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)

Here we investigate the roles of human testicular orphan receptors, TR2 and TR4, on the gene regulation of the long-terminal repeat of the human immunodeficiency virus type 1 (HIV-LTR). In gel-retardation assays, a palindromic element at the 5'-end of HIV-LTR,5'-AGGGGTCAGATATCCACTGACCTTT-3',showed high affinity to TR2 and TR4 with an equilibrium dissociation constant (Kd) of 1.11 +/- 0.48 (n = 3) and 0.52 +/- 0.12 nM (n = 3), respectively. Interestingly, each half-site of the palindromic element is sufficient to compete with the binding of the labeled palindromic element to TR2 or TR4 with an equilibrium inhibition constant (ki) around 10 nM. However, the transiently expressed TR2 or TR4 in Chinese hamster ovary (CHO) cells or Japanese quail muscle myoblasts (QM7) cells showed no activity in regulating the transcriptional activity of the chloramphenicol acetyltransferase (CAT) reporter gene inserted downstream of the HIV-LTR promoter. Although both TR2 and TR4 showed no effect on CAT activity by itself, our data showed only the TR4 could crosstalk to the chicken ovalbumin upstream protein-transcription factor (COUP-TF1) and thyroid hormone receptor (TR alpha 1), and potentiated the transcriptional activity of HIV-LTR on the CAT reporter gene regulated by COUP-TF1 and TR alpha 1. These results indicate that TR4, but not TR2, may couple to other nuclear receptors in the upregulation of the HIV replication.
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PMID:TR4 orphan receptor crosstalks to chicken ovalbumin upstream protein-transcription factor and thyroid hormone receptor to induce the transcriptional activity of the human immunodeficiency virus type 1 long-terminal repeat. 970 74

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

Several laboratories have presented evidence that HIV can productively infect CD4- cell lines. However, this data could be challenged on the basis that the target cells may express low levels of the CD4 receptor. In addition, it could be argued that assays might be detecting residual virus. In the case of cell-mediated infection, it is possible that virus detected in assays could be secreted from HIV-infected donor cells rather than the target CD4- cells. In this report we describe a CD4- epithelial cell line which has been transfected with a plasmid containing the chloramphenicol acetyltransferase (CAT) gene ligated to the HIV LTR. CAT-ELISA and immunocytochemistry indicate that target cells synthesize CAT after exposure to HIV-infected primary activated peripheral blood mononuclear cells (PBMC). Results correlate very well with p24 ELISA assays. Infection of epithelia by primary NSI strains of HIV can be blocked by patient antisera or by certain sulfated polysaccharides. Since the CAT assay is not dependent on virus production, the data reported here confirm that CD4- epithelial cells derived from the human cervix can be productively infected by HIV. The observations also support the theory that sexual transmission of HIV could be initiated by infection of genital tract epithelia. Furthermore, the findings support the suppositions that sexual transmission of HIV could be prevented by antibodies to HIV or alternately by a topical formulation containing certain sulfated polysaccharides.
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PMID:CAT-transfected epithelial cells provide evidence for a CD4 independent pathway of HIV infection. 1021 19

p73 is a recently cloned tumor suppressor gene that is highly homologous to p53, and the products of both possess similar functions in inhibiting cell growth and inducing apoptosis. Interestingly, the COOH-terminal region of p53 displays no significant homology with that of p73. Moreover, p73 has an additional segment at its COOH terminus. Recently, we have found two mutations of p73 with amino acid substitution (P405R and P425L) in primary neuroblastomas. Because the region (amino acid residues 382-491) contains a glutamine- and proline-rich domain, we hypothesized that it has a transactivation function, and the mutations found in tumors result in loss of function. To test it, we used the yeast GAL4 DNA-binding fusion system. Yeast transformants expressing a GAL4-p73(1-112) or a GAL4-p73alpha(380-513) fusion protein were grown in SD medium lacking histidine and tryptophan and exhibited a significant induction of beta-galactosidase activity. Transient transfection experiments revealed that both of fusion proteins could induce the chloramphenicol acetyltransferase activity in mammalian cells, indicating that the COOH-terminal as well as NH2-terminal regions of p73 had significantly high levels of transactivation activity. Furthermore, the former activity was severely impaired in two naturally occurring mutant forms found in neuroblastomas. These suggest that, unlike p53, p73 has two domains with transactivation function, one in the NH2-terminal region and the other in the COOH-terminal region. Loss of function mutation in the latter might be involved in tumorigenesis and/or tumor progression.
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PMID:Identification of a transactivation activity in the COOH-terminal region of p73 which is impaired in the naturally occurring mutants found in human neuroblastomas. 1038 37

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

We have previously reported the isolation of mutant cell lines from the human carcinoma line ME180 that are resistant to the antiproliferative effect of interferon-gamma (IFN-gamma). These cell lines were defective in the induction of indoleamine 2,3-dioxygenase (IDO), a key enzyme of tryptophan catabolism. One of these cell lines, 3B6A, was chosen for further study. This cell line was also defective in the ability of IFN-gamma to protect against vesicular stomatitis virus (VSV) infection. However it maintained a normal antiviral response to IFN-alpha. A promoter-chloramphenicol acetyltransferase (CAT) construct containing the promoter region of IDO, which includes IFN-gamma activation site (GAS), IFN-stimulated response element-1 (ISRE-1), and ISRE-2 regions, was not expressed in 3B6A in the presence of IFN-gamma, indicating that the defect was likely to be in either Stat1 or IFN regulatory factor-1 (IRF-1), transcription factors known to bind to these cis-acting sequences. The induction of other IFN-gamma-inducible genes, such as tryptophanyl-tRNA synthetase (hWRS), was also affected. Electrophoretic mobility shift assays (EMSA) comparing nuclear extracts from parental and mutant cells indicated that Stat1 from the mutant did not bind to GAS sequences. However, Western blot analysis indicated that Stat1 protein was present. This IDO-negative phenotype can be reversed by transfection with a Stat1 expression vector. DNA sequencing of the Stat1 cDNA from wild-type and 3B6A cells indicated that an amino acid change occurred in the Stat1 protein of the mutant at W573, a tryptophan conserved in all known Stat proteins. We hypothesize that a change in this region of the Stat protein affects the response to IFN-gamma but not to IFN-alpha.
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PMID:An indoleamine 2,3-dioxygenase-negative mutant is defective in stat1 DNA binding: differential response to IFN-gamma and IFN-alpha. 1092 4

Cyclopentenone prostaglandins inhibit virus replication in several DNA and RNA virus models. In this report we investigated the effect of prostaglandin A1 (PGA1) on HIV-1 transcription in human CD4+ Jurkat T lymphocyte cells. A dramatic reduction of HIV-1 RNA levels was detected up to seven days post infection in both unstimulated and phorbol 12-mystrate 13-acetate (PMA)-stimulated cells treated with PGA1- PGA1 treatment of cells was also effective in inhibiting the transcription of a chloramphenicol acetyltransferase (CAT) reporter gene, under the control of HIV-1 LTR, in Jurkat-Tat cells. We also show that PGA1 induced the synthesis of 70-kDa heat-shock protein (HSP70) in this cell system and the induction correlated with the drug-antiviral activity. PGA1 was also found to induce the loss of the tumor suppressor p53 protein, in the "proliferative" conformation, in a time correlation with the induction of the HSP70 As the "proliferative" p53 has been involved in the positive trans-activation of the HIV-1 LTR its depletion could contribute to the inhibitory mechanisms of PGA1 on virus transcription.
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PMID:Inhibition of HIV-1 transcription by cyclopentenone prostaglandin A1 in Jurkat T lymphocytes. 1103 55

Substance P (SP) is a potent modulator of neuroimmunoregulation. SP receptors are present on human monocytes and T lymphocytes, and SP alters the function of these immune cells. We investigated the effects of SP on HIV-1 replication in latently infected human immune cells. SP significantly enhanced HIV-1 replication in the latently infected promonocytic cell line (U1) and T lymphocyte line (ACH-2) stimulated with tumor necrosis factor (TNF-alpha). When added to these cells in combination with TNF-alpha, SP also enhanced HIV-1 gag gene expression in U1 and ACH-2 cells. This stimulatory effect of SP was associated with the activation of HIV-LTR (long terminal repeat) driven chloramphenicol acetyltransferase (CAT) gene expression, and could be blocked by pretreatment of U1 and ACH-2 cells with an SP receptor antagonist RP-67,580, indicating specific SP receptor-mediated regulation. Furthermore, the addition of SP to the cultures of latently infected peripheral blood mononuclear cells isolated from HIV-1-infected patients enhanced HIV-1 gag gene expression. Thus, SP may play a potentially important role as a positive regulator of HIV-1 replication in latently infected monocytes and lymphocytes. These observations may have significant implications toward understanding the role of neuropeptide SP in the immunopathogenesis of HIV-1 infection and AIDS.
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PMID:Substance P enhances HIV-1 replication in latently infected human immune cells. 1173 Sep 41

Coincubation of monocytoid cell line U937 cells cotransfected with HIV-1 LTR CAT plasmid and Tat expression plasmid, with Mycobacterium smegmatis, M. avium, M. bovis BCG and M. tuberculosis enhanced chloramphenicol acetyltransferase (CAT) production, indicating that these mycobacteria could activate the LTR in this cell line. The amount of CAT in the cells coincubated with M. smegmatis was higher than that infected with the other mycobacteria after 12, 24 and 48 hour time periods. However, the amount of CAT production in the cells cocultured with M. tuberculosis was higher than those coincubated with the other mycobacteria at 72 hours. These findings indicated that avirulent mycobacteria such as M. smegmatis may activate HIV replication at an early time and its effects are gradually decreased, while the effect of virulent M. tuberculosis increased gradually, and lasted for a long time resulting in an acceleration of HIV disease in patients.
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PMID:Changed activation of HIV-1 LTR in monocytoid cells by mycobacteria with temporal progression of infection. 1217 80

RNA polymerase II transcripts are confined to nuclear compartments. A detailed analysis of the nuclear topology of RNA from individual genes was performed for transcripts from the marker gene coding for chloramphenicol acetyltransferase, expressed at a high level from the HTLV-1 LTR promoter. The construct was transfected into A293 cells where the RNA was organized as an extensive reticular network. We also studied the RNA distribution from combinations of neighboring HIV and bacterial resistance genes that co-integrated within the genome of COS-7 cells-revealing spherical or track-like accumulations of RNA that were extensively branched. There were many nuclei with distinct but overlapping RNA accumulations. Since the coding genes localized at the overlapping points, the RNAs are synthesized at a common region and diverge. The correlation between the frequency of the separation of the transcripts and the physical distance of the respective genes suggests a subcompartmentalization in the microenvironment of genes on the basis of geometric parameters. Thus, the more distant the genes are on the same chromosome, the more likely they are confined to separated subcompartments of an extensive reticular system. Co-delineation of the RNA transcripts with Cajal bodies and chromosome territories indicated the organization of nuclear RNA transcripts in a reticular interchromosome domain compartment.
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PMID:Nuclear RNAs confined to a reticular compartment between chromosome territories. 1556 Nov


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