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Fausto G Hegardt - One of the best experts on this subject based on the ideXlab platform.
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Refining the diagnosis of mitochondrial HMG‐CoA Synthase deficiency
Journal of inherited metabolic disease, 2006Co-Authors: Rosa Aledo, Fausto G Hegardt, Nuria Casals, Juan Pié, Cecilia Mir, R. N. Dalton, Charles Turner, Michael ChampionAbstract:Mitochondrial HMG-CoA Synthase deficiency is an inherited metabolic disorder caused by a defect in the enzyme that regulates the formation of ketone bodies. Patients present with hypoketotic hypoglycaemia, encephalopathy and hepatomegaly, usually precipitated by an intercurrent infection or prolonged fasting. The diagnosis may easily be missed as previously reported results of routine metabolic investigations, urinary organic acids and plasma acylcarnitines may be nonspecific or normal, and a high index of suspicion is required to proceed to further confirmatory tests. We describe a further acute case in which the combination of urinary organic acids, low free carnitine and changes in the plasma acylcarnitine profile on carnitine supplementation were very suggestive of a defect in ketone synthesis. The diagnosis of mitochondrial HMG-CoA Synthase deficiency was confirmed on genotyping, revealing two novel mutations: c.614G > A (R188H) and c.971T > C (M307T). A further sibling, in whom the diagnosis had not been made acutely, was also found to be affected. The possible effects of these mutations on enzyme activity are discussed.
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The diagnosis of mitochondrial HMG-CoA Synthase deficiency☆
The Journal of pediatrics, 2002Co-Authors: Johannes Zschocke, Fausto G Hegardt, Nuria Casals, Juan Pié, Johannes M. Penzien, Rainer Bielen, Rosa Aledo, Georg F. Hoffmann, Ertan MayatepekAbstract:Deficiency of 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) Synthase, the only disorder exclusively affecting hepatic ketogenesis, is a cause of hypoglycemic coma. We report that the diagnosis can be made by typical laboratory findings (hypoketosis, elevated free fatty acids, normal acylcarnitines, specific urinary organic acids) during acute episodes.
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Genetic basis of mitochondrial HMG-CoA Synthase deficiency
Human Genetics, 2001Co-Authors: Rosa Aledo, Nuria Casals, Juan Pié, Johannes Zschocke, Georg F. Hoffmann, Ertan Mayatepek, Cecilia Mir, Sonja Fiesel, Fausto G HegardtAbstract:Deficiency of mitochondrial 3-hydroxy-3-methylglutaryl-CoA Synthase (mHMGS) is a recessive disorder of ketogenesis that has been previously diagnosed in two children with hypoglycaemic hypoketotic coma during fasting periods. Here, we report the results of molecular investigations in a third patient affected by this disease. Sequencing of the entire coding region of the HMGCS2 gene revealed two missense mutations, G212R and R500H. Mendelian inheritance was confirmed by the analysis of parental samples and neither of the mutations was found on 200 control chromosomes. Functional relevance was confirmed by in vitro expression studies in cytosolic HMGS-deficient cells. Whereas wild-type cDNA of the HMGCS2 gene reverted the auxotrophy for mevalonate, the cDNAs of the mutants did not. The disease may be recognised by specific clinical and biochemical features but it is difficult to confirm enzymatically since the gene is expressed only in liver and testis. Molecular studies may facilitate or confirm future diagnoses in affected patients.
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3-Hydroxy-3-methylglutaryl coenzyme A Synthase-1 of Blattella germanica has structural and functional features of an active retrogene
Insect Biochemistry and Molecular Biology, 2001Co-Authors: Nuria Casals, Pedro F. Marrero, Xavier Belles, Carlos Buesa, Fausto G HegardtAbstract:Blattella germanica has two cytosolic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) Synthase genes, HMG-CoA Synthase-1 and -2. HMG-CoA Synthase-1 gene shows several features of processed genes (retroposons): it contains no introns but has a short direct-repeat sequence (ATTATTATT) at both ends. An atypical feature is the presence at both ends of the gene of short inverse repeats flanked by direct repeats. There is neither a TATA box nor a CAAT box in the 5' region. Comparative analysis with other species suggests that the HMG-CoA Synthase-1 gene derives from HMG-CoA Synthase-2. Cultured embryonic B. germanica UM-BGE-1 cells express HMG-CoA Synthase-1 but not HMG-CoA Synthase-2, suggesting that the intron-less gene is functional. In addition, it can complement MEV-1 cell line, which is auxotrophic for mevalonate. We show that compactin and mevalonate do not significantly affect the mRNA levels of HMG-CoA Synthase-1 in UM-BGE-1 cells. Compactin induces a 6.7-fold increase in HMG-CoA reductase activity, which is restored to normal levels by mevalonate. HMG-CoA Synthase activity is not modified by either of these effectors, suggesting that the mevalonate pathway in this insect cell line is regulated by post-transcriptional mechanisms affecting HMG-CoA reductase but not HMG-CoA Synthase.
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Low activity of mitochondrial HMG-CoA Synthase in liver of starved piglets is due to low levels of protein despite high mRNA levels.
Archives of biochemistry and biophysics, 2001Co-Authors: Maria J. Barrero, Diego Haro, Fausto G Hegardt, Jose Ortiz, Clarice S. Alho, Pedro F. MarreroAbstract:Abstract The unusually low hepatic ketogenic capacity of piglets has been correlated with lack of expression of the mitochondrial HMG-CoA Synthase gene. However, we have shown that starvation of 2-week-old piglets increased the mRNA levels of mitochondrial HMG-CoA Synthase to a level similar to that observed in starved rats (S. H. Adams, C. S. Alho, G. Asins, F. G. Hegardt, and P. F. Marrero, 1997, Biochem. J. 324, 65–73). We now report that antibodies against pig mitochondrial HMG-CoA Synthase detected the pig enzyme in mitochondria of 2-week-old starved piglets and that the pig mitochondrial HMG-CoA Synthase cDNA encodes an active enzyme in the eukaryotic cell line Mev-1, with catalytic behavior similar to that of the rat enzyme when expressed in the same system. We also show that low activity of pig mitochondrial HMG-CoA Synthase correlates with low expression of the pig enzyme. The discrepancy in mitochondrial HMG-CoA Synthase gene expression between the high levels of mRNA and low levels of enzyme was not associated with differences in transcript maturation, which suggests that an attenuated translation of the pig mRNA is responsible for the diminished ketogenic capacity of pig mitochondria.
Diego Haro - One of the best experts on this subject based on the ideXlab platform.
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Influence of DMBA-induced mammary cancer on the liver CPT I, mit HMG-CoA Synthase and PPARα mRNA expression in rats fed a low or high corn oil diet
International journal of molecular medicine, 2004Co-Authors: Raquel Moral, Diego Haro, Montserrat Solanas, Eva Mónica Manzanares, Eduard EscrichAbstract:Hepatic mitochondrial outer membrane carnitine palmitoyltransferase I (CPT I) and mitochondrial 3-hydroxy-3-methylglutaryl-CoA Synthase (HMG-CoA Synthase) enzymes play a key role in regulation of fatty acid oxidation and in ketogenic pathways, respectively. Their expression are regulated by fatty acids mainly by the peroxisome proliferator-activated receptor alpha (PPARalpha). To investigate possible mechanisms through which cancer alters the lipid metabolism, we analyzed by Northern blot, the mRNA relative abundance of these proteins in liver from healthy and DMBA-induced mammary tumor-bearing rats fed a low or high corn oil diet. Serum levels of lipids, body weight and mass were also determined. Whereas mRNA steady-state levels of CPT I and mit HMG-CoA Synthase were unaffected by the presence of the extra-hepatic tumor, the cancer state seemed to modify the regulation of the expression of these genes by high fat diet. We hypothesize that putative changes in PPARalpha mRNA levels could have contributed to such alterations. These results, together with changes in serum lipid profiles, body weight and mass, indicate fat mobilization and non-enhanced oxidation rates despite a high-fat feeding. This effect of the cancer state could be related to tumor aggressiveness and suggest a preferential redirection of long-chain fatty acids into energetic and specific pathways of the cancer cells.
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Histone deacetylase inhibitors stimulate mitochondrial HMG‐CoA Synthase gene expression via a promoter proximal Sp1 site
Nucleic acids research, 2003Co-Authors: Nuria Camarero, Alícia Nadal, Diego Haro, Maria J. Barrero, Pedro F. MarreroAbstract:The expression of mitochondrial HMG-CoA Synthase in the colon has been correlated with the levels of butyrate present in this tissue. We report here that the effect of butyrate on mitochondrial HMG-CoA Synthase gene expression is exerted in vivo at the transcriptional level, and that trichostatin A (TSA), a specific histone deacetylase inhibitor, also induces transcriptional activity and mRNA expression of the gene in human cell lines derived from colon carcinoma. Using chromatin immunoprecipitation assays, we show that histone deacetylase 1 (HDAC1) is associated with the endogenous mitochondrial HMG-CoA Synthase promoter and that TSA induction correlates with hyperacetylation of H4 histone associated with the 5' flanking region of the gene. Overexpression of HDAC1 activity leads consistently to mitochondrial HMG-CoA Synthase promoter hypoacetylation and reduces its transcriptional activity. The effect of butyrate and TSA maps to a single Sp1 site present in the proximal promoter of the gene, which is able to bind Sp1 and Sp3 proteins. Interestingly, the binding affinity of Sp1 and Sp3 proteins to the Sp1 site correlates with the TSA responsiveness of the promoter. Using a one-hybrid system (GAL4-Sp1 and GAL4-Sp3), we show that both proteins can mediate responsiveness to TSA in CaCo-2 cells employing distinct mechanisms.
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Low activity of mitochondrial HMG-CoA Synthase in liver of starved piglets is due to low levels of protein despite high mRNA levels.
Archives of biochemistry and biophysics, 2001Co-Authors: Maria J. Barrero, Diego Haro, Fausto G Hegardt, Jose Ortiz, Clarice S. Alho, Pedro F. MarreroAbstract:Abstract The unusually low hepatic ketogenic capacity of piglets has been correlated with lack of expression of the mitochondrial HMG-CoA Synthase gene. However, we have shown that starvation of 2-week-old piglets increased the mRNA levels of mitochondrial HMG-CoA Synthase to a level similar to that observed in starved rats (S. H. Adams, C. S. Alho, G. Asins, F. G. Hegardt, and P. F. Marrero, 1997, Biochem. J. 324, 65–73). We now report that antibodies against pig mitochondrial HMG-CoA Synthase detected the pig enzyme in mitochondria of 2-week-old starved piglets and that the pig mitochondrial HMG-CoA Synthase cDNA encodes an active enzyme in the eukaryotic cell line Mev-1, with catalytic behavior similar to that of the rat enzyme when expressed in the same system. We also show that low activity of pig mitochondrial HMG-CoA Synthase correlates with low expression of the pig enzyme. The discrepancy in mitochondrial HMG-CoA Synthase gene expression between the high levels of mRNA and low levels of enzyme was not associated with differences in transcript maturation, which suggests that an attenuated translation of the pig mRNA is responsible for the diminished ketogenic capacity of pig mitochondria.
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Contribution of steroidogenic factor 1 to the regulation of cholesterol synthesis.
Biochemical Journal, 2000Co-Authors: Cristina Mascaró, Alícia Nadal, Pedro F. Marrero, Fausto G Hegardt, Diego HaroAbstract:Steroidogenic factor 1 (SF-1) is an orphan member of the nuclear receptor family expressed in steroidogenic tissues, where it has an essential role in the regulation of the steroid hormone biosynthesis, adrenal and gonadal development and endocrine responses fundamental for reproduction. Here we show that SF-1 regulates the transcription of cytosolic 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) Synthase gene, which is essential for the endogenous synthesis of cholesterol. We have identified an element located 365bp upstream of the gene for cytosolic HMG-CoA Synthase; SF-1 binds as a monomer to this element and confers SF-1 responsiveness to homologous and heterologous promoters. It has been shown that in tissues with a high demand for cholesterol to be used in steroid synthesis, there is a lack of correlation between the cholesterol levels and the activity of the limiting enzymes of the mevalonate pathway. In accord with those results, we observed that cholesterol synthesis from acetate and either cytosolic HMG-CoA mRNA expression or transcriptional activity were not changed in response to 25-hydroxycholesterol in the SF-1-expressing steroidogenic Leydig tumour MA-10 cells. Moreover, the overexpression of SF-1 in non-steroidogenic CV-1 cells renders them less sensitive to the regulatory effects of cholesterol. This observation led to the hypothesis that in steroidogenic tissues the expression of SF-1 permits high levels of endogenous synthesis of cholesterol irrespective of the intracellular levels of this metabolite.
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Sterol regulatory element binding protein-mediated effect of fluvastatin on cytosolic 3-hydroxy-3-methylglutaryl-coenzyme A Synthase transcription.
Archives of biochemistry and biophysics, 2000Co-Authors: Cristina Mascaró, Diego Haro, Jose Ortiz, Ma Mercedes Ramos, Fausto G HegardtAbstract:Abstract The effects of acute treatment with fluvastatin, a hypocholesteremic drug, on the mRNA levels of several regulatory enzymes of cholesterogenesis and of the LDL receptor were determined in rat liver. Fluvastatin increased the hepatic mRNA levels for HMG-CoA reductase up to 12-fold in 5 weeks of treatment at a daily dose of 6.3 mg/kg. The effect was less marked in cytosolic HMG-CoA Synthase, farnesyl-PP Synthase, squalene synthetase, and LDL receptor. SREBP-2 mRNA levels were also increased, but SREBP-1 were not. De novo synthesis of cholesterol in several cultured cells was reduced by increasing concentrations of fluvastatin, and the IC 50 values of fluvastatin in HepG2, CV-1, and CHO cells were respectively 0.01, 0.05, and 0.1 μM. When CHO cells stably transfected with a chimeric gene composed of the promoter of cytosolic HMG-CoA Synthase and the CAT gene as a reporter were incubated with fluvastatin, the CAT gene was overexpressed, an effect which was similar to the cotransfection with the processed form of SREBP-1a. Both ALLN and fluvastatin increased the transcriptional activity of cytosolic HMG-CoA Synthase. Mutation in either SRE or NF-Y boxes abolished the increase in transcriptional rate caused by fluvastatin in the promoter of cytosolic HMG-CoA Synthase. These results indicate that the increase in transcriptional activity in the HMG-CoA Synthase gene attributable to fluvastatin is a consequence of the activation of the proteolytic cleavage of SREBPs by reduced levels of intracellular cholesterol.
Pedro F. Marrero - One of the best experts on this subject based on the ideXlab platform.
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Histone deacetylase inhibitors stimulate mitochondrial HMG‐CoA Synthase gene expression via a promoter proximal Sp1 site
Nucleic acids research, 2003Co-Authors: Nuria Camarero, Alícia Nadal, Diego Haro, Maria J. Barrero, Pedro F. MarreroAbstract:The expression of mitochondrial HMG-CoA Synthase in the colon has been correlated with the levels of butyrate present in this tissue. We report here that the effect of butyrate on mitochondrial HMG-CoA Synthase gene expression is exerted in vivo at the transcriptional level, and that trichostatin A (TSA), a specific histone deacetylase inhibitor, also induces transcriptional activity and mRNA expression of the gene in human cell lines derived from colon carcinoma. Using chromatin immunoprecipitation assays, we show that histone deacetylase 1 (HDAC1) is associated with the endogenous mitochondrial HMG-CoA Synthase promoter and that TSA induction correlates with hyperacetylation of H4 histone associated with the 5' flanking region of the gene. Overexpression of HDAC1 activity leads consistently to mitochondrial HMG-CoA Synthase promoter hypoacetylation and reduces its transcriptional activity. The effect of butyrate and TSA maps to a single Sp1 site present in the proximal promoter of the gene, which is able to bind Sp1 and Sp3 proteins. Interestingly, the binding affinity of Sp1 and Sp3 proteins to the Sp1 site correlates with the TSA responsiveness of the promoter. Using a one-hybrid system (GAL4-Sp1 and GAL4-Sp3), we show that both proteins can mediate responsiveness to TSA in CaCo-2 cells employing distinct mechanisms.
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3-Hydroxy-3-methylglutaryl coenzyme A Synthase-1 of Blattella germanica has structural and functional features of an active retrogene
Insect Biochemistry and Molecular Biology, 2001Co-Authors: Nuria Casals, Pedro F. Marrero, Xavier Belles, Carlos Buesa, Fausto G HegardtAbstract:Blattella germanica has two cytosolic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) Synthase genes, HMG-CoA Synthase-1 and -2. HMG-CoA Synthase-1 gene shows several features of processed genes (retroposons): it contains no introns but has a short direct-repeat sequence (ATTATTATT) at both ends. An atypical feature is the presence at both ends of the gene of short inverse repeats flanked by direct repeats. There is neither a TATA box nor a CAAT box in the 5' region. Comparative analysis with other species suggests that the HMG-CoA Synthase-1 gene derives from HMG-CoA Synthase-2. Cultured embryonic B. germanica UM-BGE-1 cells express HMG-CoA Synthase-1 but not HMG-CoA Synthase-2, suggesting that the intron-less gene is functional. In addition, it can complement MEV-1 cell line, which is auxotrophic for mevalonate. We show that compactin and mevalonate do not significantly affect the mRNA levels of HMG-CoA Synthase-1 in UM-BGE-1 cells. Compactin induces a 6.7-fold increase in HMG-CoA reductase activity, which is restored to normal levels by mevalonate. HMG-CoA Synthase activity is not modified by either of these effectors, suggesting that the mevalonate pathway in this insect cell line is regulated by post-transcriptional mechanisms affecting HMG-CoA reductase but not HMG-CoA Synthase.
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Low activity of mitochondrial HMG-CoA Synthase in liver of starved piglets is due to low levels of protein despite high mRNA levels.
Archives of biochemistry and biophysics, 2001Co-Authors: Maria J. Barrero, Diego Haro, Fausto G Hegardt, Jose Ortiz, Clarice S. Alho, Pedro F. MarreroAbstract:Abstract The unusually low hepatic ketogenic capacity of piglets has been correlated with lack of expression of the mitochondrial HMG-CoA Synthase gene. However, we have shown that starvation of 2-week-old piglets increased the mRNA levels of mitochondrial HMG-CoA Synthase to a level similar to that observed in starved rats (S. H. Adams, C. S. Alho, G. Asins, F. G. Hegardt, and P. F. Marrero, 1997, Biochem. J. 324, 65–73). We now report that antibodies against pig mitochondrial HMG-CoA Synthase detected the pig enzyme in mitochondria of 2-week-old starved piglets and that the pig mitochondrial HMG-CoA Synthase cDNA encodes an active enzyme in the eukaryotic cell line Mev-1, with catalytic behavior similar to that of the rat enzyme when expressed in the same system. We also show that low activity of pig mitochondrial HMG-CoA Synthase correlates with low expression of the pig enzyme. The discrepancy in mitochondrial HMG-CoA Synthase gene expression between the high levels of mRNA and low levels of enzyme was not associated with differences in transcript maturation, which suggests that an attenuated translation of the pig mRNA is responsible for the diminished ketogenic capacity of pig mitochondria.
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Contribution of steroidogenic factor 1 to the regulation of cholesterol synthesis.
Biochemical Journal, 2000Co-Authors: Cristina Mascaró, Alícia Nadal, Pedro F. Marrero, Fausto G Hegardt, Diego HaroAbstract:Steroidogenic factor 1 (SF-1) is an orphan member of the nuclear receptor family expressed in steroidogenic tissues, where it has an essential role in the regulation of the steroid hormone biosynthesis, adrenal and gonadal development and endocrine responses fundamental for reproduction. Here we show that SF-1 regulates the transcription of cytosolic 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) Synthase gene, which is essential for the endogenous synthesis of cholesterol. We have identified an element located 365bp upstream of the gene for cytosolic HMG-CoA Synthase; SF-1 binds as a monomer to this element and confers SF-1 responsiveness to homologous and heterologous promoters. It has been shown that in tissues with a high demand for cholesterol to be used in steroid synthesis, there is a lack of correlation between the cholesterol levels and the activity of the limiting enzymes of the mevalonate pathway. In accord with those results, we observed that cholesterol synthesis from acetate and either cytosolic HMG-CoA mRNA expression or transcriptional activity were not changed in response to 25-hydroxycholesterol in the SF-1-expressing steroidogenic Leydig tumour MA-10 cells. Moreover, the overexpression of SF-1 in non-steroidogenic CV-1 cells renders them less sensitive to the regulatory effects of cholesterol. This observation led to the hypothesis that in steroidogenic tissues the expression of SF-1 permits high levels of endogenous synthesis of cholesterol irrespective of the intracellular levels of this metabolite.
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Isolation of pig mitochondrial 3-hydroxy-3-methylglutaryl-CoA Synthase gene promoter: characterization of a peroxisome proliferator-responsive element.
Biochemical Journal, 1999Co-Authors: Jose Ortiz, Diego Haro, Joan C. Rodríguez, Fausto G Hegardt, Judith Mallolas, Carine Nicot, Josep Bofarull, Pedro F. MarreroAbstract:Low expression of the mitochondrial 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) Synthase gene during development correlates with an unusually low hepatic ketogenic capacity and lack of hyperketonaemia in piglets. Here we report the isolation and characterization of the 5' end of the pig mitochondrial HMG-CoA Synthase gene. The 581 bp region proximal to the transcription start site permits transcription of a reporter gene, confirming the function of the promoter. The pig mitochondrial HMG-CoA Synthase promoter is trans-activated by the peroxisomal proliferator-activated receptor (PPAR), and a functional response element for PPAR (PPRE) has been localized in the promoter region. Pig PPRE is constituted by an imperfect direct repeat (DR-1) and a downstream sequence, both of which are needed to confer PPAR-sensitivity to a thymidine kinase promoter and to form complexes with PPAR.retinoid X receptor heterodimers. A role of PPAR trans-activation in starvation-associated induction of gene expression is suggested.
Dolors Serra - One of the best experts on this subject based on the ideXlab platform.
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The effect of dexamethasone treatment on the expression of the regulatory genes of ketogenesis in intestine and liver of suckling rats.
Molecular and Cellular Biochemistry, 1998Co-Authors: Gladys Arias, Fausto G Hegardt, Guillermina Asins, Dolors SerraAbstract:The influence of the injection of dexamethasone on ketogenesis in 12 day old suckling rats was studied in intestine and liver by determining mRNA levels and enzyme activity of the two genes responsible for regulation of ketogenesis: carnitine palmitoyl transferase I (CPT 1) and mitochondrial HMG-CoA Synthase. Dexamethasone produced a 2 fold increase in mRNA and activity of CPT I in intestine, but led to a decrease in mitt HMG-CoA Synthase. In liver the mRNA levels and activity of both CPT I and mitt HMG-CoA Synthase decreased. Comparison of these values with the ketogenic rate of both tissues following dexamethasone treatment suggests that mitt HMG-CoA Synthase could be the main gene responsible for the regulation of ketogenesis in suckling rats. The changes produced in serum ketone bodies by dexamethasone, with a profile that is more similar to the ketogenic rate in the liver than that in the intestine, indicate that liver contributes more to ketone body synthesis in suckling rats. Two day treatment with dexamethasone produced no change in mRNA or activity levels for CPT I in liver or intestine. While mRNA levels for mitt HMG-CoA Synthase changed little, the enzyme activity is decreased in both tissues.
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The effect of fasting/refeeding and insulin treatment on the expression of the regulatory genes of ketogenesis in intestine and liver of suckling rats.
Archives of biochemistry and biophysics, 1997Co-Authors: Gladys Arias, Fausto G Hegardt, Guillermina Asins, Dolors SerraAbstract:The influence of fasting/refeeding and insulin treatment on ketogenesis in 12-day-old suckling rats was studied in intestine and liver by determining mRNA levels and enzyme activity of the two genes responsible for regulation of ketogenesis: carnitine palmitoyl transferase I (CPT I) and mitochondrial HMG-CoA Synthase. Fasting produced hardly any change in mRNA or activity of CPT 1 in intestine, but led to a decrease in mitochondrial (mit.) HMG-CoA Synthase. In liver, while mRNA levels and activity for CPT I increased, neither parameter was changed in HMG-CoA Synthase. The comparison of these values with the ketogenic rate of both tissues under the fasting/refeeding treatment shows that HMG-CoA Synthase could be the main gene responsible for regulation of ketogenesis in suckling rats. The small changes produced in serum ketone bodies in fasting/refeeding, with a profile similar to the ketogenic rate of the liver, indicate that liver contributes most to ketone body synthesis in suckling rats under these experimental conditions. Short-term insulin treatment produced increases in mRNA levels and activity in CPT I in intestine, but it also decreased both parameters in mit. HMG-CoA Synthase. In liver, graphs of mRNA and activity were nearly identical in both genes. There was a marked decrease in mRNA levels and activity, resembling those values observed in adult rats. As in fasting/refeeding, the ketogenic rate correlated better to mit. HMG-CoA Synthase than CPT I, and liver was the main organ regulating ketogenesis after insulin treatment. Serum ketone body concentrations were decreased by insulin but recovered after the second hour. Long-term insulin treatment had little effect on the mRNA levels for CPT I or mit. HMG-CoA Synthase, but both the expressed and total activities of mit. HMG-CoA Synthase were reduced by half in both intestine and liver. The ketogenic rate of both organs was decreased to 40% by long-term insulin treatment. The different effects of refeeding and insulin treatment on the expression of both genes, on the ketogenic rate, and on ketone body concentrations are discussed.
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The effect of etomoxir on the mRNA levels of enzymes involved in ketogenesis and cholesterogenesis in rat liver
Biochemical pharmacology, 1994Co-Authors: Guillermina Asins, Dolors Serra, Fausto G HegardtAbstract:The effects of acute treatment with 2-[6-(4-chlorophenoxy)hexyl]-oxirane-2-carboxylate (etomoxir), an antiketonaemic and antidiabetic drug, on the mRNA levels of several regulatory enzymes of ketogenesis, cholesterogenesis, and fatty acid synthesis in rats were determined. In rats treated with etomoxir, mRNA levels for mitochondrial 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) Synthase and carnitine palmitoyl transferase I (CPT I) remained unchanged, while mRNA levels for carnitine palmitoyl transferase II (CPT II) significantly increased 2-fold. Injection of etomoxir produced no effect on the mRNA levels of cytosolic HMG-CoA Synthase but increased the mRNA levels of HMG-CoA reductase 2.5-fold. Etomoxir led to a 3-fold increase in the mRNA levels of fatty acid Synthase of rats under acute treatment. Rats fed with a fat diet significantly increased the expression of mitochondrial HMG-CoA Synthase, CPT I and CPT II 3-fold in all cases, while 2-(diethylhexyl)phthalate (DEHP) produced increases in the expression of these genes (5-, 4- and 12-fold, respectively). The mRNA levels of HMG-CoA reductase were not changed by either DEHP or fat diet, while DEHP increased cytosolic HMG-CoA Synthase 2.5-fold. DEHP did not change the mRNA levels for fatty acid Synthase. It was concluded that etomoxir does not produce its hypoketonaemic, hypocholesteraemic or hypolipogenic effects through changes in the genetic expression of the regulatory enzymes of these pathways, but probably due to the shortage of their common substrate, acetyl-CoA, because of the inhibitory action on CPT I.
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Gene expression of enzymes regulating ketogenesis and fatty acid metabolism in regenerating rat liver.
Biochemical Journal, 1994Co-Authors: Guillermina Asins, José Ayté, Gabriel Gil-gómez, Dolors Serra, Jose Luis Rosa, Ramon Bartrons, Albert Tauler, Fausto G HegardtAbstract:Levels of mRNA for mitochondrial 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) Synthase, carnitine palmitoyltransferase I (CPT I) and carnitine palmitoyltransferase II (CPT II), fatty acid Synthase (FAS) and actin were analysed during liver regeneration. mRNA levels for mitochondrial HMG-CoA Synthase decreased rapidly, reaching a minimum 12 h after partial hepatectomy and returning to normal at 24-36 h. In contrast, CPT I, CPT II and FAS mRNAs increased throughout the period examined. Expression of actin increased significantly during regeneration. Levels of mRNA for mitochondrial HMG-CoA Synthase also decreased as a result of surgical stress, although the effect of hepatectomy was much greater. We determined the levels of mitochondrial HMG-CoA Synthase using specific antibodies. The amount of protein rapidly decreased, although less markedly than the corresponding mRNA levels. These results show that the decrease described in ketogenesis in partially hepatectomized rats correlated with the decrease in the expression of mitochondrial HMG-CoA Synthase, suggesting that this enzyme may also be a control point in ketogenesis in the regenerating liver, as it is in normal and diabetic rats.
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Regulation of mitochondrial 3-hydroxy-3-methylglutaryl-coenzyme A Synthase protein by starvation, fat feeding, and diabetes
Archives of biochemistry and biophysics, 1993Co-Authors: Dolors Serra, Nuria Casals, T. Royo, Carlos J. Ciudad, Guillermina Asins, Fausto G HegardtAbstract:We have determined the levels of mitochondrial 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) Synthase under different metabolic situations to examine its potential role as a regulatory protein in the ketogenic pathway. We used specific antibodies directed against a peptide of the amino acid sequence of the protein as deduced from the cDNA sequence. The amount of mitochondrial HMG-CoA Synthase protein rapidly increased in response to cyclic AMP, dexamethasone, starvation, fat feeding, and diabetes, whereas it was decreased by insulin and refeeding. Insulin was also able to counteract the increase in mitochondrial HMG-CoA Synthase levels observed under the diabetic condition. Furthermore, the finding that quantitative changes in HMG-CoA Synthase protein were less marked than those in the corresponding mRNA in starved and diabetic rats suggests either translational control or increased degradation of either mRNA or protein. All these results indicate that mitochondrial HMG-CoA Synthase is a regulatory element in the ketogenic process.
Nuria Casals - One of the best experts on this subject based on the ideXlab platform.
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Refining the diagnosis of mitochondrial HMG‐CoA Synthase deficiency
Journal of inherited metabolic disease, 2006Co-Authors: Rosa Aledo, Fausto G Hegardt, Nuria Casals, Juan Pié, Cecilia Mir, R. N. Dalton, Charles Turner, Michael ChampionAbstract:Mitochondrial HMG-CoA Synthase deficiency is an inherited metabolic disorder caused by a defect in the enzyme that regulates the formation of ketone bodies. Patients present with hypoketotic hypoglycaemia, encephalopathy and hepatomegaly, usually precipitated by an intercurrent infection or prolonged fasting. The diagnosis may easily be missed as previously reported results of routine metabolic investigations, urinary organic acids and plasma acylcarnitines may be nonspecific or normal, and a high index of suspicion is required to proceed to further confirmatory tests. We describe a further acute case in which the combination of urinary organic acids, low free carnitine and changes in the plasma acylcarnitine profile on carnitine supplementation were very suggestive of a defect in ketone synthesis. The diagnosis of mitochondrial HMG-CoA Synthase deficiency was confirmed on genotyping, revealing two novel mutations: c.614G > A (R188H) and c.971T > C (M307T). A further sibling, in whom the diagnosis had not been made acutely, was also found to be affected. The possible effects of these mutations on enzyme activity are discussed.
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The diagnosis of mitochondrial HMG-CoA Synthase deficiency☆
The Journal of pediatrics, 2002Co-Authors: Johannes Zschocke, Fausto G Hegardt, Nuria Casals, Juan Pié, Johannes M. Penzien, Rainer Bielen, Rosa Aledo, Georg F. Hoffmann, Ertan MayatepekAbstract:Deficiency of 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) Synthase, the only disorder exclusively affecting hepatic ketogenesis, is a cause of hypoglycemic coma. We report that the diagnosis can be made by typical laboratory findings (hypoketosis, elevated free fatty acids, normal acylcarnitines, specific urinary organic acids) during acute episodes.
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Genetic basis of mitochondrial HMG-CoA Synthase deficiency
Human Genetics, 2001Co-Authors: Rosa Aledo, Nuria Casals, Juan Pié, Johannes Zschocke, Georg F. Hoffmann, Ertan Mayatepek, Cecilia Mir, Sonja Fiesel, Fausto G HegardtAbstract:Deficiency of mitochondrial 3-hydroxy-3-methylglutaryl-CoA Synthase (mHMGS) is a recessive disorder of ketogenesis that has been previously diagnosed in two children with hypoglycaemic hypoketotic coma during fasting periods. Here, we report the results of molecular investigations in a third patient affected by this disease. Sequencing of the entire coding region of the HMGCS2 gene revealed two missense mutations, G212R and R500H. Mendelian inheritance was confirmed by the analysis of parental samples and neither of the mutations was found on 200 control chromosomes. Functional relevance was confirmed by in vitro expression studies in cytosolic HMGS-deficient cells. Whereas wild-type cDNA of the HMGCS2 gene reverted the auxotrophy for mevalonate, the cDNAs of the mutants did not. The disease may be recognised by specific clinical and biochemical features but it is difficult to confirm enzymatically since the gene is expressed only in liver and testis. Molecular studies may facilitate or confirm future diagnoses in affected patients.
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3-Hydroxy-3-methylglutaryl coenzyme A Synthase-1 of Blattella germanica has structural and functional features of an active retrogene
Insect Biochemistry and Molecular Biology, 2001Co-Authors: Nuria Casals, Pedro F. Marrero, Xavier Belles, Carlos Buesa, Fausto G HegardtAbstract:Blattella germanica has two cytosolic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) Synthase genes, HMG-CoA Synthase-1 and -2. HMG-CoA Synthase-1 gene shows several features of processed genes (retroposons): it contains no introns but has a short direct-repeat sequence (ATTATTATT) at both ends. An atypical feature is the presence at both ends of the gene of short inverse repeats flanked by direct repeats. There is neither a TATA box nor a CAAT box in the 5' region. Comparative analysis with other species suggests that the HMG-CoA Synthase-1 gene derives from HMG-CoA Synthase-2. Cultured embryonic B. germanica UM-BGE-1 cells express HMG-CoA Synthase-1 but not HMG-CoA Synthase-2, suggesting that the intron-less gene is functional. In addition, it can complement MEV-1 cell line, which is auxotrophic for mevalonate. We show that compactin and mevalonate do not significantly affect the mRNA levels of HMG-CoA Synthase-1 in UM-BGE-1 cells. Compactin induces a 6.7-fold increase in HMG-CoA reductase activity, which is restored to normal levels by mevalonate. HMG-CoA Synthase activity is not modified by either of these effectors, suggesting that the mevalonate pathway in this insect cell line is regulated by post-transcriptional mechanisms affecting HMG-CoA reductase but not HMG-CoA Synthase.
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Coordinated Expression and Activity of 3- Hydroxy-3-Methylglutaryl Coenzyme A Synthase and Reductase in the Fat Body of Blattella germanica (L.) During Vitellogenesis
Insect biochemistry and molecular biology, 1996Co-Authors: Nuria Casals, Pedro F. Marrero, Maria-dolors Piulachs, Carlos Buesa, Joana Caban˜ó, Xavier Belle´s, Fausto G HegardtAbstract:Levels of mRNA for the two 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) Synthases, (HMG-S1 and HMG-S2), and for HMG-CoA reductase (HMG-R) of Blattella germanica were analyzed in the fat body during the first gonadotrophic cycle. HMG-S2 and HMG-R showed the highest mRNA levels on day 0 and decreased thereafter, whereas HMG-S1, showed faint expression. Western blot using specific antibodies for HMG-S1 and HMG-S2 showed no detectable levels for HMG-S1 but a clear pattern for HMG-S2. Both results point to a very limited role for HMG-CoA Synthase-1 in B. germanica fat body and suggest that the functional enzyme in this organ is HMG-CoA Synthase-2. HMG-CoA reductase and Synthase proteins shared a cyclic pattern (maximum levels at day 4 and minimum levels on days 0 and 8), which was coincident with the pattern of activity. The delay between gene transcription and protein synthesis suggests a finely regulated translation mechanism. Moreover, the pattern of mevalonate synthesis parallels that of vitellogenin production, suggesting a coordinate mechanism between the mevalonate pathway and the production of viteilogenin. Copyright © 1996