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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Degradation of a protein via the ubiquitin proteolytic pathway involves two successive steps. Covalent attachment of ubiquitin to the target protein and degradation of the tagged substrate by the 26S proteasome. Most native cellular proteins that are targeted by the ubiquitin system are short-lived transcriptional activators and growth and cell cycle regulators, as well as unstable membrane proteins. In the present study we demonstrate the involvement of the system in the degradation of tyrosine aminotransferase (TAT), a key enzyme in intermediary metabolism. In vitro, we have shown that the native enzyme is conjugated and degraded in a system that requires ATP and ubiquitin. Degradation was monitored by following the decrease of catalytic activity as well as disappearance of the protein molecule. The enzyme could be protected from degradation by association with its specific cofactor, pyridoxal phosphate (PLP). In vivo, we prepared cell extracts from livers of animals in which TAT was induced by starvation and corticosteroid administration. The dramatic increase in the level of the enzyme was accompanied by a concomitant increase in the level of specific TAT-ubiquitin adducts.
Mol Biol Rep 1997 Mar
PMID:Ubiquitin-mediated degradation of tyrosine aminotransferase (TAT) in vitro and in vivo. 922 77

The participation of tyrosine kinase in the regulation of the glucocorticoid receptor (GR) was studied in primary cultured rat hepatocytes using the tyrosine kinase inhibitor herbimycin A. Herbimycin A decreased the number of high-affinity binding sites of glucocorticoids in the cytosolic fraction and increased the equilibrium dissociation constant (Kd). Western blot analysis revealed that it also decreased the amount of GR protein. On the other hand, cycloheximide did not affect the GR protein level. Although herbimycin A slightly increased the amount of GR protein in the nuclear fraction, the increase was much lower than that of its decrease in the cytosolic fraction. Therefore, the decrease of GR protein in the cytosolic fraction was not caused by the inhibition of GR protein synthesis nor the translocation of GR from cytosol to nuclei. As herbimycin A also suppressed the dexamethasone (Dex)-dependent induction of tyrosine aminotransferase (TAT) activity, the decrease of GR protein was followed by the suppression of the GR-mediated biological response. These findings indicate that tyrosine kinase is necessary for the maintenance of the level of GR protein and its affinity of binding sites in the cytosolic fraction. Our results also suggested that the increase of GR protein stability is the most probable explanation for the maintenance of its level.
J Steroid Biochem Mol Biol 1997 Apr
PMID:Regulation of glucocorticoid receptor by the tyrosine kinase inhibitor herbimycin A in the cytosolic fraction of primary cultured rat hepatocytes. 932 11

In the rat, the conformationally highly bent steroid 21-hydroxy-6, 19-oxidoprogesterone efficiently displaces [3H]corticosterone from thymus-glucocorticoid receptors and blocks type II receptors in kidney cytosols but competes with neither [3H]aldosterone for kidney-mineralocorticoid receptors nor [3H]progesterone for uterus-progesterone receptors. It evokes Na+ retention only at very high doses (approximately 100 microg/100 g of rat weight) and is unable to induce tyrosine aminotransferase or to increase glycogen deposits in rat liver. When coincubated with corticosterone or dexamethasone, 2.5 microM 21OH-6OP inhibits 80% of tyrosine aminotransferase induction. It may therefore be used experimentally as an antiglucocorticoid virtually lacking mineralocorticoid or glucocorticoid properties as well as affinity for mineralocorticoid or progesterone receptors.
Mol Pharmacol 1997 Oct
PMID:21-Hydroxy-6,19-oxidoprogesterone: a novel synthetic steroid with specific antiglucocorticoid properties in the rat. 938 39

A single dose of 50 microg of trans-retinoic acid administered to rats significantly raised the level of hepatic tyrosine after a few hours. This effect was compared with that of dexamethasone and a possible correlation between these effectors was also investigated. An equal increase in enzyme activity level caused by retinoic acid was observed in adrenalectomized rats, leading to the suggestion that the effect of retinoic acid on liver tyrosine aminotransferase is independent of glucocorticoids. However, the study of the binding activity of the liver nuclear glucocorticoid receptors vs dexamethasone showed that this activity is favoured by retinoic acid, whereas no variation was evidenced for retinoic acid receptors caused by dexamethasone. In the adrenalectomized rat, the synergistic effect produced by the association of retinoic acid and dexamethasone at the lowest doses used led us to conclude that retinoic acid is an efficient effector of liver tyrosine aminotransferase. It probably affects tyrosine aminotransferase activity in a dependent and an independent way, regulated respectively by the glucorticoid status and by the provision of retinoic acid.
J Steroid Biochem Mol Biol
PMID:Correlation between the effects of retinoic acid and dexamethasone on liver tyrosine aminotransferase. 944 5

The winged helix transcription factor hepatocyte nuclear factor 3gamma (HNF3gamma) is expressed in embryonic endoderm and its derivatives liver, pancreas, stomach, and intestine, as well as in testis and ovary. We have generated mice carrying an Hnf3g-lacZ fusion which deletes most of the HNF3gamma coding sequence as well as 5.5 kb of 3' flanking region. Mice homozygous for the mutation are fertile, develop normally, and show no morphological defects. The mild phenotype change of the Hnf3g-/- mice can be explained in part by an upregulation of HNF3alpha and HNF3beta in the liver of the mutant animals. Analysis of steady-state mRNA levels as well as transcription rates showed that levels of expression of several HNF3 target genes (phosphoenolpyruvate carboxykinase, transferrin, tyrosine aminotransferase) were reduced by 50 to 70%, indicating that HNF3gamma is an important activator of these genes in vivo.
Mol Cell Biol 1998 Jul
PMID:Targeted disruption of the gene encoding hepatocyte nuclear factor 3gamma results in reduced transcription of hepatocyte-specific genes. 963 8

Fatty acid transport protein (FATP) was identified by expression cloning strategies (Schaffer, J. E., and Lodish, H. F. (1994) Cell 79, 427-436) and shown by transfection analysis to catalyze the transfer of long-chain fatty acids across the plasma membrane of cells. It is expressed highly in tissues exhibiting rapid fatty acid metabolism such as skeletal muscle, heart, and adipose. FATP mRNA levels are down-regulated by insulin in cultured 3T3-L1 adipocytes and up-regulated by nutrient depletion in murine adipose tissue (Man, M. Z., Hui, T. Y., Schaffer, J. E., Lodish, H. F., and Bernlohr, D. A. (1996) Mol. Endocrinol. 10, 1021-1028). To determine the molecular mechanism of insulin regulation of FATP transcription, we have isolated the murine FATP gene and its 5'-flanking sequences. The FATP gene spans approximately 16 kilobases and contains 13 exons, of which exon 2 is alternatively spliced. S1 nuclease and RNase protection assays revealed the presence of multiple transcription start sites; the DNA sequence upstream of the predominant transcription start sites lacks a typical TATA box. By transient transfection assays in 3T3-L1 adipocytes, the inhibitory action of insulin on FATP transcription was localized to a cis-acting element with the sequence 5'-TGTTTTC-3' from -1347 to -1353. This sequence is very similar to the insulin response sequence found in the regulatory region of other genes negatively regulated by insulin such as those encoding phosphoenolpyruvate carboxykinase, tyrosine aminotransferase, and insulin-like growth factor-binding protein 1. Fluorescence in situ hybridization analysis revealed that the murine FATP gene is localized to chromosome 8, band 8B3.3. Interestingly, this region of chromosome 8 contains a cluster of three other genes important for fatty acid homeostasis, lipoprotein lipase, the mitochondrial uncoupling protein 1 (UCP1) and sterol regulatory element-binding protein 1. These results characterize the murine FATP gene and its insulin responsiveness as well as present a framework for future studies of its role in lipid metabolism, obesity, and type II diabetes mellitus.
 ...
PMID:Characterization of the murine fatty acid transport protein gene and its insulin response sequence. 976 71

A broad specificity aminotransferase (BSAT), with high activity with both, aromatic amino acids and aspartate as substrates, was purified to homogeneity from promastigotes of Leishmania mexicana by a method involving chromatography on DEAE-cellulose, Red-120-Sepharose and Mono Q, and gel filtration on Sephacryl S-200. The purified enzyme showed a single band in SDS-polyacrylamide gel electrophoresis, with an apparent molecular mass of 45 kDa. Since the apparent molecular mass of the native enzyme, determined by gel filtration, was 90 kDa, the native enzyme is a dimer of similar subunits. The amino acid composition was determined, as well as the sequence of four internal peptides obtained by tryptic digestion. Two of these peptides, consisting of 49 amino acid residues in total, showed high similarity (57%) with corresponding sequences of plant aspartate aminotransferases, whereas they had only 33% identity with the aromatic aminotransferase of Escherichia coli, and 16% identity with the tyrosine aminotransferase from the related parasite Trypanosoma cruzi. The BSAT contained only one 1/2 Cys residue per monomer. The optimal pH for the enzyme reaction, with tyrosine and alpha-oxoglutarate as substrates, was 7.0. The apparent Km values for tyrosine, phenylalanine, tryptophan and glutamate, with oxaloacetate as co-substrate, were 1.3, 0.9, 0.9 and 171.8 mM, respectively; the value for aspartate with alpha-oxoglutarate as co-substrate was 2.5 mM, and that for alanine with alpha-oxoglutarate as co-substrate was 216 mM. The values for pyruvate, alpha-oxoglutarate and oxaloacetate, with tyrosine as co-substrate, were 5.6, 0.71 and 0.12 mM, respectively. These results suggest that the enzyme is a broad-specificity aminotransferase, able to transaminate the aromatic amino acids, aspartate, and to a lower extent alanine, with high sequence similarity to aspartate aminotransferases.
Mol Biochem Parasitol 1998 Oct 30
PMID:Isolation and partial characterization of a broad specificity aminotransferase from Leishmania mexicana promastigotes. 985 9

The glutamate synthetase gene (GS) contains a composite glucocorticoid response element (cGRE) comprised of a GRE and an adjacent element with features of both a cAMP-response element (CRE) and a 12-O-tetradecanoylphorbol 13-acetate (TPA) response element (TRE). The CRE/TRE element of the cGRE contributed to two modes of transcriptional activation: 1) enhancement of the response to cortisol and 2) a synergistic response to cortisol and increased cAMP. COS-7 cells transfected with a cGRE-luciferase construct show minimal expression under basal conditions or forskolin treatment. After cortisol treatment, luciferase activity from the cGRE is enhanced 4- to 8-fold greater than the GRE portion of the cGRE or a GRE from the tyrosine aminotransferase gene. Treatment with both forskolin and cortisol produced a 2- to 4-fold synergistic response over cortisol alone. Synergy is also seen with 8-bromo-cAMP, is specific for the cGRE, and occurs in a number of established cell lines. Elimination of the GRE or CRE/TRE reduces the synergy by 70-100%. Altering the CRE/TRE to GRE spacing changed both enhancement and synergy. Moving the elements 3 bp closer or extending 15 bp reduced enhancement. Synergy was markedly reduced when elements were one half of a helical turn out of phase. Western blots verified that CREB (cAMP-responsive binding protein) and ATF-1 (activating transcription factor-1) binds to the cGRE sequence. A specific dominant negative inhibitor of the CREB family, A-CREB, reduced synergy by 50%. These results suggest that the GS cGRE can potentially integrate signaling from both the cAMP and glucocorticoid receptor transduction pathways and that CREB/ATF-1 may play an important role in this process.
Mol Endocrinol 1999 Apr
PMID:Cyclic adenosine-3',5'-monophosphate-mediated activation of a glutamine synthetase composite glucocorticoid response element. 1019 61

Glucocorticoid and androgen receptors have been shown to function through the same palindromic glucocorticoid response element (GRE) and yet have differential effects on gene transcription. In this study, we examined the functional and structural relationship of the androgen and glucocorticoid receptors with the androgen responsive region (ARR) of the probasin (PB) gene containing two androgen receptor binding sites, ARBS-1 and ARBS-2. Transfection studies indicated that one copy of each cis-acting DNA element was essential for maximal androgen-induced chloramphenicol acetyltransferase (CAT) activity and that androgen selectivity was maintained when multiple copies of the minimal wild type (wt) androgen responsive region containing both ARBS-1 and ARBS-2 (-244 to -96) were subcloned in front of the thymidine kinase promoter. Furthermore, replacing the androgen response region with 1, 2 or 3 copies of either ARBS-1 or ARBS-2 restored less than 4% of the biological activity seen with the wt PB ARR. Multiple copies of either ARBS-1 or ARBS-2 did not result in glucocorticoid-induced CAT gene activity. By comparison, 1 or 2 copies of the tyrosine aminotransferase (TAT) GRE, as well as the mouse mammary tumour virus GRE, were strong inducers of CAT activity in response to both androgen and glucocorticoid treatment. In addition, band shift assays demonstrated that although the synthetic glucocorticoid receptor, GR-DNA binding domain (GR-DBD), and the synthetic androgen receptor, AR2, could interact with the TAT GRE (dissociation constants Kd of 63.9 and 14.1 respectively), only AR2 but not GR-DBD binding could be detected on ARBS-1 and ARBS-2. Our findings provide further evidence that androgen-induced regulation of gene transcription can occur through androgen-specific DNA binding sites that are distinct from the common GRE.
J Mol Endocrinol 1999 Jun
PMID:Selective activation of the probasin androgen-responsive region by steroid hormones. 1034 90

The natural steroid 11beta-hydroxyprogesterone is not only a modulator of 11beta-hydroxy-steroid dehydrogenase activity, but also an efficient inducer of tyrosine aminotransferase activity in hepatocytes. In contrast with the low affinity for the mineralocorticoid receptor. 11beta-hydroxyprogesterone binds well to both the glucocorticoid receptor and the carrier protein transcortin. It is accepted that the introduction of a 1:ene double bond into 3-keto 4:ene steroids increases the glucocorticoid potency, so that 3-keto-1,4:diene steroids show improved chemical stability and are more potent glucocorticoids than their respective 4:ene analogs. The steroid pregna-1,4-diene-11beta-ol-3,20-dione (deltaHOP) had previously been described as an anti-inflamatory compound and an inhibitor of macromolecular biosynthesis in thymocytes and lymphocytes. In such studies, deltaHOP also exhibited some particular glucocorticoid properties which made it attractive as a tool for the study of the mechanism of action of glucocorticoids. In the present paper we show that deltaHOP possesses some classical biological actions of glucocorticoids such as deposition of glycogen in rat liver, induction of TAT activity in hepatocytes, and inhibition of the uptake of leucine and thymidine by thymocytes. It also exhibits minimal sodium-retaining properties. Consistent with these biological effects, deltaHOP shows a 70 times lower relative binding affinity for the mineralocortioid receptor than aldosterone, but a reasonable affinity for the glucocorticoid receptor, and is as efficient as dexamethasone in dissociating the 90 kDa heat shock protein from the glucocorticoid receptor heterocomplex. However, the inhibition of the uptake of amino acids and nucleotides observed in the presence of deltaHOP is not efficiently blocked when thymocytes are coincubated in the presence of steroids with known antiglucocorticoid activity. deltaHOP is similarly inefficient in inducing chloramphenicol-acetyl transferase activity in cells transfected with a plasmid that possesses two canonical glucocorticoid-responsive elements. Unlike most glucocorticoids, deltaHOP does not induce the fragmentation of DNA in a regular pattern characteristic of apoptosis and it does not reduce thymus weight. This unusual dissociation of glucocorticoid parameters makes deltaHOP a useful tool to discriminate between mechanisms of action by which steroids can exert their biological effects.
Mol Cell Endocrinol 1999 Mar 25
PMID:The glucocorticoid properties of the synthetic steroid pregna-1,4-diene-11beta-ol-3,20-dione (deltaHOP) are not entirely correlated with the steroid binding to the glucocorticoid receptor. 1037 32


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