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)

One remarkable genetic feature of the class I MHC genes is their unparalleled degree of genetic polymorphism and diversity. The polymorphism is reflected by the fact that multiple loci encode class I molecules, and for each locus there are multiple alleles. In the course of investigating the regulation of HLA-A and HLA-B mRNA in human colorectal carcinoma cell lines, we have noticed a noncoordinate expression of the HLA mRNA in some of these cell lines. This observation prompted us to make use of these cell lines to study the locus-specific transcriptional regulation of HLA genes. Bandshift and footprint assays revealed at least three distinct and independent DNA-binding factors that bind to the core regulatory element of the HLA-A and HLB-B gene locus. A "novel" DNA-binding factor recognizing the CCAAT motif seems to be important for locus-specific expression of HLA-A mRNA, whereas a different factor which binds to a Sp1-like sequence is crucial for normal HLA-B mRNA expression. In certain colorectal cancer cell lines, underrepresentation of these locus-specific DNA-binding proteins correlates with the locus-specific down-regulation of HLA mRNA. This observation is further supported by experiments which demonstrated that the locus-specific suppression of exogenously introduced TK-chloramphenicol acetyltransferase DNA constructs, containing the "putative" HLA locus-specific DNA core regulatory sequence, is regulated in a locus-specific manner when introduced into these HLA-A- and HLA-B-deficient human colorectal cell lines.
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PMID:Locus-specific transcriptional control of HLA genes. 151 66

A primary site of infection by human adenoviruses is lymphoid cells. However, analysis of the viral control elements and the cellular factors that regulate adenoviral gene expression in lymphocytes has not been reported. The adenovirus early region 3 (ES) gene products are involved in the maintenance of viral persistence by complexing with the class I MHC antigens, thus preventing their cell surface expression with a resultant decrease in host immunologic destruction. To determine whether different cellular factors were involved in E3 regulation in lymphocytes as compared with HeLa cells, both DNA binding and transfection analysis with the E3 promoter in both cell types were performed. These studies detected two novel domains referred to as L1 and L2 with a variety of lymphoid but not HeLa extracts. Each of these domains possessed strong homology to motifs previously found to bind the cellular factor NF-kappa B. Transfections of E3 constructs linked to the chloramphenicol acetyltransferase gene revealed that mutagenesis of the distal NF-kappa B motif (L2) had minimal effects on promoter expression in HeLa cells, but resulted in dramatic decreases in expression by lymphoid cells. In contrast, mutagenesis of proximal NF-kappa B motif (L1) had minimal effects on gene expression in both HeLa cells and lymphoid cells but resulted in a small, but reproducible, increase in gene expression in lymphoid cells when coupled to the L2 mutation. Reversing the position and subsequent mutagenesis of the L1 and L2 domains indicated that the primary sequence of these motifs rather than their position in the E3 promoter was critical for regulating gene expression.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Lymphoid specific gene expression of the adenovirus early region 3 promoter is mediated by NF-kappa B binding motifs. 214 90

The rat alpha-myosin heavy-chain (alpha-MHC) gene is regulated by 3,5,3'-triiodo-L-thyronine (T3) in ventricular myocardium and is constitutively expressed in atrial tissue. Less is known about regulation of the human gene, but conservation of sequences in the 5'-flanking region between the rat and human alpha-MHC genes suggests that the human gene may be regulated similarly. Accordingly, T3-responsiveness and tissue-specific expression of human and rat alpha-MHC/chloramphenicol acetyltransferase fusion constructs have been compared in rat fetal heart cells, L6E9 myoblasts and myotubes, 3T3 fibroblasts, and HeLa cells. Transient transfection assays revealed a complex series of cis-regulatory elements in the 5'-flanking sequences in the human genes, including a basal promoter element with canonical TATAA and CAAT sequences, two positive regulatory element(s), and two negative regulatory elements, which markedly diminished both constitutive and T3-inducible activity. Interestingly, the human gene seemed to contain a proximal thyroid-hormone response element(s) not found in the rat gene. In L6E9 myoblasts and myotubes, the human constructs were constitutively expressed but not T3-regulated; none of the constructs were active in 3T3 or HeLa cells. We propose that interactions among the thyroid hormone responsive elements and other cis-acting elements in the human alpha-MHC 5'-flanking sequences may be sufficient to explain the characteristic features of expression of this gene in cardiac tissues.
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PMID:Thyroid hormone regulates expression of a transfected human alpha-myosin heavy-chain fusion gene in fetal rat heart cells. 229 92

In ventricular muscle, 3,5,3'-triiodo-L-thyronine (T3) stimulates the expression of the alpha-myosin heavy-chain (alpha-MHC) gene. To test for gene elements required for induction, a fragment of the alpha-MHC gene containing 2.9 kilobases of 5' flanking sequences and 420 base pairs of DNA 3' to the transcription initiation site was linked to the coding sequences of the bacterial chloramphenicol acetyltransferase (CAT) gene. The alpha-MHC fusion gene was introduced into primary cultures of fetal rat heart myocytes. Induction of the transfected gene was monitored by assaying CAT activity while endogenous alpha-MHC mRNA expression was measured by using a synthetic oligonucleotide probe complementary to sequences in the 3' untranslated region of the mRNA. Without T3, CAT activity was only slightly greater than background. When T3 at a final concentration of 10 nM was added to the cultures, CAT activity was increased 8-fold by 48 hr. The response time and doses of T3 required for induction of CAT activity and alpha-MHC mRNA in transfected cells were similar, suggesting that the synthetic and endogenous genes may have a common mechanism of control. When simian virus 40 enhancer and early promoter sequences were included in the construct, CAT activity was constitutively expressed, but it could be increased 7-fold by the addition of T3. Several deletions were introduced into the 5' flanking sequences of the alpha-MHC fragment and the effects on induction of CAT activity were examined. Progressive deletions of 5' sequences from positions -947 to -374 reduced but did not eliminate induction of CAT activity, suggesting that more than one region may be required for optimal induction by thyroid hormone. The results indicate that DNA sequences required for efficient induction by T3 are present in the 5' flanking sequences of the alpha-MHC gene.
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PMID:Thyroid hormone regulates expression of a transfected alpha-myosin heavy-chain fusion gene in fetal heart cells. 347 99

Cellular adherence is important for monocyte migration and function and is known to induce monocyte activation, leading to the production of mRNA for several proto-oncogenes and cytokines. In addition, since cellular adherence has important intracellular signalling function, it has the potential to enhance human immunodeficiency virus (HIV) replication in monocytic cells. We have investigated the effects of adhesion of the monocytic cell line THP-1 transfected with HIV1 or HIV2 long terminal repeat chloramphenicol acetyltransferase (LTR CAT) constructs. These studies have shown that adherence to tissue culture plastic or confluent endothelial cells is essential for enhanced HIV LTR CAT expression in lipopolysaccharide-stimulated cells. In addition, we have investigated the effects of engagement of specific adhesion molecules, using immobilized antibodies, on HIV replication in the promonocytic cell line OM101, which contains a single latent proviral copy of HIV. Such studies have demonstrated that engagement of CD18, the beta subunit of the lymphocyte function-related antigen-1 (LFA-1) and major histocompatibility complex class II (MHC II) enhanced HIV replication. LFA-1 is involved in both monocyte-endothelial cell interactions and monocyte-T-cell interactions, and MHC II is involved in monocyte interaction with antigen-specific T cells. These data suggest that such interactions of membrane adhesion molecules with their appropriate ligand enhance HIV replication in vivo. Thus, this study has demonstrated that cellular adherence is a key regulatory factor of HIV replication in monocytic cells.
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PMID:Cellular adherence enhances HIV replication in monocytic cells. 780 Sep 38

The molecular mechanism(s) by which three cytokines (IFN-gamma, TNF-alpha, TGF-beta) affect class II MHC gene expression in primary rat microglia was examined. IFN-gamma is a potent inducer of the class II gene, and this induction is unaffected by treatment with either TNF-alpha or TGF-beta. Transient transfection of primary rat microglia with an HLA-DRA promoter linked to the chloramphenicol acetyltransferase reporter gene (DRA-CAT) demonstrated that IFN-gamma acts at the transcriptional level to induce class II MHC gene expression, and that TNF-alpha and TGF-beta have no influence on IFN-gamma-induced promoter activity. Experiments using a series of DRA substitution mutants that individually affect the W, X1, X2, or Y elements, as well as a double mutation in both X1 and X2, indicate that all four of these elements are required for responsiveness of the DRA promoter to IFN-gamma. The effect of IFN-gamma and TNF-alpha on DNA binding proteins by microglia was examined. A constitutive complex with specificity for the X2 box was detected in extracts from unstimulated microglia. IFN-gamma treatment changed this complex to migrate with slower mobility, and TNF-alpha had no effect on either the constitutive or IFN-gamma-induced complexes. These studies provide information on the molecular regulation of the class II MHC gene in microglia, a cell type critically involved in immune regulation within the central nervous system.
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PMID:Class II MHC gene expression in microglia. Regulation by the cytokines IFN-gamma, TNF-alpha, and TGF-beta. 787 54

The hepatitis delta virus (HDV) genome consists of circular ssRNA which has extensive intramolecular complementarity and can form a dsRNA rod-like structure. If such RNA species were to exist in an unmasked form in cells, they would be expected to induce interferon (IFN) expression and activate two IFN-inducible dsRNA-dependent enzymes with anti-viral activity, namely the dsRNA-dependent protein kinase (PKR) and 2',5' oligoadenylate (2',5' A) synthetase. Since the virus replicates to high copy number for prolonged periods in infected cells it is apparently able to evade these antiviral mechanisms. The RNA genome may be masked and fail to induce or activate the antiviral response, or the virus may inhibit such a response. Treatment of a hepatoma cell line, Huh7, and a fibrosarcoma cell line, HT1080, stably transfected with a trimeric HDV cDNA construct, with IFN-alpha or IFN-gamma for up to seven days failed to influence the level of expression of genomic or antigenomic HDV RNA, or delta antigen (Ag). This is consistent with either failure of activation or inhibition of the IFN response. However the induction of several IFN-responsive genes, including PKR, 2',5' A synthetase and class I MHC is normal and cotransfection of a construct expressing delta Ag did not affect expression from an IFN-inducible chloramphenicol acetyltransferase construct. In addition, the activation of PKR is not inhibited in HDV-expressing cells and antiviral assays suggest that the ability of these cells to mount an antiviral response to at least two cytopathic viruses is unaffected. IFN-beta is inducible normally by dsRNA in cells transfected with the delta cDNA trimer. We conclude that HDV replication is not inhibited by IFN-alpha or IFN-gamma, even though the responses of cells expressing HDV RNA and antigen to IFN and dsRNA are intact.
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PMID:Hepatitis delta virus replication in vitro is not affected by interferon-alpha or -gamma despite intact cellular responses to interferon and dsRNA. 791 7

Basal transcription of the human multidrug resistance (mdr1) promoter was studied by chloramphenicol acetyltransferase (CAT) reporter fusion gene analysis in two parental and doxorubicin-resistant human tumor cell lines. Deletion of mdr1 DNA sequences to -89 relative to the start of transcription (at +1) had little effect on expression. Deletion of nucleotide sequences from -89 to -70, however, resulted in a 5-10-fold reduction in mdrCAT expression. DNase I footprint analysis demonstrated that the region from -85 to -70 was protected from nuclease digestion using nuclear extracts from these cell lines. The sequence between -82 and -73 is perfectly homologous with the 10-base pair Y-box consensus sequence found in the promoters of all major histocompatibility complex class-II (MHC II) genes. The Y-box sequence in MHC II genes is required for accurate and efficient transcription and contains the sequence CCAAT in the reverse orientation (Dorn, A., Durand, B., Marfing, C., Le Meur, M., Benoist, C., and Mathis, D. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 6249-6253). Mutations in the reverse CCAAT sequence of the Y-box consensus substantially reduced expression of an mdrCAT vector and eliminated nucleoprotein binding in an electrophoretic mobility shift assay. These results suggest that proteins which bind to the putative Y-box consensus sequence are critical for basal transcriptional regulation of the human mdr1 gene.
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PMID:A Y-box consensus sequence is required for basal expression of the human multidrug resistance (mdr1) gene. 809 99

We have investigated the regulated expression of genes injected into the heart of large mammals in situ. Reporter constructs using the chloramphenicol acetyltransferase gene under the control of muscle-specific beta-myosin heavy chain (beta-MHC) or promiscuous (mouse sarcoma virus) promoters were injected into the canine myocardium. There was a linear dose-response relation between the level of gene expression and the quantity of plasmid DNA injected between 10 and 200 micrograms per injection site. The level of reporter gene expression did not correlate with the amount of injury imposed on the cardiac tissue. There was no regional variation in expression of injected reporter genes throughout the left ventricular wall. By use of both the mouse sarcoma virus and a muscle-specific beta-MHC promoter, reporter gene expression was one to two orders of magnitude greater in the heart than in skeletal muscle. Expression in the left ventricle was threefold higher than in the right ventricle. Chloramphenicol acetyltransferase activity was detected at 3, 7, 14, and 21 days after injection, with maximal expression at 7 days after injection. Statistical analysis of coinjection experiments revealed that coinjection of a second gene construct (Rous sarcoma virus-luciferase) is useful in the control of transfection efficiency in vivo. Furthermore, using reporter constructs containing serial deletions of the 5' flanking region of the beta-MHC gene, we performed a series of experiments that demonstrate the utility of this model in mapping promoter regions and identifying important regulatory gene sequences in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Gene injection into canine myocardium as a useful model for studying gene expression in the heart of large mammals. 843 91

Class II genes of the MHC show a striking homology upstream of the transcription start site that is composed of three conserved sequences (S, X and Y boxes, each separated by 15-20 bp). The presence of the S-box sequence in the mouse MHC class II gene I-A Beta was examined for its influence on the expression of this gene. Deletion or mutation of the S box decreased the induction of chloramphenicol acetyltransferase (CAT) activity in B lymphocytes by 32%. In macrophages, deletion or mutation of the S box abolished interferon-gamma (IFN-gamma) inducibility of CAT activity. Using a gel-retardation assay, we have identified a nuclear factor whose binding site overlaps the 7-mer conserved sequence of the S box. This factor is present in lymphocytes, macrophages, mastocytes and fibroblasts. Surprisingly, binding of this nuclear factor to DNA was induced by IFN-gamma in bone-marrow-derived macrophages, but not in macrophage-like cell lines. The binding site for this factor was defined by DNase I footprinting and partially purified by using an affinity column containing double-stranded oligonucleotides containing a sequence of the S box. A prominent protein of 43 kDa was found that bound specifically to the S-box sequence.
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PMID:Identification of a transcription factor that binds to the S box of the I-A beta gene of the major histocompatibility complex. 861 Nov 49


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