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)

Two overlapping clones spanning 19 kilobase pairs (kb) of the 5' end of the alpha 1 (IV) collagen gene were isolated and found to contain a single exon which encoded the 5'-untranslated sequence and 84 base pairs of the signal peptide. The 5' end of this exon was determined to be the 5' end of the transcript by S1 nuclease protection and primer extension. The nucleotide sequence of 1 kb of the 5'-flanking DNA was extremely G + C-rich (greater than 70%) and contained two GC boxes and a putative cAMP regulatory sequence. The transcriptional regulation of the alpha 1 (IV) gene was studied with chimeric gene constructs utilizing 2.5 kb of the 5'-flanking sequence coupled to the gene for chloramphenicol acetyltransferase. Transfection of this construct into differentiating F9 cells resulted in low chloramphenicol acetyltransferase activity compared to beta-actin or Rous sarcoma virus long terminal repeat promoters, although these cells produce large amounts of collagen IV. Inclusion of a 2.7-kb sequence 2.3 kb downstream from the first exon in either orientation increased the transcription of the chloramphenicol acetyltransferase construct approximately 10-fold in F9 cells, but was not active in NIH 3T3 cells, which synthesize little collagen IV. These results indicate the presence of an enhancer within the first intron, which increases the expression of this gene.
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PMID:Characterization of the promoter for the alpha 1 (IV) collagen gene. DNA sequences within the first intron enhance transcription. 284 28

cAMP and phorbol esters mediate cellular metabolism by the activation of distinct signal transduction pathways consisting of a cascade of sequential protein phosphorylations. An important consequence of the activation of these pathways is the stimulation of gene transcription by way of interactions of specific proteins with DNA control elements. The 8-base-pair (bp) DNA consensus sequence TGACGTCA [cAMP response element (cAMP-RE)] has been shown to confer cAMP responsivity on transcription from various promoters, and the closely related 7-bp consensus sequence TGA-(C or G)TCA [phorbol 12-myristate 13-acetate response element (PMA-RE)] lends transcriptional responsiveness to phorbol esters. In the JEG-3 placental cell line we find that several variants of the cAMP-REs fused to a gonadotropin alpha promoter chloramphenicol acetyltransferase reporter gene mediate responsiveness to cAMP but not to phorbol esters. The PMA-RE is responsive to phorbol esters but also imparts submaximal sensitivity to cAMP in the JEG-3 cells and in the Hep G2 hepatoma cell line. The transcriptional activities of cAMP-RE and PMA-RE are markedly influenced by the composition of the neighboring bases, but different sequences are permissive for the activity of the cAMP-RE versus the PMA-RE. The two signaling agents together display a supraadditive effect on reporter genes containing active PMA-REs but not cAMP-REs. Gel-mobility-shift and UV cross-linking analyses show that distinct proteins bind to the two control elements. One protein of 38 kDa binds to the cAMP-RE and several proteins of 48-84 kDa bind to the PMA-RE.
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PMID:Cyclic AMP and phorbol ester-stimulated transcription mediated by similar DNA elements that bind distinct proteins. 284 47

A transcriptional cAMP-responsive enhancer element (CRE) consisting of the 8-base pair (bp) palindrome, 5' TGACGTCA 3', is found in several eukaryotic genes. We analyzed the effects on gene transcription of point mutations within the CRE, the influence of the bases surrounding the CRE, and the requirements for transcriptional synergism of tandemly repeated CREs. When inserted as an oligonucleotide with restriction enzyme linker sites, the 8-bp CRE itself is as active in conferring cAMP responsivity on an enhancerless chloramphenicol acetyltransferase reporter plasmid as is a single copy of the choriogonadotropin alpha (CG alpha), twice repeated 18-bp sequence containing the CRE. Point mutations in the first (T to A), fourth (C to G), or eighth (A to T) positions of the CRE, when contained within the CG alpha 18-bp sequence, each inhibited transcriptional activity greater than 90%. However, the identical eighth position A to T mutation occurs in the cAMP-responsive sequence of the vasoactive intestinal peptide (VIP) gene, and that mutant sequence in the context of the adjacent bases of the native VIP sequence is maximally cAMP responsive when inserted in the reporter plasmid. The substantially reduced activity of the core 8-bp CRE when synthesized as a cassette including the adjacent bases of the rat glucagon or bovine parathyroid hormone gene further emphasizes the restrictive influence of particular surrounding sequences. Active oligonucleotides containing the 8-bp palindrome and different but equally permissive contexts have comparable properties in transfected reporter genes and gel mobility-shift assays. The pair of tandemly repeated 18-bp elements containing the CRE in the CG alpha gene synergistically stimulate transcription either with paired native CREs or when one native CRE is paired with one mutant CRE, suggesting the presence of cooperative interactions. Tandem insertion of more than two 18-bp sequences, or insertion of additional sequences between the two CREs, inhibits transcription. These observations indicate that the contexts of the bases adjacent to CREs exert profound influences on the transcriptional activities mediated by the cAMP-responsive elements.
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PMID:Structural determinants for transcriptional activation by cAMP-responsive DNA elements. 284 37

The molecular mechanisms involved in the regulation of expression of the rat CRH gene have been examined in rat pheochromocytoma (PC-12) cells transiently transfected with a chimeric gene containing 1.4 kilobases of rat CRH 5'-flanking DNA fused to the bacterial reporter gene encoding chloramphenicol acetyltransferase. Cyclic AMP analogs and activators of adenylate cyclase positively regulate the expression of this chimeric gene in PC-12 cells, inducing chloramphenicol acetyltransferase activity more than 15-fold. The DNA sequence required for this response to cAMP has been localized to a 59 base pair region located between 238 and 180 base pairs 5' to the putative CRH mRNA cap site. This sequence can confer cAMP-responsiveness on a heterologous promoter in an orientation independent fashion and has homology to cAMP regulatory regions from a number of other eukaryotic genes.
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PMID:Identification of a cyclic adenosine monophosphate-responsive element in the rat corticotropin-releasing hormone gene. 285 Nov 1

We have examined the regulation of somatostatin gene expression by cAMP in PC12 rat pheochromocytoma cells transfected with the rat somatostatin gene. Forskolin at 10 microM caused a 4-fold increase in somatostatin mRNA levels within 4 hr of treatment in stably transfected cells. Chimeric genes containing the somatostatin gene promoter fused to the bacterial reporter gene encoding chloramphenicol acetyltransferase were also induced by cAMP in PC12 cells. To delineate the sequences required for response to cAMP, we constructed a series of promoter deletion mutants. Our studies defined a region between 60 and 29 base pairs upstream from the transcriptional initiation site that conferred cAMP responsiveness when placed adjacent to the simian virus 40 promoter. Within the cAMP-responsive element of the somatostatin gene, we observed an 8-base palindrome, 5'-TGACGTCA-3', which is highly conserved in many other genes whose expression is regulated by cAMP. cAMP responsiveness was greatly reduced when the somatostatin fusion genes were transfected into the mutant PC12 line A126-1B2, which is deficient in cAMP-dependent protein kinase 2. Our studies indicate that transcriptional regulation of the somatostatin gene by cAMP requires protein kinase 2 activity and may depend upon a highly conserved promoter element.
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PMID:Identification of a cyclic-AMP-responsive element within the rat somatostatin gene. 287 59

Previous studies have shown that differentiation of 3T3-L1 preadipocytes leads to the activation of transcription of an unidentified gene which encodes a 4.9-kilobase (kb) mRNA. Several cDNAs that include the complete sequence of this mRNA were obtained and used to isolate and characterize the gene. Analysis of the nucleotide and amino acid sequences of both cDNA and genomic clones revealed that the gene encodes the mouse stearoyl-CoA desaturase (SCD), an enzyme known to be expressed upon differentiation of 3T3-L1 preadipocytes. The predicted amino acid sequence (355 residues) of the mouse 3T3-L1 adipocyte SCD exhibits 92% identity to that of the rat liver SCD. There is also a high degree of nucleotide sequence identity between the mouse and rat mRNAs in their unusually long approximately 3.5-kb 3'-untranslated regions. Mice fed a diet containing unsaturated triacylglycerides express SCD mRNA only in adipose tissue, whereas mice starved and refed a fat-free diet, express SCD mRNA in both liver and adipose tissue. The mouse gene for the desaturase spans approximately 15 kb and contains 6 exons and 5 introns with all intron-exon junctions conforming to the GT/AG splicing rule. As determined by S1 nuclease mapping and primer extension analysis, the transcriptional initiation site maps 152 nucleotides upstream from the initiation methionine codon. A canonical promoter "TATA" box is located 30 base pairs upstream of the Cap site. A typical "CCAAT" box sequence is not present in the adjacent 5'-flanking region; however, there is a GC-rich sequence (at nucleotide -215) similar to the binding site for the nuclear transcription factor Sp1. Upstream from the transcriptional initiation site are elements with homology (approximately 75%) to the putative fat-specific transcriptional element FSE2 and core consensus sequences for cAMP and glucocorticoid regulatory elements. A chimeric construct, containing 363 base pairs of 5'-flanking sequence and 30 nucleotides of 5'-untranslated sequence of the mouse SCD gene ligated to the bacterial chloramphenicol acetyltransferase gene, was transfected into 3T3-L1 cells. When cells were induced to differentiate into adipocytes, expression of the SCD chloramphenicol acetyltransferase gene increased approximately 63-fold, suggesting that the SCD promoter region contains elements that mediate the response to adipogenic agents which induce differentiation.
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PMID:Differentiation-induced gene expression in 3T3-L1 preadipocytes. Characterization of a differentially expressed gene encoding stearoyl-CoA desaturase. 290 62

Stage VI Xenopus oocytes were injected with a plasmid (pBB0.6-CAT) which contains the cAMP regulatory element (CRE) from the rat liver phosphoenolpyruvate carboxykinase (PEPCK) gene fused upstream from a reporter gene [chloramphenicol acetyltransferase (CAT)]. Inhibition of the expression of the reporter gene (average = 51%) was observed in the presence of 10 microM progesterone, which is known to lead to inactivation of the oocyte cAMP dependent protein kinase (A kinase). In contrast, oocytes injected with a control plasmid (pSV2CAT), which contains no CRE, exhibited a variable increase (average = 31%) in CAT activity after progesterone treatment. Injection of the purified bovine cardiac A kinase catalytic subunit prior to exposure of oocytes injected with pBB0.6 CAT to progesterone prevents the loss of CAT activity generated by incubation with the steroid. Gel retardation analyses with oocyte lysates and a labeled synthetic oligonucleotide fragment containing the CRE from the PEPCK gene showed the existence of a complex with the same Rf and specificity as that formed with rat liver extracts. Subsequent exposure to progesterone, however, led to a rapid and extensive decrease in this binding activity. Taken together, these results are consistent with but do not prove the hypothesis that progesterone treatment and A kinase inactivation lead to a decrease in pBB0.6 CAT expression by virtue of a decline in the binding activity of an oocyte factor(s) to the CRE of the PEPCK fragment in pBB0.6-CAT, thereby decreasing transcription of the CAT gene.
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PMID:Progesterone decreases DNA binding factor activity and the expression in Xenopus oocytes of a cAMP responsive gene from rat liver. 297 91

Transcription of the vasoactive intestinal polypeptide (VIP) gene is regulated by cAMP. To identify the nucleotide sequences in the human VIP gene responsible for this regulation, we constructed chimeric genes containing different portions of the 5'-flanking region of the human VIP gene fused to the structural sequence encoding the bacterial reporter enzyme chloramphenicol acetyltransferase (CAT). The transcriptional activities of the fusion genes introduced into the rat pheochromocytoma cell line PC12 were assayed by measuring CAT activity in the cell lysates. Forskolin, an adenylate cyclase-activating agent, stimulated the expression of VIP-CAT fusion genes. Deletional analysis demonstrated that a region between -86 and -70 nucleotides upstream from the transcriptional origin of the human VIP gene was responsible for stimulation by forskolin. This region was able to confer cAMP-responsiveness to a gene that is not normally regulated by cAMP. Two copies of a 5 base pair motif, 5'-CGTCA-3', are required for activity of the VIP cAMP regulatory region. This motif is also present in the cAMP regulatory region of several other eukaryotic genes.
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PMID:Identification of a region in the human vasoactive intestinal polypeptide gene responsible for regulation by cyclic AMP. 303 25

The human chorionic gonadotropin-alpha (CG-alpha) gene is transcriptionally activated by cAMP. Sequencing the CG-alpha 5'-flanking region identified two copies of a palindrome, 5'-TGACGTCA-3', homologous to sequences in other cAMP-responsive genes. The two palindromes are contained within two identical 18-base pair (bp) sequences arranged as adjacent direct repeats. One or two synthetic copies of the 18-bp sequences were inserted into plasmids containing either the CG-alpha promoter or the SV40 promoter directing transcription of the chloramphenicol acetyltransferase gene. The 36-bp (double) element markedly enhanced chloramphenicol acetyltransferase activity in placental choriocarcinoma (JEG-3) cells when inserted in either orientation both 5' to the cap site or 3' of the coding sequence, thus defining it as an enhancer. Moreover, 8-br-cAMP stimulated the enhancer activity 30-40-fold. A single 18-bp element also stimulated chloramphenicol acetyltransferase activity, although 5-fold less than the double element, but still imparted a 35-fold transcriptional cAMP responsivity. The enhancer activates its homologous promoter much more efficiently than the SV40 promoter in JEG-3 cells. The alpha-promoter is not nearly as receptive to activation by the enhancer in baby hamster kidney fibroblasts, whereas the more modest enhancement of the SV40 promoter is less cell-specific. These studies suggest that the interaction of a 36-bp enhancer-like element with the homologous promoter represents part of the mechanism of cell-specific expression of the CG-alpha gene and that the enhancer is co-localized with a highly effective cAMP-response element.
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PMID:Cyclic AMP responsiveness of human gonadotropin-alpha gene transcription is directed by a repeated 18-base pair enhancer. Alpha-promoter receptivity to the enhancer confers cell-preferential expression. 304 Jul 32

The hallmark of "beta 2-interferon (IFN-beta 2)/hepatocyte-stimulating factor/interleukin 6" gene expression is its inducibility in different types of human cells (fibroblasts, monocytes, epithelial cells, and endothelial cells) by different stimuli, which include cytokines such as tumor necrosis factor, interleukin 1 (IL-1) and platelet-derived growth factor, different viruses, and bacterial products such as endotoxin. The activation by cytokines, viruses, and second messenger agonists of the IFN-beta 2 promoter linked to the bacterial chloramphenicol acetyltransferase (CAT) gene was studied after transfection into HeLa cells. A chimeric gene containing IFN-beta 2 DNA from -1180 to +13 linked to the CAT gene was inducible approximately 10-fold by phorbol 12-myristate 13-acetate (PMA), followed, in decreasing order, by pseudorabies and Sendai viruses (7- to 11-fold each); serum (6- to 9-fold); the cytokines tumor necrosis factor, IL-1, and epidermal growth factor (3- to 5-fold each); the cAMP agonists BrcAMP and forskolin and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (2- to 6-fold each); poly(I).poly(C) (2- to 4-fold); 1,2-diacylglycerol and the calcium ionophore A23187 (1.5- to 2-fold each). Bacterial endotoxin did not activate this IFN-beta 2/CAT fusion gene in HeLa cells. Deletion of the 5' boundary of the IFN-beta 2 DNA from -1180 to -596 in the fusion gene preserved its activation by IL-1, tumor necrosis factor, epidermal growth factor, serum, pseudorabies, and Sendai viruses and by PMA, Br-cAMP, and forskolin; deletion to -225 led to a small reduction (by a factor of 1.5-2) in the responsiveness to serum, PMA, and Sendai virus but not to the other inducers; a further deletion to -112 greatly reduced all responsiveness. Thus, the region between -225 and -113 in IFN-beta 2, which contains DNA motifs similar to the regulatory elements in the human c-fos gene, appears to contain the major cis-acting regulatory elements responsible for the activation of the IFN-beta 2 promoter by several different cytokines, viruses, and second messenger agonists.
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PMID:Activation of the human "beta 2-interferon/hepatocyte-stimulating factor/interleukin 6" promoter by cytokines, viruses, and second messenger agonists. 304 22


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