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)

The transient expression of hepatitis B virus (HBV) surface and "eJ" antigens caused by transfection of human hepatoblastoma HepG2 cells with HBV DNA was markedly inhibited by cotransfection with poly(I):poly(C). Cotransfection with poly(I):poly(C) also inhibited the expression of bacterial chloramphenicol acetyltransferase (CAT) gene which was under the control of either the HBV core promoter or the human immunodeficiency virus (HIV-1) long terminal repeat. This inhibition was much more pronounced on the expression of HBV-promoted CAT than HIV-promoted CAT. The uptake of reporter plasmid was not affected by cotransfected poly(I):poly(C). The inhibition was found to be at the steady-state CAT mRNA level and appeared to be specific for HBV and HIV regulatory sequences since CAT expression directed by other viral and cellular regulatory sequences was not inhibited. Cotransfection with a mixture of equal amounts of poly(I) and poly(C) had similar inhibitory effects whereas cotransfection with poly(l) or poly(C) alone, or other double-stranded ribo- or deoxyribonucleotides, did not have such strong effects. The addition of poly(l):poly(C) to the culture medium of cells transfected with these reporter plasmids caused little inhibition. Transfection with poly(l):poly(C) induced a minimal amount of intracellular interferon-alpha in HepG2 cells which may be involved in selective inhibition of HBV-and HIV-1-directed gene expression. 2-Aminopurine, an inhibitor of double-stranded RNA activated protein kinase known to block interferon gene induction by poly(l):poly(C), partially reversed the poly(l):poly(C)-induced inhibitory effect on HBV-CAT expression.
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PMID:Selective inhibition of hepatitis B virus and human immunodeficiency virus sequence-promoted gene expression by cotransfected poly(I):poly(C). 221 31

12-O-Tetradecanoylphorbol-13-acetate (TPA) activated the c-fos gene enhancer linked to the chloramphenicol acetyltransferase or luciferase reporter gene in the wild type PC-12 cells but not in the variant PC-12 cells that originated from the wild type cells. Transfection of the c-Ha-rasval12 complementary DNA (cDNA) or addition of dibutyryl cAMP to the wild type PC-12 cells as well as to the variant PC-12 cells activated the c-fos gene enhancer. Prolonged treatment of the wild type PC-12 cells with phorbol-12,13-dibutyrate caused down-regulation of protein kinase C. In these cells, TPA did not stimulate the c-fos gene enhancer any more, but transfection of the c-Ha-rasval12 cDNA still stimulated the c-fos gene enhancer to the same extent as induced in the control cells. Transfection of the c-Ha-rasval12 cDNA or addition of TPA to the wild type PC-12 cells stimulated the serum-response element but not the cAMP-response element. Dibutyryl cAMP stimulated both the serum-response element and the cAMP-response element in the wild type PC-12 cells. These results indicate that the c-Ha-rasval12 protein activates the serum-response element, but not the cAMP-response element in the c-fos gene enhancer, and that the signal pathway from the c-Ha-rasval12 protein to the c-fos serum-response element is independent of protein kinase C and cAMP-dependent protein kinase.
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PMID:Activation of the c-fos serum-response element by the activated c-Ha-ras protein in a manner independent of protein kinase C and cAMP-dependent protein kinase. 240 11

Interleukin-6 (IL-6) is a major systemic alarm signal that indicates the occurrence of tissue damage. The IL-6 gene is induced in various cell types by serum, inflammation-associated cytokines, viruses, and second-messenger agonists. There is an overall functional similarity between IL-6 and c-fos promoters, since transfection of excess amounts of either promoter DNA into intact HeLa cells modulates the function of the heterologous promoter construct. Furthermore, the transcription regulatory factor Fos transrepresses both the IL-6 and c-fos promoters. The 115-base pair (bp) region from -225 to -111 in the IL-6 5'-flanking region, which shares nucleotide sequence similarity with the c-fos serum response (SRE) and adjacent AP-1-like (the CGTCA motif) elements, confers responsiveness to several reagents, including serum, forskolin, and phorbol ester, upon the heterologous herpesvirus thymidine kinase (TK) promoter. In gel shift assays using nuclear extracts from HeLa cells, the 115-bp IL-6 enhancer formed several complexes that (i) were increased when extracts from induced HeLa cells were used and (ii) were inhibited most efficiently by the fos E DNA fragment (-700 to -100) and by c-fos oligonucleotides containing an intact AP-1-like site (the CGTCA motif). The 23-bp oligonucleotide designated AR1 from within the IL-6 enhancer region (-173 to -151) contains a CGTCA motif and bound nuclear proteins that also associated with c-fos oligonucleotides containing either an intact SRE or AP-1-like site. A single copy of AR1 inserted upstream of the herpesvirus TK promoter rendered this heterologous promoter inducible by IL-1 alpha, tumor necrosis factor, and serum as well as by activators of the protein kinase A (forskolin) and protein kinase C (phorbol ester) signal transduction pathways. Mutations in the AP-1-like site within AR1 (CGTCA----GTTCA) decreased inducibility of the chimeric IL-6/TK/chloramphenicol acetyltransferase gene by phorbol ester and by forskolin but not by serum, IL-1 alpha, or tumor necrosis factor. These data not only show that the AR1 segment from within the IL-6 enhancer binds nuclear proteins that also bind to c-fos regulatory elements but also demonstrate that a single copy of this 23-bp element is functionally sufficient to confer responsiveness to a variety of inducers and thus define a multiple-response element.
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PMID:A multiple cytokine- and second messenger-responsive element in the enhancer of the human interleukin-6 gene: similarities with c-fos gene regulation. 251 37

The possible role of the catalytic subunit of the cAMP-dependent protein kinase in mediating the regulation of prolactin gene transcription has been investigated through the use of a synthetic gene encoding the heat-stable inhibitor of the cAMP-dependent protein kinase. To assess the effects of protein kinase inhibitor expression on cAMP induction of prolactin gene transcription, a marker gene containing the rat prolactin promoter and adjacent 5'-flanking sequences linked to the bacterial chloramphenicol acetyltransferase gene was cotransfected with a protein kinase inhibitor-expression vector. The results demonstrate that the protein kinase inhibitor-expression vector reduced both basal and cAMP-stimulated expression of the cotransfected prolactin-chloramphenicol acetyltransferase gene. A mutant protein kinase inhibitor-expression vector, coding for an inactive inhibitor protein, did not inhibit basal or cAMP-stimulated prolactin gene transcription. Furthermore, the protein kinase inhibitor-expression vector did not inhibit zinc induction of the metallothionein promoter. Analysis of protein kinase activity in transfected cells demonstrated that the protein kinase inhibitor expression vector reduced cAMP-dependent protein kinase activity but did not reduce protein kinase C activity. Nuclease protection experiments confirmed that the effects of the inhibitor vector involved changes in correctly initiated transcripts produced from the prolactin promoter. Surprisingly, the protein kinase inhibitor-expression vector reduced the effects of several different agents including epidermal growth factor, thyrotropin-releasing hormone, phorbol esters, and estrogen on prolactin gene expression to the same extent as it altered cAMP effects.
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PMID:A protein kinase inhibitor gene reduces both basal and multihormone-stimulated prolactin gene transcription. 253 42

Transfection of the cDNA encoding the activated c-raf-1 protein or addition of 12-O-tetradecanoylphorbol-13-acetate (TPA) or dibutyryl cAMP to NIH/3T3 cells activated the c-fos gene enhancer linked to the chloramphenicol acetyltransferase or luciferase reporter gene. Prolonged treatment of NIH/3T3 cells with phorbol 12,13-dibutyrate caused down-regulation of protein kinase C. In these cells, addition of TPA did not stimulate the c-fos gene enhancer any more, but transfection of the c-raf-1 cDNA or addition of dibutyryl cAMP still stimulated the c-fos gene enhancer to the same extent as those induced in the control cells. Transfection of the c-raf-1 cDNA or addition of TPA to NIH/3T3 cells stimulated the serum response element and TPA response element but not the cAMP response element. In contrast, addition of dibutyryl cAMP to NIH/3T3 cells stimulated the cAMP response element but not the serum response element or TPA response element. These results indicate that the activated c-raf-1 protein stimulates the serum response element and TPA response element in a manner independent of protein kinase C and cAMP-dependent protein kinase. Since the c-fos gene enhancer has been shown to contain the serum response element and cAMP response element, it is most likely that the c-raf-1 protein is involved in the regulation of c-fos gene expression through the serum response element.
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PMID:Activation of the serum response element and 12-O-tetradecanoylphorbol-13-acetate response element by the activated c-raf-1 protein in a manner independent of protein kinase C. 255 85

We have constructed the expression plasmids harboring protein kinase C (PKC) mutant cDNAs with a series of deletions in the PKC coding region. These plasmids were transfected into COS7 cells to characterize the PKC mutants. Immunoblot analysis using the anti-PKC antibody identified proteins with the Mr values expected from the PKC mutant cDNAs in the extracts from COS7 cells. The wild-type PKC, when expressed in COS7 cells, conferred increased phorbol ester binding activity on intact cells; but the PKC mutants with the deletion around the C1 region did not show this activity. The wild-type PKC showed protein kinase activity dependent on phospholipid, Ca2+, and phorbol ester, whereas these PKC mutants exhibited protein kinase activity independent of the activators in a cell-free system. A PKC mutant cDNA with the deletion in the C2 region gave increased phorbol ester binding activity. Protein kinase activity of this mutant was much less dependent on Ca2+ compared with the wild-type PKC. A PKC mutant cDNA with the deletion in the C3 region conferred increased phorbol ester binding activity, but neither activator-dependent nor -independent protein kinase activity. These results indicate that elimination of the C1 region of PKC gives rise to constitutively active PKC independent of phospholipid, Ca2+, and phorbol ester and that the C1-C3 regions play distinct roles in the regulatory and catalytic function of PKC. In another series of experiments, transfection of some PKC mutant cDNAs with the deletions around the C1 region into Chinese hamster ovary and Jurkat cells activated the activator protein-1-binding element or the c-fos gene enhancer linked to the chloramphenicol acetyltransferase reporter gene in the absence of phorbol ester. Microinjection of these constructs into Xenopus oocytes induced initiation of germinal vesicle breakdown, indicating that they stimulated the PKC pathway in vivo. Thus, the phorbol ester-independent PKC mutant cDNAs could be a powerful tool to investigate the transmembrane signaling pathway mediated by PKC.
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PMID:Molecular genetic analysis of the regulatory and catalytic domains of protein kinase C. 276 32

The promoter region of mouse lactate dehydrogenase-A gene was fused with the chloramphenicol acetyltransferase gene of Escherichia coli, and the expression of this fusion gene was induced by cyclic AMP in Chinese hamster ovary wild-type cells, but not in mutants affecting the regulatory or catalytic subunit of cAMP-dependent protein kinase.
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PMID:Cyclic AMP-induced expression of the mouse lactate dehydrogenase-A promoter-cat fusion gene in Chinese hamster ovary wild-type cells, but not in cAMP-dependent protein kinase mutant cells. 282 Apr 3

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

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

We have investigated the ability of a constitutively active Gq-alpha mutant, Q209L-alpha q, to regulate target gene expression. Transient expression in GH3 pituitary cells of a rat proximal prolactin promoter-chloramphenicol acetyltransferase construct (-187)PRL-CAT, was stimulated by co-expression of Q209L alpha q, but not by wild-type alpha q. Q209L-alpha q stimulated expression of constructs driven by promoters for either rat prolactin or growth hormone, but not of a control construct driven by the thymidine kinase promoter. Thus, transcriptional effects of alpha q are specific both for the activated state of this G-alpha subunit and the promoter examined. Since both the prolactin and growth hormone promoters are activated by the pituitary cell-specific transcription factor Pit-1, we examined whether a Pit-1 binding site could direct a response to Q209L-alpha q. Two copies of prolactin promoter Pit-1 binding site 1P conferred upon a heterologous metallothionein promoter a response to Q209L-alpha q, implying an involvement of this site in the transcriptional action of Q209L-alpha q on the prolactin promoter. The phorbol ester activator of protein kinase C, 12-O-tetradecanoylphorbol-13-acetate, stimulated (-187)PRL-CAT activity, but opposed the action of Q209L-alpha q on activity of this PRL-CAT construct. Q209L-alpha q stimulation of (-187)PRL-CAT activity was inhibited by co-expression of a dominant negative Raf mutant, Raf-C4, but not by a point mutant of Raf-C4 with reduced inhibitory properties. These results imply that activated alpha q subunits can stimulate prolactin promoter activity via a pathway that involves a Pit-1 DNA binding site(s), is opposed by protein kinase C, and is mediated by a pathway in which Raf-1 kinase plays a role.
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PMID:Constitutively active Gq-alpha stimulates prolactin promoter activity via a pathway involving Raf activity. 748 29


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