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:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cytochrome p4501A induction and subsequent enzyme expression is used as a biomarker for exposure to aryl hydrocarbon receptor active contaminants in fish and other species. In the present study, CYP1A cDNA (1912 bp, GenBank accession number AF364076) was cloned, sequenced and characterized from the liver of a beta-naphthoflavone (betaNF)-treated teleost, Atlantic salmon (Salmo salar), by reverse-transcriptase polymerase chain reaction (RT-PCR). The salmon CYP1A sequence contained a 5'-flanking region of 99 bp, an open reading frame of 1566 bp that encodes a 521 amino acid protein, a stop codon, and a 3'-untranslated region of 346 bp, and a single polyadenylated signal. The theoretical molecular mass and isoelectric point was 58.6 kDa and 6.17, respectively. Comparison of the deduced amino acid sequence of salmon cDNA with reported CYP1A genes showed identities of 73-88% with fish CYP1A, 51-54% with mammalian CYP1A1, 47-51% with mammalian CYP1A2 and 54% with frog p450. Phylogenetic analysis showed that salmon CYP1A clustered in the tree with rainbow trout CYP1A1 and eel CYP1A sequences. CYP1A mRNA induction in beta-naphthoflavone-treated salmon showed differential organ expression with a distinct single transcript pattern. A new specific molecular tool has been developed for the monitoring of environmental pollution using CYP1A mRNA from salmon as a biomarker.
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PMID:Complementary DNA cloning, sequence analysis and differential organ expression of beta-naphthoflavone-inducible cytochrome P4501A in Atlantic salmon (Salmo salar). 1245 89

Oltipraz, a promising cancer chemopreventive agent, has been recognized as a monofunctional inducer selectively activating phase II carcinogen-detoxifying enzymes via the antioxidant responsive element (ARE). However, we report here that oltipraz also induces rat glutathione S-transferase A5 (GSTA5), a potent phase II detoxifying enzyme, by means of the xenobiotic responsive element (XRE). Although an ARE sequence exists in the 5' upstream of the rGSTA5 gene, this cis-acting regulatory element loses its responsiveness to oltipraz treatment because of extensive mutations in its distal-half site. Our data indicate that a XRE sequence, located downstream of the transcription initiation site of the gene, is another oltipraz-responsive element. Electrophoretic mobility shift assay showed that oltipraz steadily induces XRE-aryl hydrocarbon receptor (AhR) binding, which can be blocked specifically by excess XRE oligonucleotides or by AhR antibody. By cloning different XREs into the pGL3-promoter vector, we found that oltipraz can activate XRE enhancers from several phase II drug metabolism enzymes, including rGSTA5, rGSTA2, NAD(P)H:quinone reductase, and it also activates XRE from the phase I metabolism enzyme CYP1A1. Oltipraz's effect on XRE is AhR-dependent and is independent of the presence of active CYP1A1. Reverse transcriptase-polymerase chain reaction experiments revealed that oltipraz induces gene expression of both phase I and II drug-metabolizing enzymes in rat hepatoma cells. Thus, we conclude that, like ARE, the XRE pathway constitutes an important part of the molecular mechanism contributing to oltipraz-induced expression of the phase II metabolism enzymes. Oltipraz is a bifunctional inducer, modulating both phase I and II drug-metabolizing enzymes to enhance carcinogen detoxification.
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PMID:Oltipraz is a bifunctional inducer activating both phase I and phase II drug-metabolizing enzymes via the xenobiotic responsive element. 1286 39

The expression of the cytochrome P450 1A1 gene (cyp1a1) is regulated by the aryl hydrocarbon receptor (AhR), which is a ligand-activated transcription factor that mediates most toxic responses induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In the nucleus, ligand-activated AhR binds to the xenobiotic response elements, initiating chromatin remodeling and recruitment of coregulators, leading to the formation of preinitiation complex followed by elongation. Here, we report that ligand-activated AhR recruits the positive transcription elongation factor (P-TEFb) and RNA polymerase II (RNA PII) to the cyp1a1 promoter with concomitant phosphorylation of the RNA PII carboxyl domain (CTD). Interestingly, the serine 2 and serine 5 of the heptapeptide repeats (YSPTSPS) were sequentially phosphorylated upon TCDD treatment. Inhibition of P-TEFb kinase activity by 5,6-dichloro-1-beta-d-ribofuranosyl-benzimidazole (DRB) suppressed CTD phosphorylation (especially serine 2 phosphorylation) and abolished processive elongation without disrupting the assembly of the preinitiation complex at the cyp1a1 promoter. Remarkably, we found that activation of NF-kappaB by TNF-alpha selectively inhibited TCDD-induced serine 2 phosphorylation in mouse liver cells, suggesting that residue-specific phosphorylation of RNA PII CTD at the cyp1a1 promoter is an important regulatory point upon which signal "cross-talk" converges. Finally, we show that ligand-activated AhR associated with P-TEFb through the C terminus of cyclin T1, suggesting that AhR recruit the P-TEFb to the cyp1a1 promoter whereupon its kinase subunit phosphorylates the RNA PII CTD.
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PMID:Interactions between the aryl hydrocarbon receptor and P-TEFb. Sequential recruitment of transcription factors and differential phosphorylation of C-terminal domain of RNA polymerase II at cyp1a1 promoter. 1291 20

Regulation of gene expression by the aryl hydrocarbon (AHR) receptor is a much-studied pathway of molecular toxicology. Activation of AHR by the xenobiotic ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is hypothesized as the mechanism by which TCDD exerts its toxic and carcinogenic effects. Paradoxically, some studies have shown that TCDD acts as an antiestrogen. This has led to the hypothesis that so-called selective aryl hydrocarbon receptor modulators (SAhRMs), AHR ligands that retain the antiestrogenic effects but lack the transcriptional effects of TCDD associated with toxicity, may be utilized as cancer chemotherapeutics in conjunction with other antiestrogenic compounds such as tamoxifen. The present study attempts to further define the molecular mechanism of action of the putative SAhRMs, 6-alkyl-1,3,8-trichlorodibenzofuran (6-MCDF), and diindolylmethane (DIM), focusing particularly on the former. We tested 6-MCDF and DIM for the recruitment of AHR and RNA polymerase II (pol II) to the regulatory region of the AHR responsive gene, cytochrome P4501A1 (CYP1A1), using the chromatin immunoprecipitation (ChIP) assay in the mouse hepatoma cell line Hepa-1c1c7 (Hepa-1). We also tested the level of CYP1A1 induction in Hepa-1 cells using quantitative real-time PCR. We show no difference in the recruitment of AHR or pol II to the regulatory region of CYP1A1 in response to TCDD, 6-MCDF, or co-treatment with both TCDD and 6-MCDF. Our results also show no antagonism of CYP1A1 induction with co-treatment of Hepa-1 cells with TCDD and 6-MCDF. These data suggest that 6-MCDF exhibits agonist activity with respect to induction of CYP1A1 in the Hepa-1 cell line.
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PMID:Lack of antagonism of 2,3,7,8-tetrachlorodibenzo-p-dioxin's (TCDDs) induction of cytochrome P4501A1 (CYP1A1) by the putative selective aryl hydrocarbon receptor modulator 6-alkyl-1,3,8-trichlorodibenzofuran (6-MCDF) in the mouse hepatoma cell line Hepa-1c1c7. 1553 86

We employed DNA microarray to identify unique hepatic gene expression patterns associated with subchronic exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and other halogenated aromatic hydrocarbons (HAHs). Female Harlan Sprague-Dawley rats were exposed for 13 weeks to toxicologically equivalent doses of four different HAHs based on the toxic equivalency factor of each chemical: TCDD (100 ng/kg/day), 2,3,4,7,8-pentachlorodibenzofuran (PeCDF; 200 ng/kg/day), 3,3',4,4',5-pentachlorobiphenyl (PCB126; 1,000 ng/kg/day), or 2,2',4,4',5,5'-hexachlorobiphenyl (PCB153; 1,000 microg/kg/day). Global gene expression profiles for each exposure, which account for 8,799 gene probe sets contained on Affymetrix RGU34A GeneChips, were compared by principal components analysis. The aryl hydrocarbon receptor (AhR) ligands TCDD, PeCDF, and PCB126 produced very similar global gene expression profiles that were unique from the nonAhR ligand PCB153, underscoring the extensive impact of AhR activation and/or the resulting hepatic injury on global gene expression in female rat liver. Many genes were co-expressed during the 13-week TCDD, PeCDF, or PCB126 exposures, including classical AhR-regulated genes and some genes not previously characterized as being AhR regulated, such as carcinoembryonic-cell adhesion molecule 4 (C-CAM4) and adenylate cyclase-associated protein 2 (CAP2). Real-time reverse-transcriptase polymerase chain reaction confirmed the increased expression of these genes in TCDD-, PeCDF-, and PCB126-exposed rats as well as the up- or down-regulation of several other novel dioxin-responsive genes. In summary, DNA microarray successfully identified dioxin-responsive genes expressed after exposure to AhR ligands (TCDD, PeCDF, PCB126) but not after exposure to the non-AhR ligand PCB153. Together, these findings may help to elucidate some of the fundamental features of dioxin toxicity and may further clarify the biologic role of the AhR signaling pathway.
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PMID:Subchronic exposure to TCDD, PeCDF, PCB126, and PCB153: effect on hepatic gene expression. 1559 15

In contrast to the beneficial effects of tert-butylhydroquinone (tBHQ) as a food antioxidant, a number of studies have shown that chronic exposure to tBHQ may induce carcinogenicity. Therefore, we examined the ability of tBHQ to induce the cytochrome P450 1a1 (Cyp1a1), an enzyme known to play an important role in the chemical activation of xenobiotics to carcinogenic derivatives. A significant concentration-dependent increase in Cyp1a1 mRNA, protein, and activity occurred after treatment of murine hepatoma Hepa 1c1c7 cells with tBHQ. The increase in mRNA was apparent 3 h after treatment. The RNA polymerase inhibitor, actinomycin D, completely blocked the Cyp1a1 induction by tBHQ, indicating a requirement of de novo RNA synthesis through transcriptional activation. The protein synthesis inhibitor cycloheximide superinduced the tBHQ-mediated induction of Cyp1a1 mRNA and completely prevented the increase in Cyp1a1 activity, indicating that the induction of enzyme activity by tBHQ is dependent on de novo protein synthesis. In addition, the aryl hydrocarbon receptor (AHR) antagonist, resveratrol, inhibited the increase in Cyp1a1 activity by tBHQ. Gel electrophoretic mobility shift assays showed that tBHQ causes activation or transformation of the AHR in nuclear extracts, indicating that AHR-dependent mechanisms contributed to the Cyp1a1 induction. Similar to murine Hepa 1c1c7 cells, tBHQ caused a concentration-dependent increase in CYP1A1 at the mRNA and activity levels in human HepG2 cells. This is the first demonstration that the phenolic antioxidant, tBHQ, can directly induce Cyp1a1 gene expression in an AHR-dependent manner and may represent a novel mechanism by which tBHQ promotes carcinogenicity.
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PMID:tert-Butylhydroquinone is a novel aryl hydrocarbon receptor ligand. 1560 32

Using chromatin immunoprecipitation assays, we studied the 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-mediated recruitment of the aryl hydrocarbon receptor (AhR) and several co-regulators to the CYP1A1 promoter. AhR displayed a time-dependent recruitment, reaching a peak at 75 min and maintaining promoter occupancy for the remainder of the time course. Recruitment of AhR was followed by TIF2/SRC2, which preceded CBP, histone H3 acetylation, and RNA polymerase II (RNAPII). Simultaneous recruitment to the enhancer and the TATA box region suggests the formation of a large multiprotein complex bridging the two promoter regions. Interestingly, estrogen receptor alpha (ERalpha) displayed a TCDD- and time-dependent recruitment to the CYP1A1 promoter, which was increased by co-treatment with estradiol. Transfection in HuH7 human liver cells confirmed previously reported ERalpha enhancement of AhR activity. In contrast, TCDD did not induce the recruitment of ERalpha to the estrogen-responsive pS2 promoter, and after 120 min of co-treatment with estradiol, ERalpha is still present on the CYP1A1 promoter but no longer at pS2. RNA interference studies with T47D cells support a role for ERalpha in TCDD-dependent CYP1A1 expression. Our data suggest that ERalpha acts as a coregulator of AhR-mediated transcriptional activation and that the recruitment of ERalpha by AhR represents a novel mechanism AhR-ERalpha cross talk.
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PMID:Aryl hydrocarbon receptor-mediated transcription: ligand-dependent recruitment of estrogen receptor alpha to 2,3,7,8-tetrachlorodibenzo-p-dioxin-responsive promoters. 1596 90

Transcriptional regulation of gene expression requires posttranslational modification of histone proteins, which, in concert with chromatin-remodeling factors, modulate chromatin structure. Exposure to environmental agents may interfere with specific histone modifications and derail normal patterns of gene expression. To test this hypothesis, we coexposed cells to binary mixtures of benzo[a]pyrene (B[a]P), an environmental procarcinogen that activates Cyp1a1 transcriptional responses mediated by the aryl hydrocarbon receptor (AHR), and chromium, a carcinogenic heavy metal that represses B[a]P-inducible AHR-mediated gene expression. We show that chromium cross-links histone deacetylase 1-DNA methyltransferase 1 (HDAC1-DNMT1) complexes to Cyp1a1 promoter chromatin and inhibits histone marks induced by AHR-mediated gene transactivation, including phosphorylation of histone H3 Ser-10, trimethylation of H3 Lys-4, and various acetylation marks in histones H3 and H4. These changes inhibit RNA polymerase II recruitment without affecting the kinetics of AHR DNA binding. HDAC1 and DNMT1 inhibitors or depletion of HDAC1 or DNMT1 with siRNAs blocks chromium-induced transcriptional repression by decreasing the interaction of these proteins with the Cyp1a1 promoter and allowing histone acetylation to proceed. By inhibiting Cyp1a1 expression, chromium stimulates the formation of B[a]P DNA adducts. Epigenetic modification of gene expression patterns may be a key element of the developmental and carcinogenic outcomes of exposure to chromium and to other environmental agents.
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PMID:Chromium cross-links histone deacetylase 1-DNA methyltransferase 1 complexes to chromatin, inhibiting histone-remodeling marks critical for transcriptional activation. 1768 57

Cytochrome P450 (CYP) 1A1 and CYP1B1 are inducible by 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin) in the human breast cancer cell line, MCF-7. Since CYP1A1 was inducible to a much greater degree than CYP1B1, we hypothesized that there may be differences in coactivator recruitment to the promoter and/or enhancer regions of these genes. Dioxin treatment leads to recruitment of the aryl hydrocarbon receptor to the enhancer regions but not to the proximal promoter regions of both the CYP1A1 and CYP1B1 genes. On the other hand, dioxin treatment facilitated recruitment of RNA polymerase II to the promoters but not the enhancer regions. Dioxin treatment also elicited recruitment of the transcriptional coactivators, steroid receptor coactivator 1 (SRC-1) and steroid receptor coactivator 2 (SRC-2) and p300, which possess intrinsic histone acetyltranferase activities, to both genes, whereas Brahma (BRM)/Switch 2-related gene 1 (BRG-1), a subunit of nucleosomal remodeling factors, was recruited more robustly to CYP1A1 relative to CYP1B1. Small inhibitory RNA-mediated knockdown of p300 and SRC-2 adversely affected dioxin induction of both genes, whereas knockdown of BRM/BRG-1 reduced CYP1A1 induction but had little, if any, effect on CYP1B1 induction. These results suggest that nucleosomal remodeling is less significant for dioxin-mediated induction of CYP1B1 than that of CYP1A1 and may be related to the more modest inducibility of the former. Interestingly, simultaneous knockdown of SRC-2 and BRM/BRG-1 had no greater effect on CYP1A1 induction than knockdown of each coactivator individually, while simultaneous knockdown of p300 and BRM/BRG-1 had a much greater effect than knockdown of each individual gene, suggesting that the recruitment of SRC-2 to CYP1A1 depends upon BRM/BRG-1, while the recruitments of p300 and BRM/BRG-1 are independent of each other. These observations provide novel insights into the functional roles of the endogenous coactivators in dioxin induction of the human CYP1A1 and CYP1B1 genes in their natural chromosomal configurations.
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PMID:Roles of coactivator proteins in dioxin induction of CYP1A1 and CYP1B1 in human breast cancer cells. 1884 20

The CYP1A family of cytochrome P450s (CYPs), comprising CYP1A1, CYP1A2, and CYP1B1, plays a role in bioactivation of several procarcinogens to carcinogenic derivatives, and also in detoxification of several xenobiotic compounds. Resveratrol (3,4,5-trihydroxystelbine) is a naturally occurring compound that has been shown in a number of studies to inhibit the induction of CYP1A1 and CYP1B1 by dioxin (2,3,7,8-tetrachloro-dibenzo-p-dioxin), but the mechanism(s) of resveratrol inhibition is controversial. In the current study, 100nM dioxin treatment for 24, 48, and 72 h induced CYP1A1, CYP1A2, and CYP1B1 mRNA levels in the human breast cancer cell line MCF-7, and CYP1A1 and CYP1A2 mRNA levels in the human hepatocellular carcinoma cell line, HepG2. Simultaneous treatment with 10 microM resveratrol significantly inhibited dioxin-induced mRNA expression levels of these genes in both cell lines. Our studies are novel in that we used the chromatin immunoprecipitation assay to assay dioxin-induced recruitment of the aryl hydrocarbon receptor (AHR), and aryl hydrocarbon nuclear translocator (ARNT) to the enhancer regions and recruitment of RNA polymerase II to the promoter regions, of the CYP1A1 and CYP1B1 genes in their natural chromosomal settings. These recruitments were significantly inhibited in cells cotreated with resveratrol. Our studies thus indicate that resveratrol inhibits dioxin induction of the CYP1 family members either by directly or indirectly inhibiting the recruitment of the transcription factors AHR and ARNT to the xenobiotic response elements of the corresponding genes. The reduced transcriptional factor binding at their enhancers then results in reduced pol II recruitment at the promoters of these genes.
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PMID:Resveratrol inhibits dioxin-induced expression of human CYP1A1 and CYP1B1 by inhibiting recruitment of the aryl hydrocarbon receptor complex and RNA polymerase II to the regulatory regions of the corresponding genes. 1937 45


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