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)

Phenobarbital is an inducer of xenobiotic-metabolizing enzymes, such as cytochrome P-450, glutathione S-transferases (GSTs) and NAD(P)H:quinone reductase, as well as being a promoter of hepatocarcinogenesis. The molecular mechanisms regulating these biological activities are, however, unknown. In this paper we show that induction by phenobarbital of GST Ya and quinone reductase gene expression is mediated by regulatory elements, EpRE and ARE respectively, which are composed of two adjacent AP-1-like binding sites. EpRE was recently found to be activated by a Fos/Jun heterodimeric complex (AP-1). Here we show that phenobarbital induces an increase in AP-1 binding activity in nuclear extracts of cultured hepatoma cells. Furthermore, we observe that the induction of chloramphenicol acetyltransferase (CAT) activity from an EpRE Ya-cat gene construct and of AP-1 binding activity by phenobarbital is inhibited by the thiol compounds N-acetyl-L-cysteine and glutathione. These results suggest that the phenobarbital induction of AP-1 activity, leading to the AP-1-mediated transcriptional activation of the GST Ya and quinone reductase genes, may involve production of reactive oxygen species and an increase in intracellular oxidant levels, which is prevented by thiol compounds. In view of the involvement of AP-1 in the control of cell proliferation and transformation, the induction by phenobarbital of AP-1 binding activity observed here provides a possible molecular mechanism for the tumour-promoting activity of this drug.
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PMID:Phenobarbital induction of AP-1 binding activity mediates activation of glutathione S-transferase and quinone reductase gene expression. 845 90

We have investigated an oxidoreductive regulatory pathway for the DNA binding activity of a pleiotropic cellular transcription factor, nuclear factor kappa B (NF kappa B), has been investigated by using NF kappa B prepared from the nucleus and the cytosol of the primary human T lymphocytes. We show that a cellular reducing catalyst thioredoxin (Trx) plays a major role in activation of the DNA binding of NF kappa B in vitro and stimulation of transcription from the NF kappa B-dependent gene expression. We demonstrate evidence suggesting that redox regulation of NF kappa B by Trx might be exerted at a step after dissociation of the inhibitory molecule I kappa B, a cytosolic-anchoring protein for NF kappa B. To examine the effect of Trx in intact cells, we performed transient assay with a chloramphenicol acetyltransferase-expressing plasmid under the control of human immunodeficiency virus (HIV) long terminal repeat and an effector plasmid expressing human Trx. The promoter activity from HIV long terminal repeat was greatly augmented by co-transfecting the Trx-expressing plasmid, whose effect was dependent on the NF kappa B-binding sites. These findings have suggested that cysteine residue(s) of NF kappa B might be involved in the DNA-recognition by NF kappa B and that the redox control mechanism mediated by Trx might have a regulatory role in the NF kappa B-mediated gene expression. These results may also provide a clue to understanding of the molecular process of AIDS pathogenesis and its possible biochemical intervention.
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PMID:Oxidoreductive regulation of nuclear factor kappa B. Involvement of a cellular reducing catalyst thioredoxin. 849 88

Transgenic tobacco (Nicotiana tabacum L.) plants carrying a fusion between the nopaline synthase (nos) promoter and chloramphenicol acetyltransferase (CAT) reporter gene (caf) were tested for their response to treatment with H2O2. The nos promoter-driven CAT activity increased significantly by addition of H2O2, reaching the maximum level at 15 mM. Kinetic analysis for CAT activity showed that induction by H2O2 was similar to that of methyl jasmonate (MJ), but was much slower than induction by salicylic acid (SA). Time-course experiments for mRNA level also revealed that the response to H2O2 treatment was similar to that of MJ. The nos promoter displayed a rapid and transient induction of mRNA with SA treatment, with the maximum levels occurring at 3 h, whereas the levels induced by H2O2 or MJ treatment increased continuously during the 11-h experimental period. The antioxidants N-acetyl-L-cysteine and catechol did not alter the SA effect. The responses of the nos promoter to H2O2, MJ, and wounding were significantly reduced by deletions of the CAAT box region and the sequence between -112 and -101. However, these deletions did not significantly alter the SA response. This suggests that H2O2 may have a different mechanism from that of SA for inducing nos promotor activity.
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PMID:Induction of nopaline synthase promoter activity by H2O2 has no direct correlation with salicylic acid. 853 87

High dietary intakes of unsaturated fats may be atherogenic by disrupting normal functions of the vascular endothelium, due in part to the ability of linoleic acid (18:2n-6) to contribute to an increase in cellular oxidative stress and related injurious events. Exposing endothelial cells to 90 micromol linoleic acid/L for 6 h resulted in a significant increase in lipid hydroperoxides that coincided wih an increase in intracellular calcium concentrations. Treatment with this fatty acid caused an initial decrease in glutathione concentrations, which was followed by an increase at later time points. Most importantly, a significant activation of the oxidative stress-sensitive nuclear transcription factor-kappa B (NF-kappa B) was achieved after a 6-h exposure to 18:2n-6, which is the time point at which maximal depletion of cellular glutathione was observed. The fatty acid-mediated NF-kappa B activation was accompanied by induction of NF-kappa B-dependent transcription, as measured by chloramphenicol acetyltransferase (CAT) assay of an NF-kappa B-responsive promoter construct. Pretreatment of endothelial cells with vitamin E and N-acetyl cysteine inhibited the fatty acid-induced activation of NF-kappa B and formation of lipid hydroperoxides. These data suggest that oxidative stress-induced cellular changes are critical early events in fatty acid-mediated endothelial cell dysfunction.
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PMID:Linoleic acid activates nuclear transcription factor-kappa B (NF-kappa B) and induces NF-kappa B-dependent transcription in cultured endothelial cells. 860 87

Nuclear factor kappaB (NF-kappaB) is an important cellular regulator of human immunodeficiency virus (HIV) gene expression. In T cells, N-acetyl-L-cysteine (NAC) inhibits the induction of NF-kappaB and transcription of HIV-1. However, NAC up-regulates HIV-1 replication in monocyte-derived macrophages (MDM). In this study we demonstrate that NAC treatment of MDM transfected with a chloramphenicol acetyltransferase (CAT) construct under transcriptional control of the HIV-1 long terminal repeat resulted in an up-regulation of CAT activity. Furthermore, MDM transfected with a HIV-1-NF-kappaB-CAT construct also produced increased CAT activity after NAC treatment. In addition, electrophoretic mobility shift assays revealed that nuclei of NAC-treated MDM contained increased binding activity to wild-type, but not mutant, kappaB oligonucleotides. Components of the binding activity were identified with antibodies as the NF-kappaB subunits p50 and p65. These data indicate that NAC-induced enhancement of HIV-1 replication in MDM is regulated at the level of viral gene expression and mediated by NF-kappaB.
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PMID:N-acetyl-L-cysteine-induced up-regulation of HIV-1 gene expression in monocyte-derived macrophages correlates with increased NF-kappaB DNA binding activity. 900 May 34

LPS-induced expression of the IL-8 gene was markedly enhanced by H2O2 or by deprivation of the cellular antioxidant glutathione by L-buthionine-(S,R)-sulfoximine (BSO) in human astrocytoma U373 cells. In contrast, it was markedly suppressed by the reductant N-acetyl-L-cysteine (NAC) and other antioxidants. Transient expression analysis using the chloramphenicol acetyltransferase assay revealed that activation of the IL-8 promoter by LPS was stimulated by BSO and was suppressed by NAC; likewise LPS-induced activation of both NF kappa B and AP-1 was enhanced by BSO and inhibited by NAC. These results suggest that LPS-induced IL-8 gene expression is regulated by cellular redox via modulation of these transcription factors.
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PMID:Redox regulation of lipopolysaccharide (LPS)-induced interleukin-8 (IL-8) gene expression mediated by NF kappa B and AP-1 in human astrocytoma U373 cells. 912 24

Endogenous sulphated polysaccharides such as heparin have been shown to inhibit the infectivity of HIV-1 min vitro. However, these naturally occurring polymers, due to extensive microheterogeneity within their structure, are difficult to characterise accurately. In contrast, dextrin can be chemically sulphated to produce a series of compounds sulphated in the 2-, 3-, or 6- position, or in all 3 positions, and the use of these compounds provides an opportunity to investigate the anti-HIV-1 activity of sulphated polysaccharides. The mechanisms whereby sulphated polysaccharides exert their anti-HIV-1 activity have not been fully elucidated. The interaction of recombinant HIV-1 proteins with sulphated polysaccharides was investigated using a biotinylated derivative of dextrin 2-sulphate (D2S) in a solid phase binding system. D2S was found to bind strongly to HIV-1 tat (EC50 = 0.10 microg/mL), less strongly to CD4 (EC50 = 0.33 microg/mL), weakly to HIV-1 vif and gp160, and not at all to HIV-1 gp120 or p24. Other sulphated derivatives of dextrin, i.e. dextrin 3-sulphate, dextrin 6-sulphate and dextrin 2,3,6-trisulphate, as well as heparin and dextran sulphate, were also shown to bind to HIV-1 tat, whereas the unsulphated compound dextrin did not. Binding studies using a series of overlapping peptides representing the complete sequence of HIV-1 tat revealed that D2S bound most strongly to the core domain of HIV-1 tat, although there was also binding to the cysteine-rich domain; both of these regions are important for HIV-1 tat function. In assessing function, HIV-1 tat-mediated transactivation was measured using H938 cells, a cell line that contains the HIV-LTR (long terminal repeat) promoter linked to a chloramphenicol acetyltransferase gene. D2S significantly inhibited HIV-1 tat transactivation in a dose-dependent manner (IC50 = 0.5 microg/mL), whereas dextrin had no effect. The interaction between D2S and HIV-1 tat provides a potential mechanism of HIV-1 inhibition whereby tat is sequestered and its transactivating activity abolished, effectively inhibiting the replication cycle.
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PMID:Interaction of the transactivating protein HIV-1 tat with sulphated polysaccharides. 1007 83

Pyrrolidine dithiocarbamate (PDTC) has been widely used as an inhibitor of the nuclear factor-kappa B, (NF-kappa B) signalling pathway. Here, we show that kappa B-dependent reporter gene expression induced by low concentrations of 12-O-tetradecanoylphorbol-13-acetate (TPA) is potentiated by PDTC in the human pro-monocytic U937 cell line. The stimulatory effect of PDTC on kappa B-dependent gene expression was shown with a 4 x kappa B chloramphenicol acetyltransferase construct and required an intact kappa B element in the human immunodeficiency virus long terminal repeat (HIV-1 LTR). Unexpectedly, an HIV-1 LTR construct with a mutation of the activator protein 2 (AP-2) binding site located between the two kappa B elements was unresponsive to the stimulatory effect of PDTC with TPA. The stimulation or inhibition of kappa B-dependent gene expression was dependent on PDTC pre-treatment and the concentration of TPA. No stimulatory effect on HIV-1 LTR activity was observed with the metal chelator dipyridyl or the anti-oxidant N-acetyl-L-cysteine. These results are consistent with the hypothesis that PDTC treatment potentiated kappa B-dependent gene expression in a manner dependent on the concentration of TPA.
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PMID:Dual activity of pyrrolidine dithiocarbamate on kappa B-dependent gene expression in U937 cells: I. Regulation by the phorbol ester TPA. 1040 58

Prior studies have demonstrated that the pineal hormone, melatonin, can stimulate chloramphenicol acetyltransferase activity in Drosophila SL-3 cells transfected with a chloramphenicol acetyltransferase reporter construct containing the response element of rat bone sialoprotein (BSP). Based on these findings, studies were performed to determine whether melatonin could similarly modulate the expression of BSP in two cell lines, the MC3T3-E1(MC3T3) pre-osteoblast and rat osteoblast-like osteosarcoma 17/2.8 cell. Initial studies demonstrated that MC3T3 cells grown in the presence of 50 nM melatonin underwent cell differentiation and mineralization by day 12 instead of the 21-day period normally required for cells grown in untreated media. Melatonin increased gene expression of BSP and the other bone marker proteins, including alkaline phosphatase (ALP); osteopontin; secreted protein, acidic and rich in cysteine; and osteocalcin in MC3T3 cells in a concentration-dependent manner. Levels of melatonin as low as 10 nM were capable of stimulating transcription of these genes when cells were grown in the presence of beta-glycerophosphate and ascorbic acid. Under these conditions, melatonin induced gene expression of the bone marker proteins; however, this does not occur until the 5th day after seeding the culture dishes. Thereafter, MC3T3 cells responded to melatonin within 2 h of treatment. The fully differentiated rat osteoblast-like osteosarcoma 17/2.8 cells responded rapidly to melatonin and displayed an increase in the expression of BSP, ALP, and osteocalcin genes within 1 h of exposure to the hormone. To determine whether melatonin-induced osteoblast differentiation and bone formation are mediated via the transmembrane receptor, MC3T3 cells were treated in the presence and absence of melatonin with either luzindole, a competitive inhibitor of the binding of melatonin to the transmembrane receptors, or pertussis toxin, an uncoupler of G(i) from adenylate cyclase. Both luzindole and pertussis toxin were shown to reduce melatonin-induced expression of BSP and ALP. These results demonstrate, for the first time, that the pineal hormone, melatonin, is capable of promoting osteoblast differentiation and mineralization of matrix in culture and suggest that this hormone may play an essential role in regulating bone growth.
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PMID:Melatonin promotes osteoblast differentiation and bone formation. 1041 30

NorM, a putative efflux pump of Vibrio cholerae, is a member of the multidrug and toxic compound extrusion family of transporters. We demonstrate that NorM confers resistance to norfloxacin, ciprofloxacin, and ethidium bromide. Inactivation of norM rendered V. cholerae hypersensitive towards these fluoroquinolones. Multiple sequence alignment of members of its family identified several regions of high sequence conservation. The topology of NorM was determined using beta-lactamase and chloramphenicol acetyltransferase fusions. The amino acid residues G(184), K(185), G(187), P(189), E(190), G(192), and G(195) in the periplasmic loops and L(381), R(382), G(383), Y(384), K(385), and D(386) in the cytoplasmic loops, as well as all the acidic and cysteine residues of NorM, were mutated. Mutants G184V, G184W, K185I, P189S, E190K, and E190A lost the norfloxacin resistance-imparting phenotype characteristic of NorM. Mutants E124V, D155V, G187V, G187R, C196S, Y384H, Y384S, and Y384F exhibited partial resistance to norfloxacin. Mutants with replacements of G(184) or G(187) by A, K(185) by R, and E(190) by D retained the norfloxacin resistance phenotype of NorM. Analysis of the accumulation of norfloxacin in intact cells of Escherichia coli expressing NorM or its mutants in the presence or absence of carbonyl cyanide m-chlorophenylhydrazone supported the results obtained through susceptibility testing and argued in favor of NorM-mediated efflux as the determining factor in norfloxacin susceptibility in the genetically manipulated strains. Taken together, these results suggested that E(124), D(155), G(184), K(185), G(187), P(189), E(190), C(196), and Y(384) are likely involved in NorM-dependent norfloxacin efflux. Except for D(155), C(196), and Y(384), all of these residues are located in periplasmic loops.
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PMID:Analysis of the topology of Vibrio cholerae NorM and identification of amino acid residues involved in norfloxacin resistance. 1695 25


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