Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

3T3-F442A adipocytes express the gene encoding cytosolic phosphoenolpyruvate carboxykinase (GTP) (PEPCK). Retinoic acid (RA) caused a 5-fold induction of PEPCK mRNA within 6 h in these cells with a half-maximal effective concentration of approximately 75 microM. This effect was independent of cycloheximide and inhibited by actinomycin D. In vitro run-on experiments using isolated nuclei confirmed that the RA-induced increase was mainly due to an increased rate of transcription of the gene. Stable transfectants bearing either the region of the PEPCK promoter from -2100 to +69 fused to the chloramphenicol acetyltransferase (CAT) gene (pPL1-CAT) or -600 to +69 fused to CAT (pPL9-CAT) were used to study PEPCK gene regulation during differentiation. The same transfected cells were used to analyse the RA effect. Preadipocytes containing pPL1-CAT expressed a much lower level of CAT activity than did adipocytes. pPL9-CAT was not expressed in either preadipocytes or adipocytes. RA induced the expression of CAT activity in preadipocytes and adipocytes transfected with pPL1-CAT, but had no effect in cells transfected with pPL9-CAT. These results suggest that one or more DNA sequences located between -2100 and -600 bp of the PEPCK promoter is required for adipocyte-specific expression of this gene. RA action is independent of the state of differentiation and appears to require different elements in fat cells from those required in liver.
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PMID:Expression of the phosphoenolpyruvate carboxykinase gene in 3T3-F442A adipose cells: effects of retinoic acid and differentiation. 794 24

The cDNA for a member of the nuclear receptor family was cloned and named ubiquitous receptor (UR), since UR protein and mRNA are detected in many cell types. Rat UR/human retinoid X receptor alpha (hRXR alpha) heterodimers bound preferentially to double-stranded oligonucleotide direct repeats having the consensus half-site sequence AGGTCA and 4-nt spacing (DR-4). Coexpression of UR in COS-1 cells inhibited the stimulation of chloramphenicol acetyltransferase (CAT) reporter gene expression by hRXR alpha and human retinoic acid receptor alpha in the presence of all-trans-retinoic acid when DR-4 (but not DR-5) was present upstream of the promoter of a CAT reporter gene (DR-4-CAT). UR expression also inhibited the activation of a DR-4-CAT reporter gene by hRXR alpha and 9-cis-retinoic acid or by thyroid hormone receptor beta in the presence of thyroid hormone. However, in the absence of 9-cis-retinoic acid, UR in combination with hRXR alpha stimulation DR-4-CAT expression. Coexpression of thyroid hormone receptor markedly reduced this stimulation in the absence of thyroid hormone. UR may play an important role in normal growth and differentiation by modulating gene activation in retinoic acid and thyroid hormone signaling pathways.
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PMID:Ubiquitous receptor: a receptor that modulates gene activation by retinoic acid and thyroid hormone receptors. 797 66

Functional antagonism between retinoic acid (RA) receptors and activator protein-1 (AP-1) transcription factors might regulate expression of genes involved in the response to injury in the kidney. We designed experiments to analyze the mechanisms by which RA inhibits AP-1-directed transcriptional responses in glomerular mesangial cells. RA inhibited serum-stimulated mesangial cell proliferation as assessed by measurements of [3H]thymidine uptake and cell number. In transient transfection assays with a chloramphenicol acetyltransferase reporter, RA completely blocked transcription directed by an AP-1 cis-element in cells stimulated by serum. AP-1 DNA binding was analyzed in electrophoretic gel mobility shift assays using nuclear extracts from control or RA-pretreated cells stimulated with serum. RA did not abolish AP-1 DNA binding activity under the conditions of this assay. The apparent equilibrium dissociation constant, maximal density of binding, and association rate for the AP-1-DNA interaction were similar in serum-stimulated cells or RA-pretreated cells stimulated with serum. RA repressed serum-stimulated induction of the immediate early genes c-fos and c-jun, whose protein products dimerize to form AP-1. Repression was relatively selective for c-fos/c-jun; induction of other immediate early transcription factors (junB, c-myc, and egr-1) was not downregulated by RA. That repression of c-fos by RA might contribute to anti-AP-1 activity was suggested by experiments with an antisense c-fos expression vector, which demonstrated that c-fos induction was required for serum-stimulated AP-1 activity. Together, these data demonstrate that RA antagonizes AP-1-directed transcription without inhibiting AP-1 DNA-binding in mesangial cells. Selective repression of c-fos and c-jun might contribute to the anti-AP-1 activity of RA.
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PMID:Anti-AP-1 activity of all-trans retinoic acid in glomerular mesangial cells. 797 84

Many hormones regulate the rate of synthesis of phosphoenolpyruvate carboxykinase (PEPCK), the enzyme that governs the rate-limiting step in gluconeogenesis. In H4IIE rat hepatoma cells, glucocorticoids, retinoic acid and cyclic AMP (cAMP) increase PEPCK gene transcription whereas insulin and phorbol esters have the opposite effect. Insulin and phorbol esters are dominant as they prevent cAMP- and glucocorticoid-stimulated PEPCK gene transcription. In contrast, insulin and phorbol esters both stimulate transcription of gene 33 in the same H4IIE cells, with the same time course as seen for their inhibitory effect on PEPCK gene transcription. We now report that the protein phosphatase inhibitor, okadaic acid, mimics the action of insulin and phorbol esters on expression of both gene 33 and PEPCK gene in H4IIE cells. Okadaic acid stimulates gene 33 mRNA accumulation whereas it inhibits cAMP- and glucocorticoid-stimulated PEPCK mRNA accumulation. The effect of okadaic acid on the PEPCK gene is mediated through the PEPCK promoter as, in a cell line, HL1C, stably transfected with a PEPCK-chloramphenicol acetyltransferase (CAT) fusion gene, okadaic acid inhibits cAMP- and glucocorticoid-stimulated CAT expression. Desensitization of the protein kinase C pathway by exposure to phorbol 12-myristate 13-acetate for 16 h abolishes the subsequent action of the phorbol ester but does not markedly affect the inhibition of cAMP- and glucocorticoid-stimulated CAT expression by insulin or okadaic acid. Even though insulin and okadaic acid appear to repress PEPCK gene expression through a pathway initially distinct from that used by phorbol esters, transient-transfection studies show that the final target of the action of okadaic acid, insulin and phorbol ester is the same DNA element.
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PMID:Comparison of the effects of insulin and okadaic acid on phosphoenolpyruvate carboxykinase gene expression. 798 Apr 40

Cytosolic phosphoenolpyruvate carboxykinase (GTP) (PEPCK) is a key glyceroneogenic enzyme in adipose tissue. The regulation of PEPCK gene expression by retinoic acid (RA) and dexamethasone (DEX) was studied in 3T3-F442A adipocytes maintained in a serum-free medium. RA induced whereas DEX reduced PEPCK mRNA steady-state level. RA stimulation was about 4-fold and DEX repression was of 80% in 4 hrs. In addition to reducing basal mRNA level, DEX was able to counteract RA induction in a dominant manner. The use of the glucocorticoid antagonist RU 38486 indicated that the DEX effect was mediated by the glucocorticoid receptor. Stable transfectants bearing the region of the PEPCK promoter from -2100 to +69 fused to the chloramphenicol acetyltransferase (CAT) gene (pPL1-CAT) were used to study PEPCK gene regulation in differentiated adipocytes. In such cells, RA stimulated CAT expression 3 to 5.5 fold. DEX had no effect on basal CAT activity whereas it inhibited the stimulation induced by RA. Thus, in adipocytes, the PEPCK gene regulatory region between -2100 and +69 bp mediates both stimulation by RA and repression by DEX of RA action.
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PMID:Glucocorticoids antagonize retinoic acid stimulation of PEPCK gene transcription in 3T3-F442A adipocytes. 798 26

When human promyelocytic leukemia cell line HL60 was treated with retinoic acid (RA), considerable suppression of protooncogene myc expression was achieved before granulocytic differentiation became evident. From transient transfection experiments using the reporter plasmid containing exon 1 and its 2.3 kilobases upstream of the c-myc gene fused to the chloramphenicol acetyltransferase gene, it was indicated that this suppression was mainly attributable to the level of transcription initiation. Deletion down to 95 base pairs upstream of the P2 promoter did not change the suppressive effect of RA on c-myc gene expression. Mobility shift assays with respect to the P2 promoter region revealed that the 15-base pair fragment located between P1 and P2 promoters was responsive to the RA treatment. This fragment included the E2F binding site in the c-myc P2 promoter region, and a difference of shifted bands between RA-treated and untreated HL60 cells was due to complex formation of E2F and retinoblastoma protein. The present results suggest that E2F plays an important role in the process of cell differentiation by RA and that a change of the E2F binding pattern induced by RA contributes to the suppression of c-myc gene expression preceding granulocytic differentiation.
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PMID:Transcription from the P2 promoter of human protooncogene myc is suppressed by retinoic acid through an interaction between the E2F element and its binding proteins. 801 61

To understand the molecular basis of the phosphorylation-enhanced transcriptional activity of human thyroid hormone nuclear receptor subtype beta 1 (hTR beta 1), we studied the effect of phosphorylation on the interaction of hTR beta 1 with the retinoid X receptor beta (RXR beta), we studied the effect of phosphorylation on the interaction of hTR beta 1 with the retinoid X receptor beta (RXR beta). In vitro, the extent of hTR beta 1.RXR beta heterodimer bound to various thyroid hormone response elements (TREs) was compared before and after phosphorylation of hTR beta 1. Without phosphorylation, hTR beta 1.RXR beta heterodimer was barely detectable under the experimental conditions. After phosphorylation of hTR beta 1, heterodimer bound to (i) the chicken lysozyme gene TRE, (ii) a TRE consisting of direct repeats of half-site binding motifs separated by four gaps, and (iii) a palindromic TRE was enhanced by approximately 10-, 7-, and 6-fold, respectively. The effect of phosphorylation on hTR beta 1.RXR beta heterodimerization was reversible. Dephosphorylation of the phosphorylated hTR beta 1 by alkaline phosphatase led to loss of the ability of hTR beta 1 to form a heterodimer with RXR beta in either the absence or the presence of DNA. These results indicate that the heterodimerization is enhanced by phosphorylation. To evaluate the effect of phosphorylation on the interaction of hTR beta 1 with RXR beta in vivo, we cotransfected hTR beta 1, RXR beta and TRE-chloramphenicol acetyltransferase (CAT) expression plasmids into CV-1 cells. CAT activity was assessed in the presence or absence of okadaic acid. Okadaic acid is a potent inhibitor of phosphatases 1 and 2A and increases the in vivo phosphorylation of hTR beta 1 by approximately 10-fold. Using the CAT reporter gene under control of the TRE from the malic enzyme gene, we found that RXR beta increased the okadaic acid-enhanced hTR beta 1-mediated CAT activity by 2- to 3-fold in the presence of 3,3',5-triiodo-L-thyronine. However, 9-cis-retinoic acid did not enhance the effect of okadaic acid. Our results indicate that phosphorylation is essential for the interaction of hTR beta 1 with RXR beta. Thus, phosphorylation plays a pivotal role in the gene-regulating activity of hTR beta 1.
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PMID:Phosphorylation enhances the target gene sequence-dependent dimerization of thyroid hormone receptor with retinoid X receptor. 805 36

The molecular mechanisms by which expression of a gene is down-regulated after differentiation of F9 embryonal carcinoma cells into parietal endoderm-like cells was studied by characterizing the cis- and trans-regulatory elements of the gb110 gene. This gene encodes a putative RNA helicase, and its expression is down-regulated when F9 cells are differentiated with retinoic acid and cyclic AMP. The 5'-flanking region of the gene has all of the features of a GC-rich island promoter and seems to play only a minor role, if any, in the regulated expression. A 133-bp enhancer in the first intron was identified by transient chloramphenicol acetyltransferase assays that activated expression in undifferentiated F9 cells about 50- to 100-fold. As this enhancer was not active in differentiated F9 cells, it seems to be the prime mediator of the differentiation-specific down-regulation of the gb110 gene. Four different protein-binding sites, three of which contain GC- and GT-box motifs, were identified in the enhancer element. The fourth site, interacting with previously described transcription factor FTZ-F1/ELP, seems to be of minor importance for the activity of the enhancer. Mutational analysis showed that the cooperative interaction of several most likely related proteins with the three GC- and GT-box motifs was required for full enhancer activity. On the basis of their binding properties, at least two of these proteins seem to be identical or closely related to ubiquitous transcription factor Sp1. One of the GT-box-binding proteins was present in undifferentiated F9 cells but not, however, in its differentiated derivatives. The cell specificity of this transcription factor explains why the gb110 gene is not expressed or expressed only at low levels in parietal endoderm-like cells.
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PMID:Interaction of several related GC-box- and GT-box-binding proteins with the intronic enhancer is required for differential expression of the gb110 gene in embryonal carcinoma cells. 806 13

In this study, we demonstrated that retinoic acid (RA) up-regulated interleukin-2 receptor-alpha (IL-2R alpha) expression on two human B-cell lines, IE8.6 and SKW6.4. Deleted forms of the human IL-2R alpha promoter linked to the bacterial chloramphenicol acetyltransferase reporter gene were transfected into IE8.6 cells in order to define RA-responsive regulatory domains. Experiments using the -1.6 kb construct, which contains all known regulatory regions in the IL-2R alpha promoter, indicated that RA could induce IL-2R alpha promoter activity. The basal activity of the -471 construct was initially low, but was markedly enhanced by the addition of RA. Deletion of promoter sequences between -471 and -317 resulted in a significant augmentation of basal promoter activity and abolished promoter induction by RA. This finding revealed a requirement for sequences 5' of base -317 for RA-induced promoter activation, raising the possibility of the presence of both a RA response element and a negative regulatory element (NRE) upstream of base -317. Transfection studies with internal deletion mutants with the putative NRE removed resulted in increases in basal promoter activity and unresponsiveness to RA similar to the -317 construct. In contrast, an internal deletion mutant with the NRE intact had low basal activity and was inducible by RA similar to the -471 construct. Taken together, our results suggested that RA-induced activation of the IL-2R alpha promoter was through changes in the function of a NRE present between bases -400 and -368. This 31-base pair element may interact with an adjacent RA-responsive regulatory site as well as being responsible for down-regulation of basal IL-2R alpha expression under certain conditions.
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PMID:Transcriptional regulation by retinoic acid of interleukin-2 alpha receptors in human B cells. 815 76

We evaluated SR11237, a retinoid X receptor (RXR)-specific compound, for its pharmacologic effects on cell differentiation in F9 embryonal carcinoma cells and rhino mouse epidermis. SR11237 can cause RXR/RXR homodimers to form and transactivate a reporter gene containing a RXR-response element. We confirmed, using nuclear receptor co-transfection assays in COS-1 cells, that SR11237 is effective at transactivating a chloramphenicol acetyltransferase reporter gene through RXRs but not retinoic acid receptors. When SR11237 was tested for its ability to modulate cell differentiation, it was inactive on F9 embryonal carcinoma cells and rhino mouse skin. Because differentiation in these systems is known to be regulated by RAR-specific compounds, such as all-trans-retinoic acid and (E)-4-[2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)-1-prope nyl benzoic acid], our results with SR11237 are compatible with the concept that classical retinoid pleiotropic responses are mediated by RXR/RAR heterodimeric nuclear receptors rather than through RXR/RXR homodimers.
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PMID:A pleiotropic response is induced in F9 embryonal carcinoma cells and rhino mouse skin by All-trans-retinoic acid, a RAR agonist but not by SR11237, a RXR-selective agonist. 817 47


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