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)

Human papillomavirus type 8 (HPV-8) is one aetiological agent of macular and flat wart-like lesions in patients with epidermodysplasia verruciformis and appears to be closely linked to skin carcinogenesis. A 1.2 kb region of the genome, which was previously shown to contain a viral E2-dependent enhancer, was progressively shortened from both ends with Bal 31. The resulting fragments were tested for their ability to stimulate chloramphenicol acetyltransferase (CAT) expression from the simian virus 40 (SV 40) promoter. This analysis showed a complex interaction between cis-active, positive and negative control elements located throughout the non-coding region and the flanking reading frames. Two separate positively acting sequences significantly stimulated expression only in cooperation with a third region, which led to 12-fold, E2-dependent enhancement on its own. A major negative element was not only active in the context of HPV-8 sequences, but also down-regulated SV40 enhancer-promoter-driven CAT expression when cloned downstream of the transcription unit. It acted at the transcriptional levels as shown by RNase protection assays and can therefore be regarded as a cis-acting silencer of transcription.
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PMID:Human papillomavirus type 8 contains cis-active positive and negative transcriptional control sequences. 217 59

The 5'-region of the rat alpha 2-macroglobulin gene has been characterized. A 5.6 kb Sal I - Xba I fragment containing the first 4 exons of the alpha 2-macroglobulin gene and 1.3 kb of its 5'-flanking region was sequenced. The putative transcriptional start site was determined by RNase protection and primer extension analysis. TATA- and CAAT-box equivalent sequences were found. A potential glucocorticoid receptor binding site was located on the antisense strand. DNA sequences containing the 5'-flanking region of the rat alpha 2-macroglobulin gene were linked to the gene coding for the bacterial chloramphenicol acetyltransferase and introduced into Hep G2 cells. In these transfected Hep G2 cells CAT activity could be induced by recombinant human interleukin-6. Deletion analyses have shown that the sequences between -852 and -777 as well as between -404 and -165 relative to the cap site, contain regulatory elements involved in the interleukin-6 dependent induction of the alpha 2-macroglobulin gene.
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PMID:Identification of the promoter sequences involved in the interleukin-6 dependent expression of the rat alpha 2-macroglobulin gene. 246 33

Enhancer-like sequences have previously been identified in the promoter region of the mouse major histocompatibility complex (MHC) class I genes. We have screened for such sequences in and around a human MHC class I gene, HLA-B7. Various restriction fragments of the B7 gene were assayed for their ability to enhance transcription of a bacterial chloramphenicol acetyltransferase gene from a simian virus 40 promoter in transiently transfected mouse LTA cells. Our results demonstrate that enhancer activity is located in introns 3 and 5 as well as 5' to the transcription initiation site. RNase protection experiments corroborate the results. Preliminary experiments indicate that B7 enhancers are active in various cell types. The role of these enhancers in B7 gene expression is not known at present. We speculate that the position of the enhancer elements may be related to the occurrence of Hpa II tiny fragment islands.
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PMID:Multiple enhancer-like sequences in the HLA-B7 gene. 250 82

The human papillomavirus type 11 enhancer, when linked to the minimal simian virus 40 early promoter, has been dissected into two domains in monkey kidney CV-1 cells, one being constitutive (designated CEI) and the other inducible by trans-acting E2 proteins encoded by homologous and heterologous papillomaviruses (H. Hirochika, T.R. Broker, and L.T. Chow, J. Virol. 61:2599-2606, 1987; H. Hirochika, R. Hirochika, T.R. Broker, and L.T. Chow, Genes Dev. 2:54-67, 1988). We have demonstrated that the natural promoter regulated by this enhancer is located immediately upstream of the E6 open reading frame (the E6 promoter). We have mapped the cap site to nucleotide 99 by RNase protection. We further demonstrate a second constitutive enhancer element, CEII, which is required for transcription from the E6 promoter in the human cervical carcinoma cell lines C-33A and HeLa but not in CV-1 cells. By deletion mapping, we have localized this cell type-specific domain to 71 base pairs by using chloramphenicol acetyltransferase assays. Deletion of either CEI or CEII dramatically decreased the constitutive activity of the enhancer and the E6 promoter, whereas multimerization of either domain in the absence of the other could independently restore expression. Furthermore, when either of these elements was deleted, the full-length E2 protein of human papillomavirus type 11 abolished the remaining basal E6 promoter activity, demonstrating for the first time that the enhancer-activating E2 protein of human papillomaviruses can also function as a transcriptional repressor for the homologous E6 viral promoter. The presence of multiple copies of each element in tandem overcomes the repression by the E2 protein. The effects of CEII are at the level of transcription, without changing the cap site. By gel shift assay, we have shown that a protein present in nuclear extracts of C-33A and HeLa cervical carcinoma cells binds to the newly identified constitutive element II. This protein did not bind the simian virus 40 enhancer, nor did it bind to the enhancer region of many other papillomaviruses tested. UV cross-linking experiments revealed major 44-kilodalton and minor 34-kilodalton proteins that bound specifically to CEII. These two proteins are either related or bind to CEII with high cooperativity. We conclude that transcriptional activities directed by the enhancer and E6 promoter reflect an intricate balance among viral and cellular factors. We present a model on the regulation of the E6 promoter by host and viral transcription factors.
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PMID:Identification of a novel constitutive enhancer element and an associated binding protein: implications for human papillomavirus type 11 enhancer regulation. 254 7

We have examined the transcriptional regulation of the rabbit myosin heavy chain (HC) beta gene by using DNA-mediated transfection experiments. To analyze the activity of the myosin HC beta promoter in a myogenic background, cultured myoblasts from 12-day-old chick embryonic breast muscle were transfected with a chimeric gene containing 781 base pairs of the promoter region fused to the gene for chloramphenicol acetyltransferase (CAT). As indicated by the transient expression of chloramphenicol acetyltransferase, the activity of the promoter in myoblast cultures increased at least 32-fold following differentiation and was selectively inhibited when myogenesis was blocked with 5-bromodeoxyuridine. Furthermore, RNase protection experiments showed that the in vivo myosin HC beta transcriptional initiation (or cap) site was utilized in the transfected skeletal muscle cells and also that the regulation of the exogenous promoter was similar to the induction of the endogenous skeletal alpha-actin gene. The results indicated that the exogenous promoter is regulated in a tissue- and stage-specific manner. By creating progressive 5' deletions of the promoter, we showed that only the region extending -294 base pairs upstream from the cap site is necessary for the muscle-specific expression. Linker-scanner mutagenesis of this region indicated that the positive regulation in differentiated skeletal muscle is mediated by at least two distinct elements within the 5'-flanking region of the myosin HC beta gene.
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PMID:Muscle-specific regulation of a transfected rabbit myosin heavy chain beta gene promoter. 256 93

Previous studies have shown that transferred hybrid constructs containing the PRL promoter are expressed specifically in rat pituitary (GH) cell lines. However, it is not yet clear which DNA region(s) is primarily responsible for expression directed by this promoter in pituitary cells. In the present studies we have examined the DNA sequences required for cell type-specific transcription of the rat PRL (rPRL) promoter either during transient expression in intact cells or in nuclear chromatin extracts. RNase protection and nuclear run-on transcription assays showed directly that a PRL-chloramphenicol acetyltransferase (CAT) construct containing about two kilobase-pairs of the rPRL promoter region (pPRL-CAT) is transcribed specifically in pituitary (GH3) cells. Analysis by transient expression in GH3 cells of pPRL-CAT and its 5' deletions showed that 1) deletion of sequences between positions -1957 and -958 did not significantly affect CAT activity; 2) the first 187 basepairs (bp) of the rPRL promoter directs full CAT activity; and 3) 98% of this activity is accounted for by rPRL DNA sequences between positions -187 and -113, containing two GH3 chromatin footprinting sites. Analysis in GH3 cell nuclear extracts showed that transcription of PRL-CAT constructs is unaffected by successive 5' deletions from position -1957 to -187, and that further deletion to -75 yielded only a moderate (approximately 2-fold) decrease in transcription.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Proximal rat prolactin promoter sequences direct optimal, pituitary cell-specific transcription. 274 60

The upstream regulatory region of the human papilloma virus-16 (HPV-16) genomic DNA contains a sequence element with a large degree of homology to the partially palindromic sequence GGTACANNNTGTTCT, which is the consensus sequence of the glucocorticoid responsive elements of known genes regulated by this steroid hormone. DNase I and dimethylsulfate protection experiments reveal the binding of this sequence by rat glucocorticoid receptor protein. A 400-bp DNA segment centrally containing this sequence confers strong inducibility by dexamethasone to the promoter p97 of HPV-16 and to the Herpes simplex virus thymidine kinase promoter, as judged by chloramphenicol acetyltransferase activity and RNase protection assays. The same DNA segment, that does not contain the consensus sequences of all papilloma viruses relevant for E2 protein-mediated transcription enhancement, functions in an enhancer-like fashion in addition to its glucocorticoid responsive action. This hormone-independent transcription enhancement is absent in human MCF7 cells, but is strong in human HeLa cells where the combined activity of the constitutive and the steroid hormone-dependent enhancer elements stimulate transcription by a factor of 500. This cell type specificity of the HPV-16 enhancer may be responsible for the tissue tropism of the virus. These observations and the presence of numerous homologies to known enhancers of cellular and viral genes suggest a complex pattern of activation of the human papilloma virus-16 promoters.
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PMID:The upstream regulatory region of the human papilloma virus-16 contains an E2 protein-independent enhancer which is specific for cervical carcinoma cells and regulated by glucocorticoid hormones. 282 35

Insulin has been shown to inhibit rat growth hormone (GH) gene transcription. The effects of insulin were therefore tested on the expression of a transfected human GH gene. A 2.6-kilobase EcoRI fragment of the human GH gene was propagated in pUC18 and transfected by calcium-phosphate shock into HeLa and GC cells, respectively. Transfected cells grown in serum-free medium for 72 h expressed human GH measured by specific radioimmunoassay, incorporation of [35S] methionine into newly synthesized GH, and the presence of the appropriately sized protected transcripts seen after RNase protection assay. Immunoprecipitation analysis showed that insulin (0.7-7 nM) suppressed both the basal as well as the hydrocortisone (100 nM)-stimulated expression of newly synthesized 22-kDa GH in a dose-dependent fashion. Insulin (7 nM) also suppressed the basal and hydrocortisone-stimulated GH mRNA transcripts in these cells. Control nontransfected cells did not express human GH. Cells transfected with the truncated pOGH gene and pTKGH gene failed to respond to insulin treatment, whereas the human GH promoter was able to confer insulin responsiveness to the chloramphenicol acetyltransferase reporter gene. cis-Acting regulatory sequences residing on the 497-base pair 5'-flanking region of the human GH gene therefore appear to be a requirement for human GH gene response to the insulin signal.
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PMID:Insulin regulates expression of the human growth hormone gene in transfected cells. 290 52

The genome of human immunodeficiency virus encodes a protein that dramatically elevates amounts of viral proteins. The precise mechanism of this trans-activation remains to be established. It has been reported that trans-activation can occur without major changes in the levels of mRNA. We constructed recombinant plasmids containing those viral sequences required in cis for trans-activation linked to the chloramphenicol acetyltransferase gene. These plasmids were introduced into cultured cells in either the presence or absence of a second plasmid that directed expression of the viral trans-activator protein. Expression of the chloramphenicol acetyltransferase gene was measured at the level of protein (by enzymatic assay) and RNA (by ribonuclease protection and primer extension). Our results demonstrate that trans-activation is accompanied by large increases in mRNA levels; these increases may be sufficient to explain the elevated levels of trans-activated protein.
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PMID:Elevated levels of mRNA can account for the trans-activation of human immunodeficiency virus. 302 48

Two overlapping cosmids have been isolated containing the entire murine gene for SPARC (osteonectin), a Ca2+-binding, phosphorylated glycoprotein associated with extracellular matrix synthesis and remodeling. The gene contains 10 exons and covers 26.5 kilobase pairs of DNA. Exon analysis shows that the two N-terminal glutamic acid-rich sequences which are predicted to undergo conformational change upon binding of calcium, as well as the C-terminal EF-hand Ca2+-binding domain are each encoded by a single exon. Comparative analysis of the exon sequence does not support the idea that the SPARC gene has evolved by shuffling of exons from other Ca2+-binding proteins. The 5' flanking region of the SPARC gene, which promotes transcription when placed in front of the bacterial chloramphenicol acetyltransferase gene, contains neither "TATA" nor "CAAT" box sequences. However, unlike most other genes lacking these motifs, mapping of the 5' end of the SPARC gene by RNase protection and primer extension analysis reveals only a single major and one minor transcription start site. The upstream region to -120 includes six repeats of the sequence GGAGG, two repeats of the sequence 5' GGAGG A/C GGAGGG 3', and a potential transcription factor AP-2 binding site.
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PMID:Characterization of the mouse SPARC/osteonectin gene. Intron/exon organization and an unusual promoter region. 316 75


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