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)

A series of cDNA clones coding for the rat liver interleukin 6 receptor (IL6-R) were isolated from an acute-phase library. The identity of the clones was established (a) by DNA sequence analysis and comparison with the known human leukocyte IL6-R and (b) by demonstrating that the clones generated specific IL6 ligand binding activity and IL6-dependent regulation of acute-phase gene control elements after transfection into appropriate recipient cells. Two types of cDNA clones were obtained corresponding to two mRNA species of different length, both encoding an identical protein of 462 amino acid residues. The two prototype clones, pRIL6RC.21 and pRIL6RC.6 contained 3'-untranslated regions of 550 and 3100 nucleotides, respectively. The sequence motifs TTATTTAT and ATTTA associated with the regulation of mRNA stability and translation efficiency were present only in the longer mRNA species. The deduced amino acid sequence of the rat liver IL6-R was 53% identical with the human leukocyte IL6-R. Both receptors contained conserved structural features in their extracellular domains, including the signal peptide, a C2 domain characteristic of the immunoglobulin superfamily, and two domains shared among members of a family of cytokine and growth factor receptors. The strongly conserved intracellular portion of the rat liver IL6-R lacked recognizable signal transduction domains. The cDNA clones were used to demonstrate that rat liver IL6-R mRNA concentrations were increased 4.2-fold at 12 h after the induction of an experimental acute-phase response. Clone pRIL6R.21ex, but not clone pRIL6RC.6ex, generated specific IL6 ligand binding activity after transfection into human Jurkat cells that lack the endogenous IL6-R. By contrast, only pRIL6RC.6ex reconstituted a response of human Hep3B-2 and HepG2 hepatoma cells to mouse IL6. These human hepatoma cells were highly responsive to human IL6 but did not respond to physiologic concentrations of murine IL6. After cotransfection with pRIL6RC.6ex and plasmids containing the chloramphenicol acetyltransferase reporter gene under the control of IL6 response elements of acute-phase plasma protein genes, these cells showed a strong stimulation of the reporter gene by recombinant mouse IL6. Thus, both cell surface ligand binding activity and the complete IL6 signal cascade terminating in the transcriptional induction of IL6-dependent promoters were successfully reconstituted. Therefore, both IL6-R mRNA species code for functionally active receptor, depending on the target cell, but only the longer mRNA species coded for significant receptor levels in human hepatoma cell lines.
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PMID:Molecular cloning, characterization and functional expression of the rat liver interleukin 6 receptor. 217 54

The cytokines interleukin-1 beta (IL-1 beta) and tumor necrosis factor alpha (TNF-alpha) are released by mononuclear phagocytes in vitro after stimulation with mycobacteria and are considered to mediate pathophysiologic events, including granuloma formation and systemic symptoms. We demonstrated that the Mycobacterium tuberculosis cell wall component lipoarabinomannan (LAM) is a very potent inducer of IL-1 beta gene expression in human monocytes and investigated the mechanism of this effect. We localized the LAM-, lipopolysaccharide (LPS)-, and TNF-alpha-inducible promoter activity to a -131/+15 (positions -131 to +15) DNA fragment of the IL-1 beta gene by deletion analysis and chloramphenicol acetyltransferase assay. Within this DNA fragment, there were two novel 9-bp motifs (-90/-82 and -40/-32) with high homology to the nuclear factor-IL6 (NF-IL6) binding site. Site-directed mutagenesis demonstrated that the two NF-IL-6 motifs could be independently activated by LAM, LPS, or TNF-alpha and that they acted in an orientation-independent manner. DNA mobility shift assay revealed specific binding of nuclear protein(s) from LAM-, LPS-, or TNF-alpha-stimulated THP-1 cells to the NF-IL6 motifs. We conclude that the two NF-IL6 sites mediate induction of IL-1 beta in response to the stimuli LAM, LPS, and TNF-alpha.
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PMID:Regulation of the interleukin-1 beta (IL-1 beta) gene by mycobacterial components and lipopolysaccharide is mediated by two nuclear factor-IL6 motifs. 768 3

Tuberculosis has emerged as an epidemic fueled by the large number of individuals infected with the human immunodeficiency virus, especially those who are injecting drug users. We found a striking increase from 4- to 208-fold in p24 levels in bronchoalveolar lavage fluid from involved sites of Mycobacterium tuberculosis infection vs uninvolved sites in three HIV+ patients. We used an in vitro cell culture model to determine if tuberculosis could activate replication of HIV-1. Mononuclear phagocyte cell lines U937 and THP-1 infected with HIV-1JR-CSF, in vitro and stimulated with live M. tuberculosis H37Ra, had a threefold increase in p24 in culture supernatants. Using the HIV-1 long terminal repeat with a chloramphenicol acetyltransferase (CAT) reporter construct, live M. tuberculosis increased transcription 20-fold in THP-1 cells, and cell wall components stimulated CAT expression to a lesser extent. The nuclear factor-kappa B enhancer element was responsible for the majority of the increased CAT activity although two upstream nuclear factor-IL6 sites may also contribute to enhanced transcription. Antibodies to TNF-alpha and IL-1 inhibited the increase in CAT activity of the HIV-1 long terminal repeat by M. tuberculosis from 21-fold to 8-fold. Stimulation of HIV-1 replication by M. tuberculosis may exacerbate dysfunction of the host immune response in dually infected individuals.
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PMID:Mycobacterium tuberculosis enhances human immunodeficiency virus-1 replication by transcriptional activation at the long terminal repeat. 773 95

Revealing the regulatory mechanisms involved in P-glycoprotein expression is important to our understanding of multidrug resistance (MDR) in tumor cells. The MDR1 gene encoding P-glycoprotein contained a promoter sequence (-157 to -125) that was found to be homologous with other mdr gene promoters and that specifically interacted with a nuclear protein. The nuclear protein was identified, using a HeLa lambda gt11 cDNA expression library, to be the transcriptional regulator nuclear factor for interleukin-6 (NF-IL6), a member of the C/EBP family of transcription factors that bound an NF-IL-6-like consensus element 5'-TTTCGCAGT-3'. Furthermore, a glutathione S-transferase fusion protein (10.1-glutathione S-transferase) containing the partial NF-IL6 cDNA was also found to specifically interact with the MDR1 promoter sequence. Co-transfection of an NF-IL6 expression vector with a chloramphenicol acetyltransferase reporter gene driven by 1018 base pairs of MDR1 5'-flanking sequences demonstrated that NF-IL6 trans-activated the MDR1 promoter. This trans-activation was significantly reduced when the NF-IL6 element in the reporter gene construct was deleted or mutated. Identification of NF-IL6 as an important transcriptional regulator and the implications of its potential role in MDR1 gene induction in response to a variety of stimuli are discussed.
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PMID:NF-IL6, a member of the C/EBP family of transcription factors, binds and trans-activates the human MDR1 gene promoter. 796 62

A glucocorticoid, dexamethasone, inhibited the production of a leukocyte chemotactic cytokine, interleukin 8 (IL-8), as well as mRNA expression by a glioblastoma cell line, T98G, stimulated with interleukin 1 (IL-1). Dexamethasone also inhibited IL-8 promoter-driven chloramphenicol acetyltransferase (CAT) activities induced by IL-1, suggesting that dexamethasone inhibited IL-8 production mainly at the transcriptional level. Moreover, CAT assay revealed that the nuclear factor-kappa B (NF-kappa B) binding site was the crucial cis-element required for conferring IL-1 responsiveness in conjunction with the CCAAT enhancer binding protein/nuclear factor-IL-6 (NF-IL6) and/or the AP-1 binding site(s). Mutation of either the AP-1 or NF-IL6 binding site did not abolish IL-8 gene repression by dexamethasone, suggesting that these sites were not targets for dexamethasone. Trimerized kappa B sequence in the IL-8 gene was enough for conferring the induction by IL-1 and inhibition by dexamethasone of CAT activity. Finally, dexamethasone diminished the IL-1-induced formation of NF-kappa B complexes, which were identified immunochemically to consist of p50 and p65, without reducing the amount of translocated factors. Collectively, dexamethasone interfered with the binding of the most essential transcription factor, NF-kappa B, to its cognate cis-element, thereby suppressing the transcription of IL-8 gene.
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PMID:Novel mechanism of glucocorticoid-mediated gene repression. Nuclear factor-kappa B is target for glucocorticoid-mediated interleukin 8 gene repression. 817 59

The expression of human papillomavirus type 16 (HPV16) early genes, including E6 and E7 transforming genes, is regulated by several cellular factors binding to the noncoding region (NCR), such as the glucocorticoid receptor, NF-I, and AP1, all of which are positive regulators. We demonstrated that the nuclear factor for interleukin 6 expression (NF-IL6) specifically binds to the HPV16 NCR ranging from nucleotides 7007 to 7766 and represses the early gene expression of HPV16. The responsive element in HPV16 NCR was determined within the region ranging from nucleotides 7454 to 7766. In this region, many binding sites for other cellular transactivators, such as NF-I and AP1, have been detected. Interestingly, three of seven binding sites for NF-I and two of two binding sites for AP1 in this region overlap with the putative NF-IL6 binding sites identified by computer analysis. Competition experiments with the oligonucleotides containing such NF-I or AP1 sites indicated that NF-IL6 certainly binds to them. Furthermore, in a chloramphenicol acetyltransferase assay using mutant NF-IL6 expression vectors, the DNA binding domain of NF-IL6 was shown to be necessary for repression, whereas the functional domain was not. These findings indicate that repression may be caused by competition with other transcriptional activators, such as NF-I and AP1. Thus, NF-IL6 may play a significant role in the regulation of viral transcription as a part of the host's resistance to viral infection.
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PMID:NF-IL6 represses early gene expression of human papillomavirus type 16 through binding to the noncoding region. 838 Apr 54

Respiratory syncytial virus (RSV), the most common etiologic agent of epidemic pediatric respiratory disease, infects and replicates in the human airway epithelium, resulting in the induction of cellular gene products essential for immune and inflammatory responses. We describe the effect of RSV infection on nuclear factor-IL6 (NF-IL6) expression, a human basic domain-leucine zipper-containing transcription factor that alone in combination with other inducible transcription factors regulates the expression of cytokine and adhesion molecule genes. RSV-infected human type II pulmonary alveolar epithelial cells (A549) synthesize a single 45.7-kDa isoform of NF-IL6 rapidly and in a time-dependent manner. NF-IL6 is first detectable after 3 h of infection and continues to accumulate until 48 h (until the cells lose viability). NF-IL6 production could not be induced by UV-inactivated virus, demonstrating the requirement of viral replication for NF-IL6 synthesis. Immunoprecipitation after [35S]methionine metabolic labeling was done to investigate the mechanism for NF-IL6 production. There was robust NF-IL6 protein synthesis within RSV-infected (24 h) cells. Protein synthesis occurred without detectable changes in the abundance or size of the single 1.8-kb NF-IL6 mRNA. RNase protection assay of transfected chloramphenicol acetyltransferase reporter genes driven by either wild-type or mutated NF-IL6 binding sites show a virus-induced increase in NF-IL6-dependent transcription. These studies have demonstrated a novel inducible mechanism for translational control of NF-IL6 synthesis and identify this transcription factor as a potential effector of the host response to RSV infection.
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PMID:Inducible translational regulation of the NF-IL6 transcription factor by respiratory syncytial virus infection in pulmonary epithelial cells. 862 74

Retinoids inhibit the expression of migration inhibitory factor-related protein-8 (MRP-8), a marker of hyperproliferative or abnormal keratinocyte differentiation, in a retinoic acid receptor (RAR)-dependent manner in various cell culture systems. MRP-8 expression is also down-regulated in vivo in psoriatic lesions after topical application of an anti-psoriatic RARbeta/gamma-selective synthetic retinoid, tazarotene. We demonstrate that an MRP-8 promoter linked to a chloramphenicol acetyltransferase reporter (MRP8CAT) faithfully replicates the differentiation-specific regulation of the endogenous keratinocyte MRP-8 gene. Further, interferon gamma and serum-induced expression of MRP8CAT is inhibited by retinoid receptors in a ligand-dependent manner. We also show that NF-IL6 acts as a transcriptional enhancer of MRP-8, and that RARs inhibit MRP8CAT by inhibiting the enhancer action of nuclear factor-interleukin-6 (NF-IL6). The NF-IL6 antagonism function of RAR is a complex of the core of the DNA binding domain and the hydrophobic zipper region. This manuscript identifies NF-IL6 as another transcription factor, in addition to AP1, whose activity is inhibited by RAR in a ligand-dependent manner. The interdiction of NF-IL6-dependent signal transduction pathway by RARs may explain some of the therapeutic effects of retinoids in inflammatory and proliferative diseases.
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PMID:Retinoic acid receptor-nuclear factor-interleukin 6 antagonism. A novel mechanism of retinoid-dependent inhibition of a keratinocyte hyperproliferative differentiation marker. 932 72

The human T-cell leukemia virus type I (HTLV-I), which infects a wide variety of mammalian cells including monocytes and macrophages, encodes a transactivating protein designated as Tax. We now report that Tax induces the human prointerleukin-1beta (IL1B) gene promoter in monocytic cells. In our transient transfection assays using human THP-1 monocytic cells, a chloramphenicol acetyltransferase (CAT) construct containing the IL1B promoter sequence between positions -131 and +12 showed an approximately 90-fold increase in activity following cotransfection of a Tax expression vector. Moreover, Tax synergized with lipopolysaccharide (LPS) to induce the IL1B promoter activity. Analyses of specific nucleotide substitutions further indicated that the Tax-induced transcriptional activation requires two transcription factor binding motifs within the IL1B promoter; one is a binding site for nuclear factor (NF)-IL6 (CCAAT/enhancer binding protein beta, C/EBP beta), which belongs to the basic region-leucine zipper (bZIP) family and the other for Spi-1 (PU.1), which is an Ets family protein found principally in monocytes, macrophages, and B lymphocytes. In electrophoretic mobility shift assays (EMSA) using in vivo THP-1 nuclear extracts, Tax expression in THP-1 monocytic cells significantly increased binding of the two factors to their target IL1B promoter sequences. However, in contrast to NF-IL6 and Spi-1, DNA binding activity of Oct-1, an ubiquitously expressed octamer-binding protein was not affected by Tax. Additional EMSA using in vitro translated proteins also showed that recombinant Tax enhances DNA binding of both of recombinant NF-IL6 and Spi-1 proteins. These data were supported by our glutathione S-transferase (GST)-pull-down data, which indicated that Tax physically interacts with the two proteins. Based on the results obtained from the present study, we conclude that the IL1B promoter is a Tax-responsive sequence as a result of ability of Tax to induce binding of NF-IL6 and Spi-1 to the IL1B promoter sequence through protein-protein interaction.
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PMID:Human T-cell leukemia virus type I Tax transactivates the promoter of human prointerleukin-1beta gene through association with two transcription factors, nuclear factor-interleukin-6 and Spi-1. 937 96

Vascular smooth muscle cells (SMC) are major constituents of the medial layer of blood vessels and are involved in the development of atherosclerotic plaque. SMC secrete copious IL-6 under basal conditions that can be increased by cytokines such as tumor necrosis factor-alpha and interleukin-1beta (IL-1beta). The goal of our studies was to define the role of estrogen in IL-6 production by SMC. In a first series of experiments, the expression of specific messenger RNAs as well as the production of IL-6 bioactivity by rat SMC in culture could be demonstrated in basal and IL-1-stimulated conditions, but was unaffected by estrogen treatment. Different constructs containing deleted or mutated fragments of the human IL-6 promoter driving luciferase or chloramphenicol acetyltransferase reporter gene were then transiently transfected in these cells. A significant basal activity that was increased 2- to 4-fold after IL-1beta stimulation was observed with the total IL-6 promoter. Deletion analysis indicated that the -158/+11 region containing activator protein-1 and cAMP response element sites was apparently the minimal region of IL-6 promoter to confer both constitutive and IL-1-inducible activities. Site-directed mutagenesis experiments suggest that basal activity is dependent upon the promoter sequence -158 to -112 containing the nuclear factor (NF)-IL6(-153) and Sp1 sites, whereas IL-1beta stimulation would depend on the residual -112 nucleotides containing NF-IL6(-75) and NF-kappaB sites. In contrast to the down-regulation of IL-6 expression by estrogen described in osteoblasts, ethinyl estradiol as well as 17beta-estradiol did not influence stimulated IL-6 activity in our experimental conditions whatever the construct tested, even when either estrogen receptor alpha or beta was overexpressed. Thus, the atheroprotective properties of estrogen are probably not mediated through the regulation of IL-6 production by SMC.
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PMID:Expression of the interleukin-6 gene is constitutive and not regulated by estrogen in rat vascular smooth muscle cells in culture. 1034 80


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