic signatures revealing evolutionary adaptations to desert environments. We now present a comprehensive catalogue of the transcriptomes and proteomes of the dromedary kidney and describe how gene expression is modulated as a consequence of chronic dehydration and acute rehydration. Our analyses suggested an enrichment of the cholesterol biosynthetic process and an overrepresentation of categories related to ion transport. Thus, we further validated differentially expressed genes with known roles in water conservation which are affected by changes in cholesterol levels. Our datasets suggest that suppression of cholesterol biosynthesis may facilitate water retention in the kidney by indirectly facilitating the AQP2-mediated water reabsorption.
Assuntos
Água Corporal/metabolismo , Camelus/fisiologia , Colesterol/fisiologia , Rim/metabolismo , Animais , Aquaporina 2/fisiologia , Desidratação/metabolismo , Clima Desértico , Metabolismo dos Lipídeos , Masculino , Proteoma , ATPase Trocadora de Sódio-Potássio/fisiologia , TranscriptomaRESUMO
OBJECTIVE AND DESIGN: The exacerbate inflammatory response contributes to the progressive loss of lung function in cystic fibrosis (CF), a genetic disease that affects the osmotic balance of mucus and mucociliary clearance, resulting in a microenvironment that favors infection and inflammation. The purinergic system, an extracellular signaling pathway characterized by nucleotides, enzymes and receptors, may have a protective role in the disease, through its action in airway surface liquid (ASL) and anti-inflammatory response. MATERIALS AND METHODS: To make up this review, studies covering topics of CF, inflammation, ASL and purinergic system were selected from the main medical databases, such as Pubmed and ScienceDirect. CONCLUSION: We propose several ways to modulate the purinergic system as a potential therapy for CF, like inhibition of P2X7, activation of P2Y2, A2A and A2B receptors and blocking of adenosine deaminase. Among them, we postulate that the most suitable strategy is to block the action of adenosine deaminase, which culminates in the increase of Ado levels that presents anti-inflammatory actions and improves mucociliary clearance. Furthermore, it is possible to maintain the physiological levels of ATP to control the hydration of ASL. These therapies could correct the main mechanisms that contribute to the progression of CF.
Assuntos
Fibrose Cística/metabolismo , Desidratação/metabolismo , Purinas/metabolismo , Mucosa Respiratória/metabolismo , Animais , Humanos , Inflamação/metabolismo , Receptores Purinérgicos/metabolismoRESUMO
As the polyploidy progenitor of modern sugarcane, Saccharum spontaneum is considered to be a valuable resistance source to various biotic and abiotic stresses. However, little has been reported on the mechanism of drought tolerance in S. spontaneum. Herein, the physiological changes of S. spontaneum GXS87-16 at three water-deficit levels (mild, moderate, and severe) and after re-watering during the elongation stage were investigated. RNA sequencing was utilized for global transcriptome profiling of GXS87-16 under severe drought and re-watered conditions. There were significant alterations in the physiological parameters of GXS87-16 in response to drought stress and then recovered differently after re-watering. A total of 1569 differentially expressed genes (DEGs) associated with water stress and re-watering were identified. Notably, the majority of the DEGs were induced by stress. GO functional annotations and KEGG pathway analysis assigned the DEGs to 47 GO categories and 93 pathway categories. The pathway categories were involved in various processes, such as RNA transport, mRNA surveillance, plant hormone signal transduction, and plant-pathogen interaction. The reliability of the RNA-seq results was confirmed by qRT-PCR. This study shed light on the regulatory processes of drought tolerance in S. spontaneum and identifies useful genes for genetic improvement of drought tolerance in sugarcane.
Assuntos
Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Folhas de Planta/metabolismo , Saccharum/metabolismo , Desidratação/genética , Desidratação/metabolismo , Folhas de Planta/genética , Saccharum/genéticaRESUMO
The African clawed frog (Xenopus laevis) withstands prolonged periods of extreme whole-body dehydration that lead to impaired blood flow, global hypoxia, and ischemic stress. During dehydration, these frogs shift from oxidative metabolism to a reliance on anaerobic glycolysis. In this study, we purified the central glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to electrophoretic homogeneity and investigated structural, kinetic, subcellular localization, and post-translational modification properties between control and 30% dehydrated X. laevis liver. GAPDH from dehydrated liver displayed a 25.4% reduction in maximal velocity and a 55.7% increase in its affinity for GAP, as compared to enzyme from hydrated frogs. Under dehydration mimicking conditions (150 mM urea and 1% PEG), GAP affinity was reduced with a Km value 53.8% higher than controls. Frog dehydration also induced a significant increase in serine phosphorylation, methylation, acetylation, beta-N-acetylglucosamination, and cysteine nitrosylation, post-translational modifications (PTMs). These modifications were bioinformatically predicted and experimentally validated to govern protein stability, enzymatic activity, and nuclear translocation, which increased during dehydration. These dehydration-responsive protein modifications, however, did not appear to affect enzymatic thermostability as GAPDH melting temperatures remained unchanged when tested with differential scanning fluorimetry. PTMs could promote extreme urea resistance in dehydrated GAPDH since the enzyme from dehydrated animals had a urea I50 of 7.3 M, while the I50 from the hydrated enzyme was 5.3 M. The physiological consequences of these dehydration-induced molecular modifications of GAPDH likely suppress GADPH glycolytic functions during the reduced circulation and global hypoxia experienced in dehydrated X. laevis.
Assuntos
Proteínas de Anfíbios/química , Desidratação/metabolismo , Gliceraldeído 3-Fosfato/química , Gliceraldeído-3-Fosfato Desidrogenases/química , Fígado/enzimologia , Processamento de Proteína Pós-Traducional , Xenopus laevis/metabolismo , Acetilação , Proteínas de Anfíbios/isolamento & purificação , Proteínas de Anfíbios/metabolismo , Animais , Sítios de Ligação , Desidratação/fisiopatologia , Secas , Gliceraldeído 3-Fosfato/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/isolamento & purificação , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Glicólise/fisiologia , Cinética , Fígado/química , Masculino , Metilação , Modelos Biológicos , Modelos Moleculares , Compostos Nitrosos/química , Compostos Nitrosos/metabolismo , Fosforilação , Polietilenoglicóis/química , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Homologia Estrutural de Proteína , Especificidade por Substrato , Termodinâmica , Ureia/químicaRESUMO
Vocal folds are a viscoelastic multilayered structure responsible for voice production. Vocal fold epithelial damage may weaken the protection of deeper layers of lamina propria and thyroarytenoid muscle and impair voice production. Systemic dehydration can adversely affect vocal function by creating suboptimal biomechanical conditions for vocal fold vibration. However, the molecular pathobiology of systemically dehydrated vocal folds is poorly understood. We used an in vivo rabbit model to investigate the complete gene expression profile of systemically dehydrated vocal folds. The RNA-Seq based transcriptome revealed 203 differentially expressed (DE) vocal fold genes due to systemic dehydration. Interestingly, function enrichment analysis showed downregulation of genes involved in cell adhesion, cell junction, inflammation, and upregulation of genes involved in cell proliferation. RT-qPCR validation was performed for a subset of DE genes and confirmed the downregulation of DSG1, CDH3, NECTIN1, SDC1, S100A9, SPINK5, ECM1, IL1A, and IL36A genes. In addition, the upregulation of the transcription factor NR4A3 gene involved in epithelial cell proliferation was validated. Taken together, these results suggest an alteration of the vocal fold epithelial barrier independent of inflammation, which could indicate a disruption and remodeling of the epithelial barrier integrity. This transcriptome provides a first global picture of the molecular changes in vocal fold tissue in response to systemic dehydration. The alterations observed at the transcriptional level help to understand the pathobiology of dehydration in voice function and highlight the benefits of hydration in voice therapy.
Assuntos
Desidratação/genética , Músculos Laríngeos/metabolismo , Prega Vocal/metabolismo , Distúrbios da Voz/genética , Animais , Fenômenos Biomecânicos , Adesão Celular/genética , Proliferação de Células/genética , Desidratação/metabolismo , Desidratação/patologia , Modelos Animais de Doenças , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Regulação da Expressão Gênica/genética , Humanos , Junções Intercelulares/genética , Músculos Laríngeos/patologia , Mucosa/metabolismo , Mucosa/patologia , Coelhos , Prega Vocal/patologia , Distúrbios da Voz/patologiaRESUMO
In arid conditions, the African Clawed frog Xenopus laevis enters a state of estivation dormancy as an adaptive survival strategy. Under estivation, X. laevis experience severe dehydration stress as 25-35% of total body water is lost. Dehydration in X. laevis can lead to periods of hypoxia due to elevated blood viscosity that impedes tissue perfusion. To understand how X. laevis survives under such stress, we studied the regulation pattern of key mitogen-activated protein kinases (MAPK) and their downstream transcription factors, along with several heat shock proteins in the oxygen sensitive brain and heart tissue of X. laevis under dehydration stress. Our study revealed that the activation phosphorylation residues of MAPK including JNK and MSK and their downstream transcription factors c-Jun and ATF2 are significantly decreased in the heart under dehydration. Given that JNK, c-Jun, and ATF2 are known positive regulators of apoptosis, this regulatory pattern suggest that a state of pro-survival signals may be established in the dehydrated heart. In support of this, protein levels of HSP60, a pro-apoptotic mitochondrial chaperone, was also downregulated in the heart in response to dehydration stress. In the brain tissue, most proteins remain unchanged with the exception of the apoptosis regulating p53 transcription factor, which showed a significant decrease in its activating phosphorylation residue under dehydration. Overall, our study revealed that in the Xenopus brain and heart, a specific suppression pattern of MAPK, transcription factors, and HSP takes place to potentially establish a state of pro-survival under dehydration stress.
Assuntos
Chaperonina 60/metabolismo , Desidratação/metabolismo , Sistema de Sinalização das MAP Quinases/genética , Proteína Supressora de Tumor p53/metabolismo , Xenopus laevis/metabolismo , Fator 2 Ativador da Transcrição/metabolismo , Animais , Encéfalo/metabolismo , Estivação/fisiologia , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Resposta ao Choque Térmico/genética , Resposta ao Choque Térmico/fisiologia , MAP Quinase Quinase 4/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Fosforilação , Proteínas Proto-Oncogênicas c-jun/metabolismo , Proteínas Quinases S6 Ribossômicas 90-kDa/metabolismoRESUMO
The African clawed frog (Xenopus laevis) naturally tolerates severe dehydration using biochemical adaptation, one of which is the elevation of antioxidant defenses during whole-body dehydration. The present study investigated the role and regulation of a pathway known to regulate oxidative stress response, the Akt-FoxO signaling pathway, in clawed frog skeletal muscle, responding to medium (15%) and high (30%) dehydration. Protein levels of total and phosphorylated Akt, FoxO1, and FoxO3 were assessed via immunoblotting, in addition to the levels of the E3 ubiquitin ligase known to be associated with muscle atrophy, MAFbx. Akt activity/phosphorylation in addition to its total protein levels were decreased in the skeletal muscle during dehydration, and this corresponded with decreases in the relative phosphorylation of FoxO1 and FoxO3 as well on several residues. Akt is an inhibitor of FoxO1 and FoxO3 activity via phosphorylation, suggesting that FoxO activities were increased during dehydration stress. Furthermore, MAFbx showed decreased protein expression during high dehydration as well, suggesting that the clawed frog may exhibit some natural resistance to skeletal muscle atrophy during severe dehydration conditions. In addition to identifying that the suppression of Akt could lead to an activation of FoxO transcription factors in X. laevis during dehydration, these investigations suggest that X. laevis dehydration may implicate FoxO1 and FoxO3 in controlling skeletal muscle atrophy in X. laevis exposed to dehydration. This study implicates the Akt signaling pathway, its regulation of FoxO transcription factors, and FoxO-controlled targets, in stress adaptation against dehydration.
Assuntos
Desidratação/metabolismo , Proteína Forkhead Box O1/metabolismo , Proteína Forkhead Box O3/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas de Xenopus/metabolismo , Animais , Fosforilação , Estresse FisiológicoRESUMO
Eukaryotic translation elongation factors are implicated in protein synthesis across different living organisms, but their biological functions in the pathogenesis of cucumber mosaic virus (CMV) and tobacco rattle virus (TRV) infections are poorly understood. Here, we isolated and characterized a cDNA clone, LreEF1A4, encoding the alpha subunit of elongation factor 1, from a CMV-elicited suppression subtractive hybridization library of Lilium regale. The infection tests using CMV remarkably increased transcript abundance of LreEF1A4; however, it also led to inconsistent expression profiles of three other LreEF1A homologs (LreEF1A1-3). Protein modelling analysis revealed that the amino acid substitutions among four LreEF1As may not affect their enzymatic functions. LreEF1A4 was ectopically overexpressed in petunia (Petunia hybrida), and transgenic plants exhibited delayed leaf and flower senescence, concomitant with increased transcription of photosynthesis-related genes and reduced expression of senescence-associated genes, respectively. A compromised resistance to CMV and TRV infections was found in transgenic petunia plants overexpressing LreEF1A4, whereas its overexpression resulted in an enhanced tolerance to salt and drought stresses. Taken together, our data demonstrate that LreEF1A4 functions as a positive regulator in viral multiplication and plant adaption to high salinity and dehydration.