Effects of nonsulfur and sulfur amino acids on the regulation of hepatic enzymes of cysteine metabolism.
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To determine the role of nonsulfur vs. sulfur amino acids in regulation of cysteine metabolism, rats were fed a basal diet or diets supplemented with a mixture of nonsulfur amino acids (AA), sulfur amino acids (SAA), or both for 3 wk. Hepatic cysteine-sulfinate decarboxylase (CSDC), cysteine dioxygenase (CDO), and gamma-glutamylcysteine synthetase (GCS) activity, concentration, and mRNA abundance were measured. Supplementation with AA alone had no effect on any of these measures. Supplementation of the basal diet with SAA, with or without AA, resulted in a higher CDO concentration (32-45 times basal), a lower CSDC mRNA level (49-64% of basal), and a lower GCS-heavy subunit mRNA level (70-76%). The presence of excess SAA and AA together resulted in an additional type of regulation: a lower specific activity of all three enzymes was observed in rats fed diets with an excess of AA and SAA. Both SAA and AA played a role in regulation of these three enzymes of cysteine metabolism, but SAA had the dominant effects, and effects of AA were not observed in the absence of SAA. (+info)
Human cysteine dioxygenase gene: structural organization, tissue-specific expression and downregulation by phorbol 12-myristate 13-acetate.
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The organization of the human cysteine dioxygenase (CDO) gene was found to be similar to its rat counterpart, and the location of the introns in the protein structure was identical to the rat CDO gene. The major transcription start site, identified by primer extension, was located 260 bp upstream from the ATG codon. The sequence of the 5'-immediate upstream region was highly conserved between the human and rat CDO genes. The putative promoter region contained a TATA-box-like sequence, and many putative cis-acting elements including HNF5, GRE, TRE, CRE, CArG box, ARE, MBS, and NF-kB. A Northern blot analysis revealed that CDO mRNA was strongly expressed in the liver and placenta, and weakly in the heart, brain and pancreas. CDO mRNA was also detected in human hepatoblastoma HepG2 cells. The CDO mRNA level in HepG2 cells was decreased after 2 h and reached a minimum 6 h-8 h after a phorbol 12-myristate 13-acetate (PMA) treatment, and then gradually returned to the basal level. (+info)
C/EBPbeta, when expressed from the C/ebpalpha gene locus, can functionally replace C/EBPalpha in liver but not in adipose tissue.
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Knockout of C/EBPalpha causes a severe loss of liver function and, subsequently, neonatal lethality in mice. By using a gene replacement approach, we generated a new C/EBPalpha-null mouse strain in which C/EBPbeta, in addition to its own expression, substituted for C/EBPalpha expression in tissues. The homozygous mutant mice C/ebpalpha(beta/beta) are viable and fertile and show none of the overt liver abnormalities found in the previous C/EBPalpha-null mouse line. Levels of hepatic PEPCK mRNA are not different between C/ebpalpha(beta/beta) and wild-type mice. However, despite their normal growth rate, C/ebpalpha(beta/beta) mice have markedly reduced fat storage in their white adipose tissue (WAT). Expression of two adipocyte-specific factors, adipsin and leptin, is significantly reduced in the WAT of C/ebpalpha(beta/beta) mice. In addition, expression of the non-adipocyte-specific genes for transferrin and cysteine dioxygenase is reduced in WAT but not in liver. Our study demonstrates that when expressed from the C/ebpalpha gene locus, C/EBPbeta can act for C/EBPalpha to maintain liver functions during development. Moreover, our studies with the C/ebpalpha(beta/beta) mice provide new insights into the nonredundant functions of C/EBPalpha and C/EBPbeta on gene regulation in WAT. (+info)
Cysteine regulates expression of cysteine dioxygenase and gamma-glutamylcysteine synthetase in cultured rat hepatocytes.
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Rat hepatocytes cultured for 3 days in basal medium expressed low levels of cysteine dioxygenase (CDO) and high levels of gamma-glutamylcysteine synthetase (GCS). When the medium was supplemented with 2 mmol/l methionine or cysteine, CDO activity and CDO protein increased by >10-fold and CDO mRNA increased by 1.5- or 3.2-fold. In contrast, GCS activity decreased to 51 or 29% of basal, GCS heavy subunit (GCS-HS) protein decreased to 89 or 58% of basal, and GCS mRNA decreased to 79 or 37% of basal for methionine or cysteine supplementation, respectively. Supplementation with cysteine consistently yielded responses of greater magnitude than did supplementation with an equimolar amount of methionine. Addition of propargylglycine to inhibit cystathionine gamma-lyase activity and, hence, cysteine formation from methionine prevented the effects of methionine, but not those of cysteine, on CDO and GCS expression. Addition of buthionine sulfoximine to inhibit GCS, and thus block glutathione synthesis from cysteine, did not alter the ability of methionine or cysteine to increase CDO. GSH concentration was not correlated with changes in either CDO or GCS-HS expression. The effectiveness of cysteine was equivalent to or greater than that of its precursors (S-adenosylmethionine, cystathionine, homocysteine) or metabolites (taurine, sulfate). Taken together, these results suggest that cysteine itself is an important cellular signal for upregulation of CDO and downregulation of GCS. (+info)
The aetiology of idiopathic Parkinson's disease.
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Agents potentially involved in the aetiology of idiopathic Parkinson's disease are discussed. These include factors regulating dopaminergic neurogenesis (Nurr 1, Ptx-3, and Lmx1b) and related proteins, together with genes involved in familial Parkinson's disease (alpha synuclein, parkin, and ubiquitin carboxy terminal hydroxylase L1), and endogenous and environmental agents. (+info)
Enzymes and metabolites of cysteine metabolism in nonhepatic tissues of rats show little response to changes in dietary protein or sulfur amino acid levels.
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In liver, cysteine dioxygenase (CDO), cysteinesulfinate decarboxylase (CSD), and gamma-glutamylcysteine synthetase (GCS) play important regulatory roles in the metabolism of cysteine to sulfate, taurine and glutathione. Because glutathione is released by the liver and degraded by peripheral tissues that express gamma-glutamyl transpeptidase, some peripheral tissues may be exposed to relatively high concentrations of cysteine. Rats were fed diets that contained low, moderate or high concentrations of protein or supplemental cysteine or methionine for 2 wk, and CDO, CSD and GCS activities, concentrations and mRNA levels and the concentrations of cysteine, taurine and glutathione were measured in liver, kidney, lung and brain. All three enzymes in liver responded to the differences in dietary protein or sulfur amino acid levels, but only CSD in kidney and none of the three enzymes in lung and brain responded. Renal CSD activity was twice as much in rats fed the low protein diet as in rats fed the other diets. Changes in renal CSD activity were correlated with changes in CSD concentration. Some significant differences in cysteine concentration in kidney and lung and glutathione and taurine concentrations in kidney were observed, with higher concentrations in rats fed higher levels of protein or sulfur amino acids. In liver, the changes in cysteine level were consistent with cysteine-mediated regulation of hepatic CDO activity, and changes in taurine level were consistent with predicted changes in cysteine catabolism due to the changes in cysteine concentration and CDO activity. Changes in renal and lung cysteine, taurine or glutathione concentrations were not associated with a similar pattern of change in CDO, CSD or GCS activity. Overall, the results confirm the importance of the liver in the maintenance of cysteine homeostasis. (+info)
Functional characterization and regulation of the taurine transporter and cysteine dioxygenase in human hepatoblastoma HepG2 cells.
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We investigated the characterization and the regulation of TAUT (taurine transporter) and CDO (cysteine dioxygenase), one of the key enzymes of taurine biosynthesis, in human hepatoblastoma HepG2 cells. The activity of TAUT in the HepG2 cells was evaluated by means of a sodium- and chloride-dependent high-affinity transport system, the characteristics of which were similar to those of the beta amino-acid-specific taurine transport system described previously for various tissues [Uchida, Kwon, Yamauchi, Preston, Marumo and Handler (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 8230-8234; Ramamoorthy, Leibach, Mahesh, Han, Yang-Feng, Blakely and Ganapathy (1994) Biochem. J. 300, 893-900; and Satsu, Watanabe, Arai and Shimizu (1997) J. Biochem. (Tokyo) 121, 1082-1087]. By culturing in a hypertonic medium, the intracellular taurine content of HepG2 cells was markedly increased. Under hypertonic conditions, the activity of TAUT was up-regulated, and the results of the kinetic analysis suggested that this up-regulation was associated with an increase in the amount of TAUT. The expression level of TAUT mRNA was markedly higher than that of the control cells. The expression level of CDO mRNA was also up-regulated under the hypertonic conditions. Culturing the cells in a taurine-rich medium resulted in both the activity of TAUT and the expression level of TAUT mRNA being down-regulated in HepG2 cells. On the other hand, the expression level of CDO mRNA was not affected under a taurine-rich condition. The present results show that both TAUT and CDO were co-operatively regulated in response to hypertonicity, but did not co-operatively respond to the change in extracellular taurine concentration. Generally, the TAUT and taurine biosynthetic enzymes have independent regulatory systems, but under certain conditions, they could be regulated in harmony with each other. (+info)
Cysteine is the metabolic signal responsible for dietary regulation of hepatic cysteine dioxygenase and glutamate cysteine ligase in intact rats.
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Cysteine, rather than a precursor or metabolite of cysteine, appears to mediate the upregulation of cysteine dioxygenase (CDO) and the downregulation of glutamate cysteine ligase (GCL) in cultured primary rat hepatocytes. However, similar experiments in intact rats have not been performed to confirm in vivo that changes in hepatic cysteine levels are associated with the regulation of CDO or GCL activity. Therefore, rats were fed a low protein basal diet (100 g casein/kg diet) with or without supplemental sulfur amino acids (8 g cystine, 9 g homocystine or 10 g methionine/kg diet) and with or without propargylglycine (PPG, 1 mmol/kg), an irreversible inhibitor of cystathionine gamma-lyase. Rats were fed the assigned diet for 2 full days and up until the mid-point of the dark cycle on d 3, at which time they were killed for collection of liver. Rats fed the PPG-containing diets had hepatic cystathionine gamma-lyase activities that were approximately 16% of the uninhibited level. PPG treatment reduced CDO activity by 50 and 54%, increased GCL activity by 41 and 61% and lowered total cysteine concentration by 33 and 64% in liver of the homocystine and methionine-supplemented groups, respectively, but not in the cystine-supplemented groups or unsupplemented groups. Glutathione levels were not affected by PPG treatment in any groups. These experiments are consistent with a role for cysteine, rather than a precursor or metabolite of cysteine, in the metabolic signaling responsible for diet-induced regulation of CDO and GCL. (+info)