RESUMEN
Hexokinase 2 (HK2), the first glycolytic rate-limiting enzyme, is closely correlated with the occurrence and progression of tumors. Effective therapeutic agents targeting HK2 are urgently needed. Bergenin has exhibited various pharmacological activities, such as antitumor properties. However, the effects of bergenin on the abnormal glucose metabolism of cancer cells are yet unclear. In this study, HK2 was overexpressed in OSCC tissues, and the depletion of HK2 inhibited the growth of OSCC cells in vitro and in vivo. Moreover, these results showed that the natural compound, bergenin, exerted a robust antitumor effect on OSCC cells. Bergenin inhibited cancer cell proliferation, suppressed glycolysis, and induced intrinsic apoptosis in OSCC cells by downregulating HK2. Notably, bergenin restored the antitumor efficacy of irradiation in the radioresistant OSCC cells. A mechanistic study revealed that bergenin upregulated the protein level of phosphatase and the tensin homolog deleted on chromosome 10 (PTEN) by enhancing the interaction between PTEN and ubiquitin-specific protease 13 (USP13) and stabilizing PTEN; this eventually inhibited AKT phosphorylation and HK2 expression. Bergenin was identified as a novel therapeutic agent against glycolysis to inhibit OSCC and overcome radioresistance. Targeting PTEN/AKT/HK2 signaling could be a promising option for clinical OSCC treatment.
Asunto(s)
Neoplasias , Proteínas Proto-Oncogénicas c-akt , Humanos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal , Glucólisis/genética , Línea Celular Tumoral , Proliferación Celular , Proteasas Ubiquitina-Específicas/metabolismo , Proteasas Ubiquitina-Específicas/farmacologíaRESUMEN
The proportions of the various muscle fiber types are important in the regulation of skeletal muscle metabolism, as well as animal meat production. Four-and-a-half LIM domain protein 3 (FHL3) is highly expressed in fast glycolytic muscle fibers and differentially regulates the expression of myosin heavy chain (MyHC) isoforms at the cellular level. Whether FHL3 regulates the transformation of muscle fiber types in vivo and the regulatory mechanism is unclear. In this study, muscle-specific FHL3 transgenic mice were generated by random integration, and lentivirus-mediated gene knockdown or overexpression in muscles of mice or pigs was conducted. Functional analysis showed that overexpression of FHL3 in muscles significantly increased the proportion of fast-twitch myofibers and muscle mass but decreased muscle succinate dehydrogenase (SDH) activity and whole-body oxygen consumption. Lentivirus-mediated FHL3 knockdown in muscles significantly decreased muscle mass and the proportion of fast-twitch myofibers. Mechanistically, FHL3 directly interacted with the Yin yang 1 (YY1) DNA-binding domain, repressed the binding of YY1 to the fast glycolytic MyHC2b gene regulatory region, and thereby promoted MyHC2b expression. FHL3 also competed with EZH2 to bind the repression domain of YY1 and reduced H3K27me3 enrichment in the MyHC2b regulatory region. Moreover, FHL3 overexpression reduced glucose tolerance by affecting muscle glycolytic metabolism, and its mRNA expression in muscle was positively associated with hemoglobin A1c (HbA1c) in patients with type 2 diabetes. Therefore, FHL3 is a novel potential target gene for the treatment of muscle metabolism-related diseases and improvement of animal meat production.
Asunto(s)
Diabetes Mellitus Tipo 2 , Ratones , Porcinos , Animales , Diabetes Mellitus Tipo 2/metabolismo , Fibras Musculares de Contracción Rápida/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/metabolismo , Glucólisis/genética , Cadenas Pesadas de Miosina/genética , Cadenas Pesadas de Miosina/metabolismoRESUMEN
NRAS-mutated melanoma lacks a specific line of treatment. Metabolic reprogramming is considered a novel target to control cancer; however, NRAS-oncogene contribution to this cancer hallmark is mostly unknown. Here, we show that NRASQ61-mutated melanomas specific metabolic settings mediate cell sensitivity to sorafenib upon metabolic stress. Mechanistically, these cells are dependent on glucose metabolism, in which glucose deprivation promotes a switch from CRAF to BRAF signaling. This scenario contributes to cell survival and sustains glucose metabolism through BRAF-mediated phosphorylation of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-2/3 (PFKFB2/PFKFB3). In turn, this favors the allosteric activation of phosphofructokinase-1 (PFK1), generating a feedback loop that couples glycolytic flux and the RAS signaling pathway. An in vivo treatment of NRASQ61 mutant melanomas, including patient-derived xenografts, with 2-deoxy-D-glucose (2-DG) and sorafenib effectively inhibits tumor growth. Thus, we provide evidence for NRAS-oncogene contributions to metabolic rewiring and a proof-of-principle for the treatment of NRASQ61-mutated melanoma combining metabolic stress (glycolysis inhibitors) and previously approved drugs, such as sorafenib.
Asunto(s)
Melanoma , Proteínas Proto-Oncogénicas B-raf , Humanos , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas B-raf/metabolismo , Sorafenib/farmacología , Línea Celular Tumoral , Mutación , Melanoma/tratamiento farmacológico , Melanoma/genética , Melanoma/metabolismo , Glucólisis/genética , Glucosa/metabolismo , Estrés Fisiológico , Fosfofructoquinasa-2/metabolismo , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , GTP Fosfohidrolasas/genética , GTP Fosfohidrolasas/metabolismoRESUMEN
The active ingredient of the traditional Chinese medicine comfrey is shikonin, a naphthoquinone compound. The focus of this study was to investigate the effect of shikonin on the proliferation, invasion, migration, and chemoresistance of non-small cell lung cancer (NSCLC) cells, and to explore its underlying molecular biological mechanisms. The results show that shikonin inhibited the viability, proliferation, invasion, and migration of NSCLC cells A549 and PC9, and induced apoptosis. As the inhibitor of pyruvate kinase M2 (PKM2), a key enzyme in glycolysis, shikonin inhibited glucose uptake and the production of lactate, the final metabolite of aerobic glycolysis. In vivo chemotherapeutic assay showed that shikonin reduced the tumor volume and weight in NSCLC mice model and increased the sensitivity to cisplatin chemotherapy. Histoimmunology experiments showed the combination of shikonin and cisplatin downregulated the expression of PKM2 and its transcriptionally regulated downstream gene glucose transporter 1 (Glut1) in tumor tissue. In an assessment of glucose metabolism, micro-PET/CT data showed a combination of shikonin and cisplatin inhibited the fluorodeoxy glucose (18F-FDG) uptake into tumor. Since exosomal PKM2 affected the sensitivity to cisplatin in NSCLC cells, we also demonstrated shikonin could inhibit exosome secretion and exosomal PKM2 through the administration of exosomal inhibitor GW4869. Furthermore, shikonin sensitized cisplatin treatment by reducing the extracellular secretion of exosomal PKM2. In conclusion, we suggest that shikonin not only inhibits PKM2 intracellularly but also reduces glycolytic flux and increases cisplatin sensitivity through the exosomal pathway.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Naftoquinonas , Animales , Carcinoma de Pulmón de Células no Pequeñas/genética , Línea Celular Tumoral , Proliferación Celular , Cisplatino/farmacología , Glucólisis/genética , Neoplasias Pulmonares/genética , Ratones , Naftoquinonas/farmacología , Naftoquinonas/uso terapéutico , Tomografía Computarizada por Tomografía de Emisión de Positrones , Piruvato Quinasa/genética , Piruvato Quinasa/metabolismoRESUMEN
Melatonin is synthesized in the pineal gland at night. Since melatonin is produced in the mitochondria of all other cells in a non-circadian manner, the amount synthesized by the pineal gland is less than 5% of the total. Melatonin produced in mitochondria influences glucose metabolism in all cells. Many pathological cells adopt aerobic glycolysis (Warburg effect) in which pyruvate is excluded from the mitochondria and remains in the cytosol where it is metabolized to lactate. The entrance of pyruvate into the mitochondria of healthy cells allows it to be irreversibly decarboxylated by pyruvate dehydrogenase (PDH) to acetyl coenzyme A (acetyl-CoA). The exclusion of pyruvate from the mitochondria in pathological cells prevents the generation of acetyl-CoA from pyruvate. This is relevant to mitochondrial melatonin production, as acetyl-CoA is a required co-substrate/co-factor for melatonin synthesis. When PDH is inhibited during aerobic glycolysis or during intracellular hypoxia, the deficiency of acetyl-CoA likely prevents mitochondrial melatonin synthesis. When cells experiencing aerobic glycolysis or hypoxia with a diminished level of acetyl-CoA are supplemented with melatonin or receive it from another endogenous source (pineal-derived), pathological cells convert to a more normal phenotype and support the transport of pyruvate into the mitochondria, thereby re-establishing a healthier mitochondrial metabolic physiology.
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Glucosa/metabolismo , Melatonina/genética , Mitocondrias/metabolismo , Neoplasias/metabolismo , Aerobiosis/genética , Comunicación Celular/genética , Glucólisis/genética , Humanos , Melatonina/metabolismo , Neoplasias/genética , Neoplasias/patología , Efecto Warburg en OncologíaRESUMEN
Lung cancer is one of the most lethal diseases and therefore poses a significant threat to human health. The Warburg effect, which is the observation that cancer cells predominately produce energy through glycolysis, even under aerobic conditions, is a hallmark of cancer. 6phosphofructo2kinase/fructose2,6biphosphatase 2 (PFKFB) is an important regulator of glycolysis. Shikonin is a Traditional Chinese herbal medicine, which has been reported to exert antitumor effects. The present study aimed to investigate the anticancer activity of shikonin in lung cancer. Cell Counting Kit8 (CCK8) and colony formation assays were used to analyze proliferation in A549 and H446 cells. Wound healing and Transwell assays were used to measure migration and invasion in A549 and H446 cells. Cell apoptosis was analyzed using flow cytometry. Lactate levels, glucose uptake and cellular ATP levels were measured using their corresponding commercial kits. Western blotting was performed to analyze the protein expression levels of key enzymes involved in aerobic glucose metabolism. Reverse transcriptionquantitative PCR was used to analyze the mRNA expression levels of PFKFB2. The results of the present study revealed that PFKFB2 expression levels were significantly upregulated in NSCLC tissues. Shikonin treatment decreased the proliferation, migration, invasion, glucose uptake, lactate levels, ATP levels and PFKFB2 expression levels and increased apoptosis in lung cancer cells in a dosedependent manner. The overexpression of PFKFB2 increased the proliferation, migration, glucose uptake, lactate levels and ATP levels in lung cancer cells, while the knockdown of PFKFB2 expression exerted the opposite effects. Moreover, there were no significant differences in lung cancer cell migration, apoptosis, glucose uptake, lactate levels and ATP levels between cells with knocked down PFKFB2 expression or treated with shikonin and the knockdown of PFKFB2 in cells treated with shikonin. In conclusion, the results of the present study revealed that shikonin inhibited the Warburg effect and exerted antitumor activity in lung cancer cells, which was associated with the downregulation of PFKFB2 expression.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Naftoquinonas/farmacología , Fosfofructoquinasa-2/genética , Anciano , Apoptosis/efectos de los fármacos , Apoptosis/genética , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Línea Celular , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Relación Dosis-Respuesta a Droga , Regulación hacia Abajo/efectos de los fármacos , Femenino , Regulación Neoplásica de la Expresión Génica , Glucólisis/efectos de los fármacos , Glucólisis/genética , Humanos , Neoplasias Pulmonares/metabolismo , Masculino , Persona de Mediana Edad , Fosfofructoquinasa-2/metabolismo , Regulación hacia Arriba/genética , Efecto Warburg en Oncología/efectos de los fármacosRESUMEN
BACKGROUND: Trace elements function as essential cofactors that are involved in various biochemical processes in mammals. Autophagy is vital for nutrient supplement, which is an important Zeitegber for the circadian homeostasis in heart. Here, we considered the possibility that autophagy, as well as the cardiomyocyte clock and glycolysis are interlinked. Detrimental effects were observed when cardiac system is exposed to bromine containing drugs. This study investigated the effects and mechanisms of bromide on the circadian clock and glycolytic metabolism of H9C2 cardiomyocytes. RESULTS: In the present study, bromide does not affect cell viability and apoptosis of H9C2 cardiomyocytes. Bromide dampens the clock and glycolytic (Hk2 and Pkm2) gene expression rhythmicity in a dose-dependent manner. Additionally, bromide inhibits autophagic process in H9C2 cardiomyocytes. In contrast, rapamycin (an autophagy inducer) dramatically restores the inhibitory effect of NaBr on the mRNA expression levels of clock genes (Bmal1, Cry1 and Rorα) and glycolytic genes (Hk2 and Pkm2). CONCLUSIONS: Our results reveal that bromide represses the clock and glycolytic gene expression patterns, partially through inhibition of autophagy.
Asunto(s)
Autofagia/efectos de los fármacos , Bromuros/farmacología , Relojes Circadianos/efectos de los fármacos , Glucólisis/efectos de los fármacos , Miocitos Cardíacos , Factores de Transcripción ARNTL/genética , Factores de Transcripción ARNTL/metabolismo , Animales , Bromuros/metabolismo , Línea Celular , Relojes Circadianos/genética , Criptocromos/genética , Criptocromos/metabolismo , Expresión Génica , Glucólisis/genética , Hexoquinasa/genética , Hexoquinasa/metabolismo , Homeostasis , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Miembro 1 del Grupo F de la Subfamilia 1 de Receptores Nucleares/genética , Miembro 1 del Grupo F de la Subfamilia 1 de Receptores Nucleares/metabolismo , Piruvato Quinasa/genética , Piruvato Quinasa/metabolismo , RatasRESUMEN
FoxA2 is an essential transcription factor for liver organogenesis and homeostasis. Although reduced expression of FoxA2 has been associated with chronic liver diseases, hepatic progenitor cells (HPCs) that are activated in these circumstances express FoxA2. However, the functional effects and underlying mechanism of FoxA2 in HPCs are still unknown. As revealed by immunostaining, HPCs expressed FoxA2 in human cirrhotic livers and in the livers of choline-deficient diet supplemented with ethionine (CDE) rats. Knocking down FoxA2 in HPCs isolated from CDE rats significantly increased cell proliferation and aerobic glycolysis. Moreover, gene transcription, protein expression, and the enzyme activities of hexokinase 2 (HK2) were upregulated, and blocking HK2 activities via 2-deoxyglucose markedly reduced cell proliferation and aerobic glycolysis. Kyoto Encyclopedia of Genes and Genomes analysis revealed that FoxA2 knockdown enhanced the transcription of genes involved in the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway and triggered downstream Akt phosphorylation. Blocking the PI3K/Akt pathway by Ly294002 inhibited HK2 activities, aerobic glycolysis, and cell proliferation in FoxA2-knockdown cells. Therefore, FoxA2 plays an important role in the proliferation and inhibition of HPCs by suppressing PI3K/Akt/HK2-regulated aerobic glycolysis.
Asunto(s)
Glucólisis/genética , Factor Nuclear 3-beta del Hepatocito/genética , Hexoquinasa/genética , Hígado/metabolismo , Organogénesis/genética , Animales , Apoptosis/genética , Línea Celular Tumoral , Proliferación Celular/genética , Colina/farmacología , Deficiencia de Colina/genética , Deficiencia de Colina/metabolismo , Hepatocitos/metabolismo , Humanos , Hígado/crecimiento & desarrollo , Fosfatidilinositol 3-Quinasa/genética , Fosforilación/genética , Proteínas Proto-Oncogénicas c-akt/genética , Ratas , Células Madre/metabolismoRESUMEN
Vitamin K2 has been shown to exert remarkable anticancer activity. However, the detailed mechanism remains unclear. Here, our study was the first to show that Vitamin K2 significantly promoted the glycolysis in bladder cancer cells by upregulating glucose consumption and lactate production, whereas inhibited TCA cycle by reducing the amounts of Acetyl-CoA. Moreover, suppression of PI3K/AKT and HIF-1α attenuated Vitamin K2-increased glucose consumption and lactate generation, indicating that Vitamin K2 promotes PI3K/AKT and HIF-1α-mediated glycolysis in bladder cancer cells. Importantly, upon glucose limitation, Vitamin K2-upregulated glycolysis markedly induced metabolic stress, along with AMPK activation and mTORC1 pathway suppression, which subsequently triggered AMPK-dependent autophagic cell death. Intriguingly, glucose supplementation profoundly abrogated AMPK activation and rescued bladder cancer cells from Vitamin K2-triggered autophagic cell death. Furthermore, both inhibition of PI3K/AKT/HIF-1α and attenuation of glycolysis significantly blocked Vitamin K2-induced AMPK activation and subsequently prevented autophagic cell death. Collectively, these findings reveal that Vitamin K2 could induce metabolic stress and trigger AMPK-dependent autophagic cell death in bladder cancer cells by PI3K/AKT/HIF-1α-mediated glycolysis promotion.
Asunto(s)
Autofagia/genética , Proteínas Quinasas/genética , Neoplasias de la Vejiga Urinaria/genética , Vitamina K 2/farmacología , Quinasas de la Proteína-Quinasa Activada por el AMP , Autofagia/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Glucosa/metabolismo , Glucólisis/genética , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Diana Mecanicista del Complejo 1 de la Rapamicina/genética , Proteína Oncogénica v-akt/genética , Fosfatidilinositol 3-Quinasas/genética , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
Enterovirus 71 (EV71) infection is an endemic disease in Southeast Asia and China. We have previously shown that EV71 virus causes functional changes in mitochondria. It is speculative whether EV71 virus alters the host cell metabolism to its own benefit. Using a metabolomics approach, we demonstrate that EV71-infected Vero cells had significant changes in metabolism. Glutathione and its related metabolites, and several amino acids, such as glutamate and aspartate, changed significantly with the infectious dose of virus. Other pathways, including glycolysis and tricarboxylic acid cycle, were also altered. A change in glutamine/glutamate metabolism is critical to the viral infection. The presence of glutamine in culture medium was associated with an increase in viral replication. Dimethyl α-ketoglutarate treatment partially mimicked the effect of glutamine supplementation. In addition, the immunoblot analysis revealed that the expression of glutamate dehydrogenase (GDH) and trifunctional carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (CAD) increased during infection. Knockdown of expression of glutaminase (GLS), GDH and CAD drastically reduced the cytopathic effect (CPE) and viral replication. Furthermore, we found that CAD bound VP1 to promote the de novo pyrimidine synthesis. Our findings suggest that virus may induce metabolic reprogramming of host cells to promote its replication through interactions between viral and host cell proteins.
Asunto(s)
Dihidroorotasa/metabolismo , Enterovirus Humano A/fisiología , Infecciones por Enterovirus/metabolismo , Glutamato Deshidrogenasa/metabolismo , Glutaminasa/metabolismo , Interacciones Huésped-Patógeno/genética , Replicación Viral/efectos de los fármacos , Replicación Viral/genética , Animales , Chlorocebus aethiops , Efecto Citopatogénico Viral/efectos de los fármacos , Efecto Citopatogénico Viral/genética , Dihidroorotasa/genética , Infecciones por Enterovirus/virología , Técnicas de Silenciamiento del Gen , Glutamato Deshidrogenasa/genética , Ácido Glutámico/metabolismo , Glutaminasa/genética , Glutamina/metabolismo , Glutamina/farmacología , Glucólisis/genética , Ácidos Cetoglutáricos/farmacología , Interferencia de ARN , Transfección , Células VeroRESUMEN
BACKGROUND: Triacylglycerols (TAGs) are the main composition of plant seed oil. Long-chain acyl-coenzyme A synthetases (LACSs) catalyze the synthesis of long-chain acyl-coenzyme A, which is one of the primary substrates for TAG synthesis. In Arabidopsis, the LACS gene family contains nine members, among which LACS1 and LACS9 have overlapping functions in TAG biosynthesis. However, functional characterization of LACS proteins in rapeseed have been rarely reported. RESULTS: An orthologue of the Arabidopsis LACS2 gene (BnLACS2) that is highly expressed in developing seeds was identified in rapeseed (Brassica napus). The BnLACS2-GFP fusion protein was mainly localized to the endoplasmic reticulum, where TAG biosynthesis occurs. Interestingly, overexpression of the BnLACS2 gene resulted in significantly higher oil contents in transgenic rapeseed plants compared to wild type, while BnLACS2-RNAi transgenic rapeseed plants had decreased oil contents. Furthermore, quantitative real-time PCR expression data revealed that the expression of several genes involved in glycolysis, as well as fatty acid (FA) and lipid biosynthesis, was also affected in transgenic plants. CONCLUSIONS: A long chain acyl-CoA synthetase, BnLACS2, located in the endoplasmic reticulum was identified in B. napus. Overexpression of BnLACS2 in yeast and rapeseed could increase oil content, while BnLACS2-RNAi transgenic rapeseed plants exhibited decreased oil content. Furthermore, BnLACS2 transcription increased the expression of genes involved in glycolysis, and FA and lipid synthesis in developing seeds. These results suggested that BnLACS2 is an important factor for seed oil production in B. napus.
Asunto(s)
Brassica napus , Coenzima A Ligasas , Semillas/metabolismo , Triglicéridos/biosíntesis , Brassica napus/genética , Brassica napus/metabolismo , Coenzima A Ligasas/genética , Coenzima A Ligasas/metabolismo , Ácidos Grasos/biosíntesis , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Glucólisis/genética , Metabolismo de los Lípidos/genética , Aceites de Plantas/metabolismo , Plantas Modificadas Genéticamente/genética , Interferencia de ARN , Triglicéridos/genéticaRESUMEN
Pseudozyma antarctica is a nonpathogenic phyllosphere yeast known as an excellent producer of industrial lipases and mannosylerythritol lipids (MELs), which are multi-functional glycolipids. The fungus produces a much higher amount of MELs from vegetable oil than from glucose, whereas its close relative, Ustilago maydis UM521, produces a lower amount of MELs from vegetable oil. In the present study, we used previous gene expression profiles measured by DNA microarray analyses after culturing on two carbon sources, glucose and soybean oil, to further characterize MEL biosynthesis in P. antarctica T-34. A total of 264 genes were found with induction ratios and expression intensities under oily conditions with similar tendencies to those of MEL cluster genes. Of these, 93 were categorized as metabolic genes using the Eukaryotic Orthologous Groups classification. Within this metabolic category, amino acids, carbohydrates, inorganic ions, and secondary metabolite metabolism, as well as energy production and conversion, but not lipid metabolism, were enriched. Furthermore, genes involved in central metabolic pathways, such as glycolysis and the tricarboxylic acid cycle, were highly induced in P. antarctica T-34 under oily conditions, whereas they were suppressed in U. maydis UM521. These results suggest that the central metabolism of P. antarctica T-34 under oily conditions contributes to its excellent oil utilization and extracellular glycolipid production.
Asunto(s)
Glucolípidos/biosíntesis , Redes y Vías Metabólicas/genética , Transcriptoma , Ustilago/genética , Ustilago/metabolismo , Ciclo del Ácido Cítrico/genética , Perfilación de la Expresión Génica , Glucosa/metabolismo , Glucólisis/genética , Familia de Multigenes , Aceite de Soja/metabolismoRESUMEN
Altered cellular metabolism is believed to be an important contributor to pathogenesis of the neurodegenerative disorder Huntington's disease (HD). Research has primarily focused on mitochondrial toxicity, which can cause death of the vulnerable striatal neurons, but other aspects of metabolism have also been implicated. Most previous studies have been carried out using postmortem human brain or non-human cells. Here, we studied bioenergetics in an induced pluripotent stem cell-based model of the disease. We found decreased adenosine triphosphate (ATP) levels in HD cells compared to controls across differentiation stages and protocols. Proteomics data and multiomics network analysis revealed normal or increased levels of mitochondrial messages and proteins, but lowered expression of glycolytic enzymes. Metabolic experiments showed decreased spare glycolytic capacity in HD neurons, while maximal and spare respiratory capacities driven by oxidative phosphorylation were largely unchanged. ATP levels in HD neurons could be rescued with addition of pyruvate or late glycolytic metabolites, but not earlier glycolytic metabolites, suggesting a role for glycolytic deficits as part of the metabolic disturbance in HD neurons. Pyruvate or other related metabolic supplements could have therapeutic benefit in HD.
Asunto(s)
Cuerpo Estriado/metabolismo , Metabolismo Energético/genética , Enfermedad de Huntington/metabolismo , Neuronas/metabolismo , Adenosina Trifosfato/metabolismo , Diferenciación Celular/genética , Línea Celular , Cuerpo Estriado/patología , Glucólisis/genética , Humanos , Enfermedad de Huntington/genética , Enfermedad de Huntington/patología , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/patología , Metaboloma/genética , Mitocondrias/genética , Mitocondrias/metabolismo , Neuronas/patología , Fosforilación OxidativaRESUMEN
Citrin, encoded by SLC25A13 gene, is an inner mitochondrial transporter that is part of the malate-aspartate shuttle, which regulates the NAD+/NADH ratio between the cytosol and mitochondria. Citrullinemia type II (CTLN-II) is an inherited disorder caused by germline mutations in SLC25A13, manifesting clinically in growth failure that can be alleviated by dietary restriction of carbohydrates. The association of citrin with glycolysis and NAD+/NADH ratio led us to hypothesize that it may play a role in carcinogenesis. Indeed, we find that citrin is upregulated in multiple cancer types and is essential for supplementing NAD+ for glycolysis and NADH for oxidative phosphorylation. Consequently, citrin deficiency associates with autophagy, whereas its overexpression in cancer cells increases energy production and cancer invasion. Furthermore, based on the human deleterious mutations in citrin, we found a potential inhibitor of citrin that restricts cancerous phenotypes in cells. Collectively, our findings suggest that targeting citrin may be of benefit for cancer therapy.
Asunto(s)
Carcinogénesis/genética , Mitocondrias/genética , Proteínas de Transporte de Membrana Mitocondrial/genética , Neoplasias/genética , Carbohidratos/genética , Citrulinemia/genética , Citrulinemia/metabolismo , Citosol/metabolismo , Citosol/patología , Regulación Neoplásica de la Expresión Génica/genética , Mutación de Línea Germinal/genética , Glutamatos/farmacología , Ácido Glutámico/análogos & derivados , Ácido Glutámico/farmacología , Glucólisis/genética , Humanos , Mitocondrias/metabolismo , Mitocondrias/patología , Proteínas de Transporte de Membrana Mitocondrial/antagonistas & inhibidores , Terapia Molecular Dirigida , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Fosforilación Oxidativa/efectos de los fármacosRESUMEN
Antiretroviral therapy extends survival but does not eliminate HIV from its cellular reservoirs. Between immune and stromal cells in the tissue microenvironment, a dynamic intercellular communication might influence host viral immune responses via intercellular transfer of extracellular vehicles (EVs) (microvesicles, exosome, or apoptotic bodies). It is increasingly recognized that HIV-infected macrophage-secreted nucleotide-rich exosomes might play a critical role in mediating communication between macrophages and other structural cells; however, molecular mechanisms underlying cell-cell crosstalk remain unknown. Here we show that HIV-1-infected macrophages and HIV-1 proteins Tat or gp120-treated macrophages express high levels of microRNAs, including miR-23a and miR-27a. Identical miRNAs expression patterns were detected in macrophage-secreted exosomes isolated from bronchoalveolar lavage fluid of HIV transgenic rats. Tat-treated macrophage-derived exosomal miR-23a attenuated posttranscriptional modulation of key tight junction protein zonula occludens (ZO-1) 3'-UTR in epithelial cells. In parallel, exosomal miR-27a released from Tat-treated macrophages altered the mitochondrial bioenergetics of recipient lung epithelial cells by targeting peroxisome proliferator-activated receptor gamma (PPARγ), while simultaneously stimulating glycolysis. Together, exosomal miRNAs shuttle from macrophages to epithelial cells and thereby explain in part HIV-mediated lung epithelial barrier dysfunction. These studies suggest that targeting miRNAs may be of therapeutic value to enhance lung health in HIV.
Asunto(s)
Pulmón/metabolismo , MicroARNs/genética , Mitocondrias/metabolismo , Movimiento Celular/efectos de los fármacos , Metabolismo Energético/genética , Células Epiteliales/metabolismo , Células Epiteliales/patología , Células Epiteliales/virología , Vesículas Extracelulares/genética , Glucólisis/genética , Proteína gp120 de Envoltorio del VIH/genética , Proteína gp120 de Envoltorio del VIH/farmacología , VIH-1/genética , VIH-1/patogenicidad , Humanos , Pulmón/patología , Pulmón/virología , Macrófagos/metabolismo , Macrófagos/patología , Macrófagos/virología , Mitocondrias/patología , Mitocondrias/virología , PPAR gamma/genética , Proteína de la Zonula Occludens-1/genética , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/farmacologíaRESUMEN
In the last decade, carbon monoxide-releasing molecules (CORMs) have been shown to act against several pathogens and to be promising antimicrobials. However, the understanding of the mode of action and reactivity of these compounds on bacterial cells is still deficient. In this work, we used a metabolomics approach to probe the toxicity of the ruthenium(II) complex Ru(CO)3Cl(glycinate) (CORM-3) on Escherichia coli By resorting to 1H nuclear magnetic resonance, mass spectrometry, and enzymatic activities, we show that CORM-3-treated E. coli accumulates larger amounts of glycolytic intermediates, independently of the oxygen growth conditions. The work provides several evidences that CORM-3 inhibits glutamate synthesis and the iron-sulfur enzymes of the tricarboxylic acid (TCA) cycle and that the glycolysis pathway is triggered in order to establish an energy and redox homeostasis balance. Accordingly, supplementation of the growth medium with fumarate, α-ketoglutarate, glutamate, and amino acids cancels the toxicity of CORM-3. Importantly, inhibition of the iron-sulfur enzymes glutamate synthase, aconitase, and fumarase is only observed for compounds that liberate carbon monoxide. Altogether, this work reveals that the antimicrobial action of CORM-3 results from intracellular glutamate deficiency and inhibition of nitrogen and TCA cycles.
Asunto(s)
Antibacterianos/farmacología , Monóxido de Carbono/farmacología , Ciclo del Ácido Cítrico/efectos de los fármacos , Escherichia coli/efectos de los fármacos , Regulación Bacteriana de la Expresión Génica , Nitrógeno/metabolismo , Compuestos Organometálicos/farmacología , Aconitato Hidratasa/antagonistas & inhibidores , Aconitato Hidratasa/genética , Aconitato Hidratasa/metabolismo , Antibacterianos/química , Monóxido de Carbono/química , Ciclo del Ácido Cítrico/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Fumarato Hidratasa/antagonistas & inhibidores , Fumarato Hidratasa/genética , Fumarato Hidratasa/metabolismo , Fumaratos/metabolismo , Glutamato Sintasa/antagonistas & inhibidores , Glutamato Sintasa/genética , Glutamato Sintasa/metabolismo , Ácido Glutámico/metabolismo , Glucólisis/efectos de los fármacos , Glucólisis/genética , Ácidos Cetoglutáricos/metabolismo , Espectroscopía de Resonancia Magnética , Metabolómica/métodos , Compuestos Organometálicos/química , Oxidación-ReducciónRESUMEN
Yin-Yang 1 (YY1) has been identified as playing critical roles in multiple diseases. However, little is known regarding its roles and mechanisms in cerebral ischemia/reperfusion (I/R) injury. This study is aimed to explore the roles of YY1 in regulating neuronal apoptosis in cerebral I/R injury and its underlying mechanisms. Primary mouse cerebral cortical neurons were isolated and subjected to OGD/R to mimic cerebral I/R injury in vitro. The roles of YY1 on OGD/R-induced neuronal injury were investigated by performing western blotting, quantitative real-time polymerase chain reaction, TUNEL, RNA-binding protein immunoprecipitation, chromatin immunoprecipitation, chromatin isolation by RNA purification assay, glucose uptake assay, lactate production assay, and extracellular acidification rate assay. YY1-binding long non-coding RNAs (LncRNAs) in neurons subjected to OGD/R were identified by RIP and RNA sequencing. The roles of YY1 on cerebral I/R in vivo were detected by assessing neuronbehaviour, infarct size, and neuronal apoptosis. We found that YY1 expression is downregulated, and LncRNA GAS5 is upregulated in neurons subjected to OGD/R. OGD/R treatment promotes YY1 interacting with GAS5 in neurons, and YY1 negatively regulates GAS5 expression by binding to GAS5 promoter to repress its transcription. Besides, YY1 and GAS5 bind to the same region of PFKFB3 promoter to promote PFKFB3 expression and strengthen neuronal glycolysis, resulting in aggravating OGD/R-induced neuronal apoptosis. Knockdown of YY1 or GAS5 protects against I/R-induced ischemic brain damage and improves overall neurological functions in vivo. Overall, YY1 interacts with LncRNA GAS5 to promote PFKFB3 transcription to enhance neuronal glycolysis, resulting in aggravating cerebral I/R injury.
Asunto(s)
Isquemia Encefálica/genética , Glucosa/metabolismo , Glucólisis/genética , Neuronas/metabolismo , Fosfofructoquinasa-2/genética , ARN Largo no Codificante/genética , Daño por Reperfusión/genética , Factor de Transcripción YY1/genética , Animales , Apoptosis/genética , Isquemia Encefálica/metabolismo , Corteza Cerebral/citología , Inmunoprecipitación de Cromatina , Inmunoprecipitación , Etiquetado Corte-Fin in Situ , Masculino , Ratones , Cultivo Primario de Células , ARN Largo no Codificante/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Daño por Reperfusión/metabolismo , Regulación hacia Arriba , Factor de Transcripción YY1/metabolismoRESUMEN
Skeletal muscle satellite cells (SCs) are adult muscle stem cells responsible for muscle regeneration after acute or chronic injuries. The lineage progression of quiescent SC toward activation, proliferation, and differentiation during the regeneration is orchestrated by cascades of transcription factors (TFs). Here, we elucidate the function of TF Yin Yang1 (YY1) in muscle regeneration. Muscle-specific deletion of YY1 in embryonic muscle progenitors leads to severe deformity of diaphragm muscle formation, thus neonatal death. Inducible deletion of YY1 in SC almost completely blocks the acute damage-induced muscle repair and exacerbates the chronic injury-induced dystrophic phenotype. Examination of SC revealed that YY1 loss results in cell-autonomous defect in activation and proliferation. Mechanistic search revealed that YY1 binds and represses mitochondrial gene expression. Simultaneously, it also stabilizes Hif1α protein and activates Hif1α-mediated glycolytic genes to facilitate a metabolic reprogramming toward glycolysis which is needed for SC proliferation. Altogether, our findings have identified YY1 as a key regulator of SC metabolic reprogramming through its dual roles in modulating both mitochondrial and glycolytic pathways.
Asunto(s)
Reprogramación Celular/genética , Músculo Esquelético/fisiología , Regeneración/genética , Células Satélite del Músculo Esquelético/fisiología , Factor de Transcripción YY1/fisiología , Animales , Diferenciación Celular/genética , Células Cultivadas , Glucólisis/genética , Células HEK293 , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Mitocondrias Musculares/genética , Mitocondrias Musculares/metabolismo , Desarrollo de Músculos/genética , Cicatrización de Heridas/genéticaRESUMEN
How Mycobacterium tuberculosis (Mtb) rewires macrophage energy metabolism to facilitate survival is poorly characterized. Here, we used extracellular flux analysis to simultaneously measure the rates of glycolysis and respiration in real time. Mtb infection induced a quiescent energy phenotype in human monocyte-derived macrophages and decelerated flux through glycolysis and the TCA cycle. In contrast, infection with the vaccine strain, M. bovis BCG, or dead Mtb induced glycolytic phenotypes with greater flux. Furthermore, Mtb reduced the mitochondrial dependency on glucose and increased the mitochondrial dependency on fatty acids, shifting this dependency from endogenous fatty acids in uninfected cells to exogenous fatty acids in infected macrophages. We demonstrate how quantifiable bioenergetic parameters of the host can be used to accurately measure and track disease, which will enable rapid quantifiable assessment of drug and vaccine efficacy. Our findings uncover new paradigms for understanding the bioenergetic basis of host metabolic reprogramming by Mtb.
Asunto(s)
Ciclo del Ácido Cítrico/genética , Ácidos Grasos/metabolismo , Glucosa/metabolismo , Glucólisis/genética , Interacciones Huésped-Patógeno , Macrófagos/microbiología , Mycobacterium tuberculosis/metabolismo , Diferenciación Celular/efectos de los fármacos , Respiración de la Célula , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Macrófagos/metabolismo , Metaboloma , Mitocondrias/metabolismo , Mycobacterium bovis/crecimiento & desarrollo , Mycobacterium bovis/metabolismo , Mycobacterium tuberculosis/crecimiento & desarrollo , Células THP-1 , Acetato de Tetradecanoilforbol/farmacologíaRESUMEN
Fatty acid amide hydrolase (FAAH) is an important enzyme for lipid metabolism and an interesting pharmacological target, given its role in anandamide breakdown. The FAAH-/- genotype is the most widely used mouse model to investigate the effects of a complete pharmacological inhibition of this enzyme. In this paper, we explore, by means of label-free SWATH proteomics, the changes in protein expression occurring in the liver of FAAH-/- knockout (KO) mice. We identified several altered biological processes and pathways, like fatty acid synthesis and glycolysis, which explain the observed phenotype of this mouse. We also observed the alteration of other proteins, like carboxylesterases and S-methyltransferases, apparently not immediately related to FAAH, but known to have important biological roles. Our study, reporting more than 3000 quantified proteins, offers an in-depth analysis of the liver proteome of this model.