Ah Receptor-Based Chemical Screening Bioassays: Application and Limitations for the Detection of Ah Receptor Agonists

Seidel, Shawn D.; Li, Violet; Winter, G.M.; Rogers, William J.; Martinez, Eugenio I.; Denison, Michael S.

doi:10.1093/toxsci/55.1.107

Cited by 99 publications

(53 citation statements)

References 39 publications

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“…Greater fold induction was observed with cells expressing zfAHR2 (4.4-to 10-fold) when using a luciferase reporter driven by the rainbow trout CYP1A promoter prt1Aluc [described previously in Abnet et al (1999a)] (data not shown). In an effort to screen other potential ligands for zfAHR1, the assay was repeated with maximal-effect concentrations of other known inducers of the AHR pathway (BNF, BaP, DMBA, HxCDD, I3C, 3MC, TCDF, TBDD, and I3AA) (Postlind et al, 1993;Heath-Pagliuso et al, 1998;Abnet et al, 1999b;Seidel et al, 2000;Stephensen et al, 2000;Henry et al, 2001). Luciferase activity was elevated in cells expressing zfAHR2 and exposed to TCDD, BaP, DMBA, HxCDD, 3MC, I3AA, TCDF, TBDD, and I3AA (Fig.…”

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confidence: 99%

The Zebrafish (Danio rerio) Aryl Hydrocarbon Receptor Type 1 Is a Novel Vertebrate Receptor

et al. 2002

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Fish are known to have two distinct classes of aryl hydrocarbon receptors, and their roles in mediating xenobiotic toxicity remain unclear. In this study, we have identified and characterized a cDNA tentatively named zebrafish AHR1 (zfAHR1). Analysis of the deduced amino acid sequence reveals that the protein is distinct from zfAHR2 and is more closely related to the mammalian aryl hydrocarbon receptor (AHR). zfAHR1 and zfAHR2 share 40% amino acid identity overall and 58% in the N-terminal half. The zfAHR1 gene maps to linkage group 16 in a region that shares conserved synteny with human chromosome 7 containing the human AHR, suggesting that the zfAHR1 is the ortholog of the human AHR. zfAHR2 maps to a separate linkage group (LG22). Both zfAHR mRNAs are expressed in early development, but they are differentially expressed in adult tissues. zfAHR2 can dimerize with zfARNT2b and binds with specificity to dioxin-responsive elements (DREs). Under identical conditions, zfAHR1/zfARNT2b/DRE complexes are formed; however, the interactions are considerably weaker. In COS-7 cells expressing zfARNT2b and zfAHR2, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure leads to a significant induction of dioxin-responsive reporter genes. In identical experiments, TCDD exposure fails to induce the reporter gene in zfAHR1-expressing cells. Ligand-binding experiments suggested that the differential zfAHR activities are attributable to differences in TCDD binding because only zfAHR2 exhibits high-affinity binding to [ 3 H]TCDD or ␤-naphthoflavone. Finally, using chimeric zfAHR1/zfAHR2 constructs, the lack of TCDDmediated transcriptional activity was localized to the ligandbinding and C-terminal domains of zfAHR1.The aryl hydrocarbon receptor (AHR) is a member of the basic helix-loop-helix PAS family of proteins. Members of this family include the aryl hydrocarbon receptor nuclear translocators (ARNT, ARNT2, and ARNT3), hypoxia-inducible factor 1␣, Drosophila melanogaster single-minded, D. melanogaster period, Clock, and others. These proteins are involved in mediating responses to environmental contaminants, low oxygen tension and glucose, circadian rhythm, and various other cues (for review, see Gu et al., 2000). The basic components of the AHR signal transduction pathway are well understood (for review, see Schmidt and Bradfield, 1996). The cytosolic AHR is complexed with at least three chaperone proteins, two molecules of HSP90 and the aryl hydrocarbon interacting protein (AIP, also known as ARA9 or XAP2), enabling proper conformation for ligand binding. Once bound by ligands such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), the cytosolic AHR translocates to the nucleus, dissociates from the chaperone proteins, and dimerizes with ARNT. This heterodimeric AHR/ARNT complex associates with specific DNA sequences termed dioxin-response elements (DRE, alternatively known as xenobiotic response el-

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confidence: 99%

The Zebrafish (Danio rerio) Aryl Hydrocarbon Receptor Type 1 Is a Novel Vertebrate Receptor

et al. 2002

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“…We also found differences in the relatively efficacy of diuron to activate AhR-dependent gene expression in guinea pig cells as compared to its ability to bind to and stimulate transformation and DNA binding of the guinea pig AhR in vitro. This is not surprising, given that we have previously found most AhR agonists to be significantly more potent in in vitro, cell-free AhR assays when compared to cell-based induction assays [15,16]. In cell-free AhR assays (ligand and DNA binding), the agonist has direct access to the AhR in the incubation condition and if it can bind, it will, resulting in a positive response.…”

Section: Discussionmentioning

confidence: 87%

Interaction of diuron and related substituted phenylureas with the Ah receptor pathway

Zhao

Baston

Hammock

et al. 2006

J Biochem & Molecular Tox

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The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that mediates many of the biological and toxicological actions of structurally diverse chemicals, including the ubiquitous environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin. Here, we have examined the ability of diuron, a widely used herbicide, and several structurally related substituted phenylureas to bind to and activate/inhibit the AhR and AhR signal transduction. Diuron induced CYP1A1 mRNA levels in mouse hepatoma (Hepa1c1c7) cells and AhR-dependent luciferase reporter gene expression in stably transfected mouse, rat, guinea pig, and human cell lines. In addition, ligand binding and gel retardation analysis demonstrated the ability of diuron to competitively bind to and stimulate AhR transformation and DNA binding in vitro and in intact cells. Several structurally related substituted phenylureas competitively bound to the guinea pig hepatic cytosolic AhR, inhibited 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced AhR-dependent luciferase reporter gene expression in a species-specific manner and stimulated AhR transformation and DNA binding, consistent with their role as partial AhR agonists. These results demonstrate not only that diuron and related substituted phenylureas are AhR ligands but also that exposure to these chemicals could induce/inhibit AhR-dependent biological effects.

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“…Bioassays results are very sensitive to the procedures used to cleanup and extract samples (Seidel et al, 2000;Windal et al, 2005). Bioassays that are marketed commercially for the measurement of PCDD/Fs and PCBs in foods, soils, or tissues typically utilize sample cleanup procedures that include an activated-carbon column (e.g., the XDS ''XCARB'' affinity matrix) (Brown et al, 2001) in series with an acid-silica column.…”

Section: Discussionmentioning

confidence: 99%

AH receptor agonist activity in human blood measured with a cell-based bioassay: Evidence for naturally occurring AH receptor ligands in vivo

Connor¹,

Harris²,

Edwards

et al. 2007

J Expo Sci Environ Epidemiol

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In the present study, an aryl hydrocarbon receptor (AHR)-driven reporter gene bioassay was used to measure the activity, measured as an induction equivalent (IEQ) as compared to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), or IEQ concentration in human blood samples from 10 volunteers under different dietary regimens. Blood concentrations of polychlorinated dibenzo-p-dioxins/furans (PCDD/Fs) and polychlorinated biphenyls (PCBs), as determined by analytical chemistry (HR-GC/MS), and expressed as toxic equivalents (TEQs) with the use of TCDD equivalency factors (TEFs), were within a range that has been reported in the general US population, ranging from 0.022 to 0.119 ppt (whole blood basis). However, the human blood IEQ measured directly via bioassay ranged from 13.4 to 218 ppt (whole blood basis). These order of magnitude greater IEQs compared to the TEQs for dioxins, furans, and certain PCBs suggests that human blood contains a relatively high level of AHR agonists able to activate the CYP1A1 dioxin response element (DRE)-linked reporter gene bioassay and that this AHR activity is not accounted for by PCDDs/Fs and dioxin-like PCBs based on standard HR-GC/MS and TEF analysis. When study participants switched from a ''baseline'' to a high-vegetable diet, increases in bioassay IEQ were observed that were statistically significant (Po0.05). In addition, IEQ activity was elevated above levels observed following dietary intervention in two subjects given indole-3-carbinol (I3C) supplements. We conclude that a substantial portion of the IEQ activity occurred as a result of the increased intake of natural AHR agonists (NAHRAs) present in many fruits, vegetables. and herbs. Our findings also suggest that dietary NAHRAs constitute a substantial daily dietary intake of AHR-active compounds, and these NAHRAs could influence AHR status in humans and play a role in a basal level of AHR activation.

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Ah Receptor-Based Chemical Screening Bioassays: Application and Limitations for the Detection of Ah Receptor Agonists

Cited by 99 publications

References 39 publications

The Zebrafish (Danio rerio) Aryl Hydrocarbon Receptor Type 1 Is a Novel Vertebrate Receptor

The Zebrafish (Danio rerio) Aryl Hydrocarbon Receptor Type 1 Is a Novel Vertebrate Receptor

Interaction of diuron and related substituted phenylureas with the Ah receptor pathway

AH receptor agonist activity in human blood measured with a cell-based bioassay: Evidence for naturally occurring AH receptor ligands in vivo

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