RESUMEN
Methionine is important for intestinal development and homeostasis in various organisms. However, the underlying mechanisms are poorly understood. Here, we demonstrate that the methionine adenosyltransferase gene Mat2a is essential for intestinal development and that the metabolite S-adenosyl-L-methionine (SAM) plays an important role in intestinal homeostasis. Intestinal epithelial cell (IEC)-specific knockout of Mat2a exhibits impaired intestinal development and neonatal lethality. Mat2a deletion in the adult intestine reduces cell proliferation and triggers IEC apoptosis, leading to severe intestinal epithelial atrophy and intestinal inflammation. Mechanistically, we reveal that SAM maintains the integrity of differentiated epithelium and protects IECs from apoptosis by suppressing the expression of caspases 3 and 8 and their activation. SAM supplementation improves the defective intestinal epithelium and reduces inflammatory infiltration sequentially. In conclusion, our study demonstrates that methionine metabolism and its intermediate metabolite SAM play essential roles in intestinal development and homeostasis in mice.
Asunto(s)
Metionina Adenosiltransferasa , S-Adenosilmetionina , Ratones , Animales , S-Adenosilmetionina/metabolismo , Metionina Adenosiltransferasa/genética , Metionina Adenosiltransferasa/metabolismo , Mucosa Intestinal/metabolismo , Metionina , Suplementos DietéticosRESUMEN
Folic acid, served as dietary supplement, is closely linked to one-carbon metabolism and methionine metabolism. Previous clinical evidence indicated that folic acid supplementation displays dual effect on cancer development, promoting or suppressing tumor formation and progression. However, the underlying mechanism remains to be uncovered. Here, we report that high-folate diet significantly promotes cancer development in mice with hepatocellular carcinoma (HCC) induced by DEN/high-fat diet (HFD), simultaneously with increased expression of methionine adenosyltransferase 2A (gene name, MAT2A; protein name, MATIIα), the key enzyme in methionine metabolism, and acceleration of methionine cycle in cancer tissues. In contrast, folate-free diet reduces MATIIα expression and impedes HFD-induced HCC development. Notably, methionine metabolism is dynamically reprogrammed with valosin-containing protein p97/p47 complex-interacting protein (VCIP135) which functions as a deubiquitylating enzyme to bind and stabilize MATIIα in response to folic acid signal. Consistently, upregulation of MATIIα expression is positively correlated with increased VCIP135 protein level in human HCC tissues compared to adjacent tissues. Furthermore, liver-specific knockout of Mat2a remarkably abolishes the advocating effect of folic acid on HFD-induced HCC, demonstrating that the effect of high or free folate-diet on HFD-induced HCC relies on Mat2a. Moreover, folate and multiple intermediate metabolites in one-carbon metabolism are significantly decreased in vivo and in vitro upon Mat2a deletion. Together, folate promotes the integration of methionine and one-carbon metabolism, contributing to HCC development via hijacking MATIIα metabolic pathway. This study provides insight into folate-promoted cancer development, strongly recommending the tailor-made folate supplement guideline for both sub-healthy populations and patients with cancer expressing high level of MATIIα expression.
Asunto(s)
Ácido Fólico , Metionina Adenosiltransferasa , Animales , Dieta , Ácido Fólico/farmacología , Neoplasias Hepáticas Experimentales/genética , Neoplasias Hepáticas Experimentales/metabolismo , Neoplasias Hepáticas Experimentales/patología , Metionina/metabolismo , Metionina Adenosiltransferasa/genética , Metionina Adenosiltransferasa/metabolismo , RatonesRESUMEN
Common buckwheat (Fagopyrum esculentum Moench), a pseudocereal crop, produces a large number of flowers, but this does not guarantee high seed yields. This species demonstrates strong abortion of flowers and embryos. High temperatures during the generative growth phase result in an increase in the degeneration of embryo sacs. The aim of this study was to investigate proteomic changes in flowers and leaves of two common buckwheat accessions with different degrees of heat tolerance, Panda and PA15. Two-dimensional gel electrophoresis and mass spectrometry techniques were used to analyze the proteome profiles. Analyses were conducted for flower buds, open flowers capable of fertilization, and wilted flowers, as well as donor leaves, i.e., those growing closest to the inflorescences. High temperature up-regulated the expression of 182 proteins. The proteomic response to heat stress differed between the accessions and among their organs. In the Panda accession, we observed a change in abundance of 17, 13, 28, and 11 proteins, in buds, open and wilted flowers, and leaves, respectively. However, in the PA15 accession there were 34, 21, 63, and 21 such proteins, respectively. Fifteen heat-affected proteins were common to both accessions. The indole-3-glycerol phosphate synthase chloroplastic-like isoform X2 accumulated in the open flowers of the heat-sensitive cultivar Panda in response to high temperature, and may be a candidate protein as a marker of heat sensitivity in buckwheat plants.
Asunto(s)
Fagopyrum/metabolismo , Flores/metabolismo , Regulación de la Expresión Génica de las Plantas , Hojas de la Planta/metabolismo , Proteoma , Termotolerancia/genética , Electroforesis en Gel Bidimensional , Fagopyrum/embriología , Fagopyrum/genética , Fagopyrum/crecimiento & desarrollo , Respuesta al Choque Térmico/genética , Calor , Indol-3-Glicerolfosfato Sintasa/biosíntesis , Indol-3-Glicerolfosfato Sintasa/genética , Metionina Adenosiltransferasa/biosíntesis , Metionina Adenosiltransferasa/genética , Proteínas de Plantas/biosíntesis , Proteínas de Plantas/genética , Espectrometría de Masas en Tándem , Regulación hacia ArribaRESUMEN
Methionine metabolism is critical for epigenetic maintenance, redox homeostasis, and animal development. However, the regulation of methionine metabolism remains unclear. Here, we provide evidence that SIRT1, the most conserved mammalian NAD+-dependent protein deacetylase, is critically involved in modulating methionine metabolism, thereby impacting maintenance of mouse embryonic stem cells (mESCs) and subsequent embryogenesis. We demonstrate that SIRT1-deficient mESCs are hypersensitive to methionine restriction/depletion-induced differentiation and apoptosis, primarily due to a reduced conversion of methionine to S-adenosylmethionine. This reduction markedly decreases methylation levels of histones, resulting in dramatic alterations in gene expression profiles. Mechanistically, we discover that the enzyme converting methionine to S-adenosylmethionine in mESCs, methionine adenosyltransferase 2a (MAT2a), is under control of Myc and SIRT1. Consistently, SIRT1 KO embryos display reduced Mat2a expression and histone methylation and are sensitive to maternal methionine restriction-induced lethality, whereas maternal methionine supplementation increases the survival of SIRT1 KO newborn mice. Our findings uncover a novel regulatory mechanism for methionine metabolism and highlight the importance of methionine metabolism in SIRT1-mediated mESC maintenance and embryonic development.
Asunto(s)
Desarrollo Embrionario/genética , Epigénesis Genética , Metionina Adenosiltransferasa/genética , Metionina/metabolismo , Células Madre Embrionarias de Ratones/metabolismo , Sirtuina 1/genética , Acetilación , Animales , Apoptosis , Diferenciación Celular , Embrión de Mamíferos , Histonas/genética , Histonas/metabolismo , Metabolómica , Metionina/administración & dosificación , Metionina Adenosiltransferasa/metabolismo , Metilación , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Análisis por Micromatrices , Células Madre Embrionarias de Ratones/citología , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , S-Adenosilmetionina/metabolismo , Sirtuina 1/deficienciaRESUMEN
S-Adenosylmethionine is widely used in a variety of biological reactions and participates in the methionine (Met) metabolic pathway. In Arabidopsis (Arabidopsis thaliana), one of the four S-adenosylmethionine synthetase genes, METHIONINE ADENOSYLTRANSFERASE3 (MAT3), is highly expressed in pollen. Here, we show that mat3 mutants have impaired pollen tube growth and reduced seed set. Metabolomics analyses confirmed that mat3 pollen and pollen tubes overaccumulate Met and that mat3 pollen has several metabolite profiles, such as those of polyamine biosynthesis, which are different from those of the wild type. Additionally, we show that disruption of Met metabolism in mat3 pollen affected transfer RNA and histone methylation levels. Thus, our results suggest a connection between metabolism and epigenetics.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimología , Metionina Adenosiltransferasa/metabolismo , Tubo Polínico/enzimología , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Histonas/metabolismo , Metabolómica/métodos , Metionina/metabolismo , Metionina Adenosiltransferasa/genética , Metilación , Microscopía Fluorescente , Mutación , Plantas Modificadas Genéticamente , Polen/enzimología , Polen/genética , Polen/crecimiento & desarrollo , Tubo Polínico/genética , Tubo Polínico/crecimiento & desarrollo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , S-Adenosilmetionina/metabolismo , Semillas/genética , Semillas/crecimiento & desarrollo , Semillas/metabolismoRESUMEN
Metabolizable methionine (Met) concentrations can be increased by feeding rumen-protected dl-Met or the isopropyl ester of 2-hydroxy-4-(methylthio) butanoic acid (HMBi). Hepatic responses to increasing concentrations of metabolizable Met as a result of supplementation of different Met sources have not been comparatively examined. The objective of this experiment was to examine the regulation of key genes for Met metabolism, gluconeogenesis, and fatty acid oxidation in response to increasing concentrations of dl-Met or 2-hydroxy-4-(methylthio) butanoic acid (HMB) in bovine primary hepatocytes. Hepatocytes isolated from 4 Holstein calves less than 7d old were maintained as monolayer cultures for 24h before addition of treatments. Cells were then exposed to treatments of dl-Met or HMB (0, 10, 20, 40, or 60 µM) in Met-free medium for 24h and collected for RNA isolation and quantification of gene expression by quantitative PCR. Expression of betaine-homocysteine methyltransferase (BHMT), 5-methyltetrahydrofolate-homocysteine methyltransferase (MTR), and 5,10 methylenetetrahydrofolate reductase (MTHFR) genes, which catalyze regeneration of Met from betaine and homocysteine, decreased linearly with increasing dl-Met concentration. We observed similar effects with increasing HMB concentration, except expression of MTHFR, which was not altered. Expression of Met adenosyltransferase 1A (MAT1A), which catalyzes the first step of Met metabolism to generate S-adenosylmethionine (SAM), a primary methyl donor, was decreased with increasing dl-Met or HMB concentration. Expression of S-adenosylhomocysteine hydrolase (SAHH) was decreased linearly with increasing HMB concentration, but not altered by dl-Met. Increasing concentrations of dl-Met and HMB decreased cytosolic phosphoenolpyruvate carboxykinase (PCK1) expression, but did not alter the expression of mitochondrial phosphoenolpyruvate carboxykinase (PCK2) or pyruvate carboxylase (PC). Expression of glucose-6-phosphatase(G6PC) decreased linearly with increasing HMB concentration, but not altered by dl-Met. Neither dl-Met nor HMB altered the expression of carnitine palmitoyltransferase 1A(CPT1a). These findings demonstrate reduced necessity for Met regeneration with increased Met concentrations in the medium, regardless of the Met source. The lack of upregulation of gluconeogenesis indicates that increased dl-Met or HMB is not prioritized for glucose synthesis in primary bovine hepatocytes.
Asunto(s)
Hígado/efectos de los fármacos , Metionina/análogos & derivados , Metionina/farmacología , 5-Metiltetrahidrofolato-Homocisteína S-Metiltransferasa/genética , Adenosilhomocisteinasa/genética , Animales , Animales Recién Nacidos , Betaína/metabolismo , Betaína-Homocisteína S-Metiltransferasa/genética , Carnitina O-Palmitoiltransferasa/genética , Bovinos , Regulación hacia Abajo , Gluconeogénesis/genética , Glucosa-6-Fosfatasa/genética , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Homocisteína/metabolismo , Hígado/metabolismo , Metionina Adenosiltransferasa/genética , Metilenotetrahidrofolato Reductasa (NADPH2)/genética , Fosfoenolpiruvato Carboxiquinasa (GTP)/genética , S-Adenosilmetionina/metabolismo , Regulación hacia ArribaRESUMEN
As an important biological methyl group donor, S-adenosyl-L-methionine is used as nutritional supplement or drug for various diseases, but bacterial strains that can efficiently produce S-adenosyl-L-methionine are not available. In this study, Corynebacterium glutamicum strain HW104 which can accumulate S-adenosyl-L-methionine was constructed from C. glutamicum ATCC13032 by deleting four genes thrB, metB, mcbR and Ncgl2640, and six genes metK, vgb, lysC(m), hom(m), metX and metY were overexpressed in HW104 in different combinations, forming strains HW104/pJYW-4-metK-vgb, HW104/pJYW-4-SAM2C-vgb, HW104/pJYW-4-metK-vgb-metYX, and HW104/pJYW-4-metK-vgb-metYX-hom(m)-lysC(m). Fermentation experiments showed that HW104/pJYW-4-metK-vgb produced more S-adenosyl-L-methionine than other strains, and the yield achieved 196.7 mg/L (12.15 mg/g DCW) after 48h. The results demonstrate the potential application of C. glutamicum for production of S-adenosyl-L-methionine without addition of L-methionine.
Asunto(s)
Corynebacterium glutamicum/metabolismo , S-Adenosilmetionina/biosíntesis , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Vías Biosintéticas , Corynebacterium glutamicum/genética , Fermentación , Eliminación de Gen , Genes Bacterianos , Ingeniería Metabólica/métodos , Metionina/metabolismo , Metionina Adenosiltransferasa/genéticaRESUMEN
RNA-Seq has proven to be a very powerful tool in the analysis of the Plum pox virus (PPV, sharka disease)/Prunus interaction. This technique is an important complementary tool to other means of studying genomics. In this work an analysis of gene expression of resistance/susceptibility to PPV in apricot is performed. RNA-Seq has been applied to analyse the gene expression changes induced by PPV infection in leaves from two full-sib apricot genotypes, "Rojo Pasión" and "Z506-7", resistant and susceptible to PPV, respectively. Transcriptomic analyses revealed the existence of more than 2,000 genes related to the pathogen response and resistance to PPV in apricot. These results showed that the response to infection by the virus in the susceptible genotype is associated with an induction of genes involved in pathogen resistance such as the allene oxide synthase, S-adenosylmethionine synthetase 2 and the major MLP-like protein 423. Over-expression of the Dicer protein 2a may indicate the suppression of a gene silencing mechanism of the plant by PPV HCPro and P1 PPV proteins. On the other hand, there were 164 genes involved in resistance mechanisms that have been identified in apricot, 49 of which are located in the PPVres region (scaffold 1 positions from 8,050,804 to 8,244,925), which is responsible for PPV resistance in apricot. Among these genes in apricot there are several MATH domain-containing genes, although other genes inside (Pleiotropic drug resistance 9 gene) or outside (CAP, Cysteine-rich secretory proteins, Antigen 5 and Pathogenesis-related 1 protein; and LEA, Late embryogenesis abundant protein) PPVres region could also be involved in the resistance.
Asunto(s)
Regulación de la Expresión Génica de las Plantas/inmunología , Genes de Plantas , Interacciones Huésped-Patógeno/genética , Enfermedades de las Plantas/genética , Virus Eruptivo de la Ciruela/fisiología , Prunus armeniaca/genética , Prunus domestica/genética , Transportadoras de Casetes de Unión a ATP/genética , Transportadoras de Casetes de Unión a ATP/inmunología , Susceptibilidad a Enfermedades , Pleiotropía Genética , Genotipo , Interacciones Huésped-Patógeno/inmunología , Oxidorreductasas Intramoleculares/genética , Oxidorreductasas Intramoleculares/inmunología , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/inmunología , Metionina Adenosiltransferasa/genética , Metionina Adenosiltransferasa/inmunología , Anotación de Secuencia Molecular , Enfermedades de las Plantas/inmunología , Enfermedades de las Plantas/virología , Inmunidad de la Planta/genética , Hojas de la Planta/genética , Hojas de la Planta/inmunología , Hojas de la Planta/virología , Proteínas de Plantas/genética , Proteínas de Plantas/inmunología , Virus Eruptivo de la Ciruela/patogenicidad , Prunus armeniaca/inmunología , Prunus armeniaca/virología , Prunus domestica/inmunología , Prunus domestica/virología , Ribonucleasa III/genética , Ribonucleasa III/inmunología , Transcriptoma/inmunologíaRESUMEN
S-adenosyl-L-methionine (SAM) synthase (SAMS) catalyze the biosynthesis of SAM, which is a precursor for ethylene and polyamines, and a methyl donor for a number of biomolecules. A full-length cDNA of SAMS from Solanum brevidens was expressed in Arabidopsis thaliana to study its physiological function. RT-PCR analysis showed that SbSAMS expression was enhanced significantly in S. brevidens leaves upon treatment with salt, mannitol, ethephon, IAA and ABA. The transgenic SbSAMS overexpression lines accumulated higher levels S-adenosyl homocysteine (SAHC) and ethylene concomitantly with increased SAM level. Expression levels of genes related to ethylene biosynthesis such as ACC synthase, but not polyamine biosynthesis genes were enhanced in SbSAMS overexpressing Arabidopsis lines. In addition, ABA responsive, wound and pathogen-inducible genes were upregulated in SbSAMS transgenic Arabidopsis plants. Transgenic Arabidopsis lines exhibited higher salt and drought stress tolerance compared to those of vector control. Based on these results we conclude that SbSAMS is expressed under abiotic stress to produce SAM as a broad-spectrum signal molecule to upregulate stress-related genes including ethylene and ABA biosynthetic pathway genes responsible for ABA, pathogen and wound responses.
Asunto(s)
Arabidopsis/crecimiento & desarrollo , Metionina Adenosiltransferasa/biosíntesis , Proteínas de Plantas/biosíntesis , Plantas Modificadas Genéticamente/crecimiento & desarrollo , Solanum/genética , Estrés Fisiológico , Arabidopsis/genética , Metionina Adenosiltransferasa/genética , Presión Osmótica , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente/genética , Solanum/enzimologíaRESUMEN
S-Adenosylmethionine synthetase (SAMS) catalyzes the synthesis of S-adenosylmethionine (SAM), a precursor for ethylene and polyamine biosynthesis. Here, we report the isolation of the 1498 bp full-length cDNA sequence encoding tetraploid black locust (Robinia pseudoacacia L.) SAMS (TrbSAMS), which contains an open reading frame of 1179 bp encoding 392 amino acids. The amino acid sequence of TrbSAMS has more than 94% sequence identity to SAMSs from other plants, with a closer phylogenetic relationship to SAMSs from legumes than to SAMS from other plants. The TrbSAMS monomer consists of N-terminal, central, and C-terminal domains. Subcellular localization analysis revealed that the TrbSAMS protein localizes mainly to in the cell membrane and cytoplasm of onion epidermal cells and Arabidopsis mesophyll cell protoplasts. Indole-3-butyric acid (IBA)-treated cuttings showed higher levels of TrbSAMS transcript than untreated control cuttings during root primordium and adventitious root formation. TrbSAMS and its downstream genes showed differential expression in shoots, leaves, bark, and roots, with the highest expression observed in bark. IBA-treated cuttings also showed higher SAMS activity than control cuttings during root primordium and adventitious root formation. These results indicate that TrbSAMS might play an important role in the regulation of IBA-induced adventitious root development in tetraploid black locust cuttings.
Asunto(s)
Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Genes de Plantas , Indoles/farmacología , Metionina Adenosiltransferasa/genética , Raíces de Plantas/crecimiento & desarrollo , Robinia/genética , Tetraploidía , Secuencia de Aminoácidos , Clonación Molecular , Etilenos/metabolismo , Metionina Adenosiltransferasa/química , Metionina Adenosiltransferasa/aislamiento & purificación , Metionina Adenosiltransferasa/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/genética , Poliaminas/metabolismo , Estructura Secundaria de Proteína , Robinia/anatomía & histología , Robinia/efectos de los fármacos , Robinia/enzimología , Alineación de Secuencia , Análisis de Secuencia de Proteína , Fracciones Subcelulares/enzimologíaRESUMEN
Methionine adenosyltransferase (MAT) is an essential enzyme that is responsible for the biosynthesis of S-adenosylmethionine (SAMe), the principal methyl donor and precursor of polyamines. MAT1A is expressed in normal liver and MAT2A is expressed in all extrahepatic tissues. MAT2A expression is increased in human colon cancer and in colon cancer cells treated with mitogens, whereas silencing MAT2A resulted in apoptosis. The aim of the current work was to examine the mechanism responsible for MAT2A-dependent growth and apoptosis. We found that in RKO (human adenocarcinoma cell line) cells, MAT2A siRNA treatment lowered cellular SAMe and putrescine levels by 70-75%, increased apoptosis and inhibited growth. Putrescine supplementation blunted significantly MAT2A siRNA-induced apoptosis and growth suppression. Putrescine treatment (100pmol/L) raised MAT2A mRNA level to 4.3-fold of control, increased the expression of c-Jun and c-Fos and binding to an AP-1 site in the human MAT2A promoter and the promoter activity. In human colon cancer specimens, the expression levels of MAT2A, ornithine decarboxylase (ODC), c-Jun and c-Fos are all elevated as compared to adjacent non-tumorous tissues. Overexpression of ODC in RKO cells also raised MAT2A mRNA level and MAT2A promoter activity. ODC and MAT2A are also overexpressed in liver cancer and consistently, similar MAT2A-ODC-putrescine interactions and effects on growth and apoptosis were observed in HepG2 cells. In conclusion, there is a crosstalk between polyamines and MAT2A. Increased MAT2A expression provides more SAMe for polyamines biosynthesis; increased polyamine (putrescine in this case) can activate MAT2A at the transcriptional level. This along with increased ODC expression in cancer all feed forward to further enhance the proliferative capacity of the cancer cell.
Asunto(s)
Neoplasias del Colon/metabolismo , Neoplasias Hepáticas/metabolismo , Metionina Adenosiltransferasa/metabolismo , Putrescina/metabolismo , Anciano , Apoptosis , Línea Celular Tumoral , Proliferación Celular , Neoplasias del Colon/enzimología , Femenino , Humanos , Neoplasias Hepáticas/enzimología , Masculino , Metionina Adenosiltransferasa/genética , Persona de Mediana Edad , Ornitina Descarboxilasa/genética , Ornitina Descarboxilasa/metabolismo , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas c-fos/genética , Proteínas Proto-Oncogénicas c-fos/metabolismo , Proteínas Proto-Oncogénicas c-jun/genética , Proteínas Proto-Oncogénicas c-jun/metabolismo , ARN Interferente Pequeño , S-Adenosilmetionina/metabolismo , Factor de Transcripción AP-1/metabolismo , Transcripción GenéticaRESUMEN
Methionine adenosyltransferase I/III (MAT I/III) deficiency, caused by mutations in the MAT1A gene, is an inherited metabolic disorder characterized by persistent hypermethioninemia, usually detected by newborn mass screening. There is a wide range of clinical manifestations, from completely asymptomatic to neurological problems associated with brain demyelination. Physiological role of S-adenosylmethionine (SAM), the metabolic product of methionine catalyzed by MAT, in the central nervous system has been investigated in vivo and in vitro, and case reports demonstrated an effectiveness of supplementary treatment of SAM in the improvement of neurological development and myelination. Methionine restriction can be an additional therapeutic strategy because hypermethioninemia alone may be neurotoxic; however, lowering methionine carries a risk to decrease the synthesis of SAM.
Asunto(s)
Errores Innatos del Metabolismo de los Aminoácidos/metabolismo , Encéfalo/metabolismo , Enfermedades Desmielinizantes/metabolismo , Metionina Adenosiltransferasa/metabolismo , Metionina/metabolismo , S-Adenosilmetionina/metabolismo , Alelos , Errores Innatos del Metabolismo de los Aminoácidos/dietoterapia , Errores Innatos del Metabolismo de los Aminoácidos/genética , Errores Innatos del Metabolismo de los Aminoácidos/patología , Encéfalo/patología , Enfermedades Desmielinizantes/dietoterapia , Enfermedades Desmielinizantes/genética , Enfermedades Desmielinizantes/patología , Dieta , Pruebas Genéticas , Glicina N-Metiltransferasa/deficiencia , Humanos , Recién Nacido , Isoenzimas/genética , Isoenzimas/metabolismo , Metionina Adenosiltransferasa/deficiencia , Metionina Adenosiltransferasa/genética , Mutación , S-Adenosilmetionina/uso terapéutico , Índice de Severidad de la EnfermedadRESUMEN
Reported is a female patient with methionine adenosyltransferase I/III (MAT I/III) deficiency, who was found to have pronounced hypermethioninemia on newborn mass spectroscopy screening, and had two compound heterozygous missense mutations in the gene encoding human MAT1A protein. Hypermethioninemia persisted and her mental development was deficient. At 4 years and 8 months, we started with the supplementary treatment of S-adenosylmethionine, the metabolic product of methionine catalyzed by MAT, which was effective in her neurological development.
Asunto(s)
Errores Innatos del Metabolismo de los Aminoácidos/tratamiento farmacológico , Metionina Adenosiltransferasa/deficiencia , Metionina Adenosiltransferasa/genética , S-Adenosilmetionina/uso terapéutico , Errores Innatos del Metabolismo de los Aminoácidos/diagnóstico , Errores Innatos del Metabolismo de los Aminoácidos/genética , Niño , Femenino , Humanos , Metionina/deficiencia , Metionina/metabolismo , Mutación MissenseRESUMEN
Methionine S-adenosyltransferase (MAT) catalyzes the only reaction that produces the major methyl donor in mammals. Low-dose methotrexate is the most commonly used disease-modifying antirheumatic drug in human rheumatic conditions. The present study was conducted to test the hypothesis that methotrexate inhibits MAT expression and activity in vitro and in vivo. HepG2 cells were cultured under folate restriction or in low-dose methotrexate with and without folate or methionine supplementation. Male C57BL/6J mice received methotrexate regimens that reflected low-dose clinical use in humans. S-adenosylmethionine and MAT genes, proteins and enzyme activity levels were determined. We found that methionine or folate supplementation greatly improved S-adenosylmethionine in folate-depleted cells but not in cells preexposed to methotrexate. Methotrexate but not folate depletion suppressed MAT genes, proteins and activity in vitro. Low-dose methotrexate inhibited MAT1A and MAT2A genes, MATI/II/III proteins and MAT enzyme activities in mouse tissues. Concurrent folinate supplementation with methotrexate ameliorated MAT2A reduction and restored S-adenosylmethionine in HepG2 cells. However, posttreatment folinate rescue failed to restore MAT2A reduction or S-adenosylmethionine level in cells preexposed to methotrexate. Our results provide both in vitro and in vivo evidence that low-dose methotrexate inhibits MAT genes, proteins, and enzyme activity independent of folate depletion. Because polyglutamated methotrexate stays in the hepatocytes, if methotrexate inhibits MAT in the liver, then the efficacy of clinical folinate rescue with respect to maintaining hepatic S-adenosylmethionine synthesis and normalizing the methylation reactions would be limited. These findings raise concerns on perturbed methylation reactions in humans on low-dose methotrexate. Future studies on the clinical physiological consequences of MAT inhibition by methotrexate and the potential benefits of S-adenosylmethionine supplementation on methyl group homeostasis in clinical methotrexate therapies are warranted.
Asunto(s)
Antirreumáticos/farmacología , Metionina Adenosiltransferasa/antagonistas & inhibidores , Metotrexato/farmacología , Animales , Dactinomicina , Dexametasona , Inhibidores Enzimáticos/farmacología , Ácido Fólico/metabolismo , Ácido Fólico/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Células Hep G2 , Humanos , Masculino , Metionina/farmacología , Metionina Adenosiltransferasa/genética , Metionina Adenosiltransferasa/metabolismo , Ratones , Ratones Endogámicos C57BL , ARN Mensajero/metabolismo , S-Adenosilmetionina/metabolismoRESUMEN
Tissues of the mucosa are lined by an epithelium that provides barrier and transport functions. It is now appreciated that inflammatory responses in inflammatory bowel diseases are accompanied by striking shifts in tissue metabolism. In this paper, we examined global metabolic consequences of mucosal inflammation using both in vitro and in vivo models of disease. Initial analysis of the metabolic signature elicited by inflammation in epithelial models and in colonic tissue isolated from murine colitis demonstrated that levels of specific metabolites associated with cellular methylation reactions are significantly altered by model inflammatory systems. Furthermore, expression of enzymes central to all cellular methylation, S-adenosylmethionine synthetase and S-adenosylhomocysteine hydrolase, are increased in response to inflammation. Subsequent studies showed that DNA methylation is substantially increased during inflammation and that epithelial NF-κB activity is significantly inhibited following treatment with a reversible S-adenosylhomocysteine hydrolase inhibitor, DZ2002. Finally, these studies demonstrated that inhibition of cellular methylation in a murine model of colitis results in disease exacerbation while folate supplementation to promote methylation partially ameliorates the severity of murine colitis. Taken together, these results identify a global change in methylation, which during inflammation, translates to an overall protective role in mucosal epithelia.
Asunto(s)
Colitis/metabolismo , Inflamación/metabolismo , Mucosa Intestinal/metabolismo , Metabolómica/métodos , Adenina/análogos & derivados , Adenina/farmacología , Adenosilhomocisteinasa/genética , Adenosilhomocisteinasa/metabolismo , Animales , Western Blotting , Butiratos/farmacología , Línea Celular Tumoral , Colitis/genética , Colon/efectos de los fármacos , Colon/metabolismo , Colon/patología , Metilación de ADN/efectos de los fármacos , Sulfato de Dextran/farmacología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Perfilación de la Expresión Génica/métodos , Células HeLa , Humanos , Inflamación/genética , Interferón gamma/metabolismo , Interferón gamma/farmacología , Mucosa Intestinal/patología , Espectroscopía de Resonancia Magnética , Metionina Adenosiltransferasa/genética , Metionina Adenosiltransferasa/metabolismo , Metilación/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Mucositis/genética , Mucositis/metabolismo , FN-kappa B/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The S(MK) (SAM-III) box is an S-adenosylmethionine (SAM)-responsive riboswitch found in the 5' untranslated region of metK genes, encoding SAM synthetase, in many members of the Lactobacillales. SAM binding causes a structural rearrangement in the RNA that sequesters the Shine-Dalgarno (SD) sequence by pairing with a complementary anti-SD (ASD) sequence; sequestration of the SD sequence inhibits binding of the 30S ribosomal subunit and prevents translation initiation. We observed a slight increase in the half-life of the metK transcript in vivo when Enterococcus faecalis cells were depleted for SAM, but no significant change in overall transcript abundance, consistent with the model that this riboswitch regulates at the level of translation initiation. The half-life of the SAM-S(MK) box RNA complex in vitro is shorter than that of the metK transcript in vivo, raising the possibility of reversible binding of SAM. We used a fluorescence assay to directly visualize reversible switching between the SAM-free and SAM-bound conformations. We propose that the S(MK) box riboswitch can make multiple SAM-dependent regulatory decisions during the lifetime of the transcript in vivo, acting as a reversible switch that allows the cell to respond rapidly to fluctuations in SAM pools by modulating expression of the SAM synthetase gene.
Asunto(s)
Proteínas Bacterianas/genética , Enterococcus faecalis/enzimología , Regulación Enzimológica de la Expresión Génica , Metionina Adenosiltransferasa/genética , Elementos de Respuesta , Riboswitch , Regiones no Traducidas 5' , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Enterococcus faecalis/química , Enterococcus faecalis/genética , Enterococcus faecalis/metabolismo , Regulación Bacteriana de la Expresión Génica , Metionina Adenosiltransferasa/química , Metionina Adenosiltransferasa/metabolismo , Conformación de Ácido Nucleico , Estabilidad del ARN , ARN Bacteriano/química , ARN Bacteriano/genética , ARN Bacteriano/metabolismo , S-Adenosilmetionina/metabolismoRESUMEN
In a previous work it was shown that ethylene participates in the up-regulation of several Fe acquisition genes of Arabidopsis, such as AtFIT, AtFRO2, and AtIRT1. In this work the relationship between ethylene and Fe-related genes in Arabidopsis has been looked at in more depth. Genes induced by Fe deficiency regulated by ethylene were searched for. For this, studies were conducted, using microarray analysis and reverse transcription-PCR (RT-PCR), to determine which of the genes up-regulated by Fe deficiency are simultaneously suppressed by two different ethylene inhibitors (cobalt and silver thiosulphate), assessing their regulation by ethylene in additional experiments. In a complementary experiment, it was determined that the Fe-related genes up-regulated by ethylene were also responsive to nitric oxide (NO). Further studies were performed to analyse whether Fe deficiency up-regulates the expression of genes involved in ethylene biosynthesis [S-adenosylmethionine synthetase, 1-aminocyclopropane-1-carboxylate (ACC) synthase, and ACC oxidase genes] and signalling (AtETR1, AtCTR1, AtEIN2, AtEIN3, AtEIL1, and AtEIL3). The results obtained show that both ethylene and NO are involved in the up-regulation of many important Fe-regulated genes of Arabidopsis, such as AtFIT, AtbHLH38, AtbHLH39, AtFRO2, AtIRT1, AtNAS1, AtNAS2, AtFRD3, AtMYB72, and others. In addition, the results show that Fe deficiency up-regulates genes involved in both ethylene synthesis (AtSAM1, AtSAM2, AtACS4, AtACS6, AtACS9, AtACO1, and AtACO2) and signalling (AtETR1, AtCTR1, AtEIN2, AtEIN3, AtEIL1, and AtEIL3) in the roots.
Asunto(s)
Arabidopsis/genética , Etilenos/metabolismo , Regulación de la Expresión Génica de las Plantas , Hierro/metabolismo , Óxido Nítrico/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Genes de Plantas , Homeostasis/genética , Metionina Adenosiltransferasa/genética , Metionina Adenosiltransferasa/metabolismo , Raíces de Plantas/genética , Raíces de Plantas/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , S-Adenosilmetionina/genética , S-Adenosilmetionina/metabolismo , Activación Transcripcional , Regulación hacia ArribaRESUMEN
OBJECTIVE: The objective of this study was to investigate the regulatory effect of fish oil rich in omega-3 polyunsaturated fatty acids (PUFAs) on critical enzyme activity and mRNA expression involved in homocysteine (Hcy) metabolism. METHODS: Thirty-six male Sprague-Dawley rats aged 3 wk, weighing 120 +/- 10 g, were randomly divided into three groups: the olive oil (OO) group, the tuna oil (TO) group, and the salmon oil (SO) group. The oil was orally administered every day using a stomach tube. Eight weeks later, plasma Hcy, phospholipids, omega-3 PUFAs, enzyme activity, and mRNA expression in tissues were determined. RESULTS: Compared with the control group, phospholipids, total omega-3 PUFAs, and omega-3/omega-6 PUFAs in the liver and lung were significantly elevated in the TO and SO groups; 22:6omega-3 in the liver and lung was significantly increased in the TO group; and 20:5omega-3 in the two tissues was significantly elevated in the SO group. The level of plasma Hcy was significantly decreased with TO; methionine adenosyl transferase (MAT) activity was significantly increased and MAT mRNA expression was significantly upregulated with TO; cystathionine-gamma-lyase mRNA expression in TO was significantly upregulated; however, cystathionine beta-synthase and S-adenosylhomocysteine hydrolases were not significantly changed when compared with control. CONCLUSION: TO rich in 22:6omega-3 decreases the concentration of Hcy despite increasing MAT activity and upregulating MAT mRNA expression through compensatory cystathionine-gamma-lyase mRNA expression, both of which are involved in Hcy metabolism.
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Ácidos Docosahexaenoicos/farmacología , Enzimas/metabolismo , Expresión Génica/efectos de los fármacos , Homocisteína/sangre , Metabolismo de los Lípidos/efectos de los fármacos , Metionina/metabolismo , Adenosilhomocisteinasa/genética , Adenosilhomocisteinasa/metabolismo , Animales , Cistationina betasintasa/genética , Cistationina betasintasa/metabolismo , Cistationina gamma-Liasa/genética , Cistationina gamma-Liasa/metabolismo , Ácidos Docosahexaenoicos/administración & dosificación , Enzimas/genética , Aceites de Pescado/farmacología , Hígado/metabolismo , Pulmón/metabolismo , Masculino , Metionina/genética , Metionina Adenosiltransferasa/genética , Metionina Adenosiltransferasa/metabolismo , Aceite de Oliva , Aceites de Plantas , ARN Mensajero/metabolismo , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , Salmón , Atún , Regulación hacia ArribaRESUMEN
S'adenosyl-L: -methionine (SAM) is a ubiquitous methyl donor and a precursor in the biosynthesis of ethylene, polyamines, biotin, and nicotianamine in plants. Only limited information is available regarding its synthesis (SAM cycle) and its concentrations in plant tissues. The SAM concentrations in flowers of Nicotiana suaveolens were determined during day/night cycles and found to fluctuate rhythmically between 10 and 50 nmol g(-1) fresh weight. Troughs of SAM levels were measured in the evening and night, which corresponds to the time when the major floral scent compound, methyl benzoate, is synthesized by a SAM dependent methyltransferase (NsBSMT) and when this enzyme possesses its highest activity. The SAM synthetase (NsSAMS1) and methionine synthase (NsMS1) are enzymes, among others, which are involved in the synthesis and regeneration of SAM. Respective genes were isolated from a N. suaveolens petal cDNA library. Transcript accumulation patterns of both SAM regenerating enzymes matched perfectly those of the bifunctional NsBSMT; maximum mRNA accumulations of NsMS1 and NsSAMS1 were attained in the evening. Ethylene, which is synthesized from SAM, reached only low levels of 1-2 ppbv in N. suaveolens flowers. It is emitted in a burst at the end of the life span of the flowers, which correlates with the increased expression of the 1-aminocyclopropane-1-carboxylate oxidase (NsACO).
Asunto(s)
Etilenos/metabolismo , Flores/metabolismo , Nicotiana/metabolismo , Proteínas de Plantas/metabolismo , S-Adenosilmetionina/metabolismo , 5-Metiltetrahidrofolato-Homocisteína S-Metiltransferasa/clasificación , 5-Metiltetrahidrofolato-Homocisteína S-Metiltransferasa/genética , 5-Metiltetrahidrofolato-Homocisteína S-Metiltransferasa/metabolismo , Aminoácido Oxidorreductasas/clasificación , Aminoácido Oxidorreductasas/genética , Aminoácido Oxidorreductasas/metabolismo , Northern Blotting , Clonación Molecular , ADN Complementario/química , ADN Complementario/genética , Flores/genética , Regulación del Desarrollo de la Expresión Génica , Regulación Enzimológica de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Biblioteca de Genes , Metionina Adenosiltransferasa/clasificación , Metionina Adenosiltransferasa/genética , Metionina Adenosiltransferasa/metabolismo , Datos de Secuencia Molecular , Filogenia , Proteínas de Plantas/genética , Análisis de Secuencia de ADN , Factores de Tiempo , Nicotiana/genética , Nicotiana/crecimiento & desarrolloRESUMEN
In order to increase the yield of S-adenosylmethionine (SAM) in recombinant Pichia pastoris, a strategy of adding oxygen vectors and supplemental carbon sources was described. Three organic solutions were used as oxygen vectors for SAM accumulation at different concentrations and addition times. Firstly, n-hexane (0.5%) or n-heptane (1.0%) was added after 72 h of cultivation to improve SAM production. Carbon metabolism was scarce during the induction phase because of low methanol concentration. Secondly, sorbitol (1.2%), selected from three candidates (glycerol, lactic acid, and sorbitol), was used as the supplemental carbon source. The yield of SAM was improved significantly (53.26%) at 1.0%n-heptane added at 72 h (48 h induction), 1.2% sorbitol added at 72, 96, and 120 h of cultivation and 1.0% methanol added every 24 h during cultivation.