Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

We have previously reported that addition of 8-bromocyclic AMP enhances the stimulatory effect of dexamethasone on the expression of the angiotensinogen gene in mouse hepatoma cells in vitro. Isoproterenol is known to stimulate the synthesis of hepatic intracellular cyclic AMP via beta-adrenergic receptors. To study the possible effect of beta-adrenergic receptors on the expression of the angiotensinogen gene in mouse hepatoma cells, we transiently transfected them with a fusion gene with the 5'-flanking region of the angiotensinogen gene linked to a bacterial chloramphenicol acetyltransferase coding sequence as a reporter, pOCAT (ANG N-1498/+18). The addition of isoproterenol (10(-9) to 10(-5) mol/L) alone had no stimulatory effect on the expression of pOCAT (ANG N-1498/+18). In the presence of dexamethasone (10(-6) mol/L), however, isoproterenol enhanced the stimulatory effect on the dexamethasone on the expression of pOCAT (ANG N-1498/+18). The enhancing effect of isoproterenol was inhibited by the presence of propranolol (beta 1- and beta 2-adrenergic receptor antagonist) and ICI 118,551 (beta 2-adrenergic receptor antagonist) but not by the presence of atenolol (beta 1-adrenergic receptor antagonist). Furthermore, the addition of Rp-cAMP (an inhibitor of protein kinase A I and II) blocked the enhancing effect of isoproterenol. These studies demonstrated that isoproterenol enhances the stimulatory effect of dexamethasone on the expression of the angiotensinogen gene in mouse hepatoma cells via beta 2-adrenergic receptor and cyclic AMP-dependent protein kinase pathways. Our data may be important in understanding the molecular mechanism(s) of the stimulatory effect of catecholamines/glucocorticoid-induced expression of the angiotensinogen gene in the liver.
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PMID:Beta-adrenergic receptors and angiotensinogen gene expression in mouse hepatoma cells in vitro. 784 40

Previously, we reported that mesangial cells increased fibronectin, laminin and type IV collagen synthesis when cultured in the presence of high glucose (30 mM). Although mRNA levels for all three extracellular matrix (ECM) proteins were also increased in high glucose conditions, the mechanism for this increase was not known. In order to determine whether increased transcription was involved in the observed increase in fibronectin mRNA levels mesangial cells were transfected with a construct containing the 5'-flanking region of the fibronectin (FN) gene [position +69 to -510 base pairs (bp)] fused to the coding region of the chloramphenicol acetyltransferase (CAT) gene [FN-CAT (-510)]. Cells were transiently and stably transfected with this construct. Under serum-free conditions, high glucose increased CAT activity only in the presence of TGF beta 1 (referred to as TGF beta). The experiments were performed without serum because FN-CAT (-510) contains a serum responsive element. The increase in CAT was approximately twofold in transiently transfected cells and threefold in stably transfected cells. TGF beta alone increased CAT activity approximately 30%. Stimulation of fibronectin gene expression appeared to occur at the level of a cAMP response element (CRE) located -170 bp of the FN gene because cells transfected with a construct containing an oligonucleotide encoding for this CRE fused to a minimal fibronectin promoter (-56 bp) and a CAT reporter gene [CRE (-170) FN-CAT] displayed similar increments of CAT activity after treatment with high glucose and TGF beta. Gel shift mobility assays with a CRE oligonucleotide revealed multiple complexes with mesangial cell nuclear proteins.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:High glucose and TGF beta 1 stimulate fibronectin gene expression through a cAMP response element. 786 96

The growth regulatory activity of transforming growth factor-beta 1 (TGF beta 1) was studied in a clonal strain of thyroid papillary carcinoma cell (NPA). Despite the presence of TGF beta 1 and its receptor messenger RNA in thyroid carcinoma, the molecular mechanism of TGF beta 1 action on cell growth of thyroid carcinoma has not yet been elucidated. Exogenously added TGF beta 1 inhibited DNA synthesis and cell growth in a dose- and time-dependent manner at concentrations of 0.1-10 ng/ml. TGF beta 1 inhibited not only basal but also fetal bovine serum-stimulated cell proliferation. Steady state levels of c-myc messenger RNA transcripts were inhibited by TGF beta 1 after 0.5-h treatment. Antisense, but not sense, c-myc oligodeoxynucleotides also caused suppression of NPA cell growth in a dose-responsive manner. Transfection studies of the 5'-up-stream flanking region (UFR) of c-myc/chloramphenicol acetyltransferase chimera genes suggest the presence of a TGF beta 1-responsive DNA element in the 2.3-kilobase c-myc 5'-UFR. Deletion mutant studies indicate the element lies between -106 to 70 relative to the P1 transcription start site. Studies with the gel mobility shift assay using 23-basepair double strand DNA showed the presence of at least two nuclear factors in NPA cell. TGF beta 1 treatment did not cause any alteration in TGF beta 1-induced mobility; however, the reduction of a positive band was selectively observed during 30 min to 2 h after treatment with TGF beta 1. In contrast, the position and intensity of another band were not altered by TGF beta 1 treatment. These results demonstrate that the inhibition of a nuclear factor binding to the c-myc 5'-UFR and subsequent suppression of c-myc gene expression are directly involved in the antiproliferative action of TGF beta 1 in NPA cell growth.
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PMID:Correlation between suppression of c-myc and antiproliferative effect of transforming growth factor-beta 1 in thyroid carcinoma cell growth. 792

Transforming growth factor beta (TGF-beta) is a potent modulator of cell growth in many systems. In normal rat kidney fibroblasts, TGF-beta 1 increases epidermal growth factor (EGF) receptor gene transcription and synergizes with EGF to stimulate growth in soft agar, a characteristic of the transformed phenotype. In order to identify the target of TGF-beta 1 action, we have used a series of 5' deletion mutants of the EGF receptor promoter linked to a chloramphenicol acetyltransferase reporter gene (ERCAT). The TGF-beta response element(s) was localized to a cis-regulatory region which resides between positions -919 and -860 relative to the ATG translation initiation codon of the EGF receptor promoter. This 60-base pair region contains a repressor of the EGF receptor promoter and a TGF-beta inhibitory element that mediates TGF-beta 1 suppression of transin/stromelysin gene transcription through binding of a Fos-containing protein complex. Cotransfection of c-fos, c-jun, or both expression vectors with the intact or 5'-deleted ERCAT constructs identified several Fos-responsive inhibitory regions within the EGF receptor promoter, but these did not localize to the -919 to -860 promoter region. Mobility shift assays showed binding of the 60-base pair DNA fragment to proteins in extracts from untreated normal rat kidney cells; the binding was specifically competed by oligonucleotides containing a CAGATG sequence but not by oligonucleotides containing the EGF receptor repressor or the TGF-beta inhibitory element. TGF-beta 1 treatment but not anti-Fos antibody caused a decrease in specific 60-base pair DNA-protein complex formation.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Induction of epidermal growth factor receptor gene transcription by transforming growth factor beta 1: association with loss of protein binding to a negative regulatory element. 798 46

Autoregulation of the human thyroid hormone receptor beta 1 (hTR beta 1) promoter was assessed by chloramphenicol acetyltransferase and luciferase reporter assays of transient transfections into COS1 and GH3 cells, DNase I footprinting, and gel shift assays. A 5'-deletional analysis of the promoter showed that the region between -906 and -839 and the sequence from -438 to -130 were positively regulated by T3 in COS1 cells cotransfected with an hTR beta 1 expression vector. We also transfected deletion constructs into GH3 cells and showed similar effects of T3 on the trans-activation of the reporters. DNase I footprinting showed a protected inverted palindromic thyroid response element (TRE) at position -890 to -866 in the distal fragment and a direct repeat at position -190 to -166 in the proximal fragment, which were protected by TRs. Mutation of each TRE significantly decreased the trans-activation of the promoter by T3. Gel mobility shift assays showed both proximal and distal TREs formed a retarded band with hTR alpha 1 or hTR beta 1 expressed in COS1 cells and reticulocyte lysates. The bands formed on the distal TRE and the proximal TRE appear to be preferentially formed by a TR homodimer and a heterodimer, respectively. Furthermore, the band formed on the distal TRE disappeared after adding T3 but that on the proximal TRE did not. These results indicate that hTR beta 1 expression is directly regulated by hTR alpha 1, beta 1, and their ligand through two TREs. The different structure of the TREs in this promoter suggests their physiological role in transcriptional regulation may be different.
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PMID:Two thyroid hormone response elements are present in the promoter of human thyroid hormone receptor beta 1. 801 48

Transgenic mice were used to locate the cis-acting DNA elements that are essential for tissue-specific and inducible expression of the rat proline-rich protein gene, R15. Chimeric genes with up to 10 kb of R15 5'-flanking region fused to chloramphenicol acetyltransferase (CAT) or polyomaviral large T-antigen (PyLT) reporter genes were tested. Our results demonstrate that (1) the isoproterenol/tannin-inducible, parotid-specific transgene expression requires an upstream cis-regulatory domain, namely the parotid control region, which extends from -6 to -1.7 kb of the R15 gene; (2) this parotid control region functions with a heterologous promoter and is indispensable for achieving a reproducible chromosomal position-independent transgene expression; (3) deletion of the R15 5'-flanking region up to -1.7 kb results in a pleiotropic effect on the transgene expression, which includes ectopic (nonsalivary) reporter expression and lack of inducibility by either the beta-agonist isoproterenol or dietary tannin stimulation; (4) when the -10 to -6 kb region from the R15 gene is deleted in the construct, the inducible expression in the parotid glands of the transgenic mice decreases by over 30-fold, but position-independent and tissue-specific transgene expression is retained. Moreover, the mechanism of induction by either catecholamine isoproterenol or dietary tannin appears to be through a beta 1-adrenergic receptor-mediated pathway for both normal (non-transgenic) and transgenic animals.
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PMID:Isoproterenol/tannin-dependent R15 expression in transgenic mice is mediated by an upstream parotid control region. 801 29

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

To gain a further understanding of the regulation of human type I collagen gene expression under physiologic and pathologic conditions, we characterized 5.3 kilobase pairs (kb) of the human alpha 1(I) procollagen gene promoter. A series of deletion constructs containing portions of the alpha 1(I) procollagen 5'-flanking region (with end points from -5.3 kb to -84 base pairs (bp)) ligated to the chloramphenicol acetyltransferase (CAT) reporter gene were transiently transfected into NIH/3T3 cells. Maximal CAT activity was observed with constructs having 5' end points from -804 to -174 bp. A further 5' deletion to -84 bp caused a marked reduction in CAT activity. Cells transfected with plasmids containing longer 5'-flanking fragments of the alpha 1(I) procollagen gene (-2.3 or -5.3 kb) showed reduced CAT activity compared with the -804 bp construct. The activity of the alpha 1(I) procollagen promoter was much lower in cells that do not normally express type I collagen (HeLa cells) compared with collagen-producing NIH/3T3 cells. The CAT activity of deletion constructs containing longer 5' regions than -84 bp was increased by approximately 2-fold in NIH/3T3 cells treated with transforming growth factor beta 1 (TGF beta 1). Electrophoretic mobility shift assays indicated that protein-DNA complex formation with a probe corresponding to the -170 to -80 bp fragment of the alpha 1(I) procollagen promoter was markedly enhanced in nuclear extracts prepared from TGF beta 1-treated fibroblasts as compared with untreated fibroblasts. The DNA binding activity stimulated by TGF beta 1 was specific for an Sp1-like sequence at positions -164 to -142 bp in the promoter. These results demonstrate that 1) there are both positive and negative cis-acting regulatory elements in the human alpha 1(I) procollagen promoter, 2) these regulatory regions function differently in collagen-producing and -nonproducing cells, 3) the alpha 1(I) procollagen promoter contains TGF beta 1-responsive sequences located between -174 and -84 bp from the transcription start site, and 4) TGF beta 1 caused marked stimulation of the DNA binding activity of a nuclear factor interacting with an Sp1-like binding site located within a region encompassing -164 to -142 bp of the alpha 1(I) procollagen promoter.
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PMID:Functional analysis of human alpha 1(I) procollagen gene promoter. Differential activity in collagen-producing and -nonproducing cells and response to transforming growth factor beta 1. 817 78

Expression of the Na(+)-K(+)-adenosinetriphosphatase (ATPase) gene family in rat intestinal epithelial cells was examined using RNA blot hybridization analyses. Rat intestinal epithelial cells express only the alpha 1- and beta 1-subunit mRNAs. A gradient in expression of alpha 1- and beta 1-subunit mRNA was seen along the villus-crypt unit in both jejunum and ileum, i.e., villus tip >> crypt cells. Regional differences in expression were observed along the intestine. alpha 1- and beta 1-subunit mRNA abundance was similar in jejunum, ileum, and colon while enzymatic activity was highest in the jejunum and lowest in the ileum. Administration of thyroid hormone to thyroidectomized rats increased the expression of alpha 1- and beta 1-subunit mRNAs in jejunum but not in colon. Hypothyroidism had no effect on subunit mRNA expression. The human intestinal cell line Caco-2 was also studied. These cells also expressed only the alpha 1- and beta 1-isoform mRNAs and demonstrated a developmental profile in both mRNA and enzymatic activity. Furthermore, in Caco-2 cells both alpha 1- and beta 1-mRNAs and Na(+)-K(+)-ATPase enzymatic activity were stimulated by thyroid hormone. Caco-2 cells transfected with 5' flanking regions of the human Na(+)-K(+)-ATPase beta 1-gene linked to the chloramphenicol acetyltransferase (CAT) reporter gene responded to 3,5,3'-triiodothyronine (T3) treatment with increased expression of CAT activity. This suggests that the 5' flanking region of the beta 1-gene contains a thyroid hormone response element and that T3 upregulation occurs at the transcriptional level.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Na(+)-K(+)-ATPase gene expression in rat intestine and Caco-2 cells: response to thyroid hormone. 823 61

C-erbA receptors and v-erbA have been shown to functionally interact with 12-O-tetradecanoyl-phorbol-13-acetate (TPA)-inducible gene expression. These proteins enhance trans-activation by c-jun, and the c-erbA receptors in the presence of thyroid hormone repress TPA and c-jun induction of transcription. Also, v-erbA can abrogate T3-mediated repression. We have examined how dominant negative (S and CL) and nondominant negative (G-H) receptors cloned from various patients with thyroid hormone resistance syndromes affect expression of the collagenase promoter induced with TPA. The CL receptor (ARG315HIS mutation) has a 2-fold reduction in T3-binding affinity compared with human c-erbA beta 1 wild-type (WT) receptor, whereas the G-H receptor (ARG311HIS) and S receptor (deletion, THR codon 332) have T3-binding affinities reduced by 100-fold and greater than 100-fold, respectively. These mutant receptors were cotransfected with a collagenase promoter (-1200 to +63 base pairs) chloramphenicol acetyltransferase reporter gene (Col-CAT) into COS-7 cells. Levels of CAT reporter gene expression after transient transfection were determined in the presence or absence of 3-10 nM T3 and the presence or absence of 100 nM TPA. Unoccupied CL receptor and G-H and S receptors stimulated TPA-induced Col-CAT expression 1.5- to 9-fold. The CL receptor with thyroid hormone totally repressed TPA induction of the collagenase receptor. In the presence of thyroid hormone, the enhancing effects by S and G-H receptors on TPA-induced Col-CAT expression were unaffected and minimally diminished, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Dominant and nondominant negative C-erbA beta 1 receptors associated with thyroid hormone resistance syndromes augment 12-O-tetradecanoyl-phorbol-13-acetate induction of the collagenase promoter and exhibit defective 3,5,3'-triiodothyronine-mediated repression. 824 13


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