The goal of this proposal is to characterize the Ah receptor - a soluble protein present in many tissues of vertebrate species and a presumed member of the erbA superfamily of receptors, which mediates the pleiotropic effects of halogenated aromatic hydrocarbons such as 2,3,7,8- tetrachlorodibenzo-p-dioxin. 1) We propose to clone and sequence the gene for the Ah receptor from cDNA libraries by using degenerate oligonucleotide probes and rabbit polyclonal antibodies to a synthetic peptide that corresponds to the N-terminal sequence of the receptor. We plan to analyze the structure-function analysis of this protein using mutagenesis and an expression assay. 2) We propose to characterize the four allelic variants of the Ah receptor among inbred mouse strains: a) in vitro by their binding kinetics K-D, B-MAX and ease of activation and b) in vivo by genetic analysis of the differences in functional phenotypes especially Ah(b-1)/Ah(d) vs. Ah(b-2)/Ah(d) and correlation of this with quantitative nuclear extract-gel retardation experiments. 3) We propose to expand on our photoaffinity labeling and immunologic methodology; a) to screen lower vertebrate and invertebrate species for the presence of the Ah receptor, b) to prepare new monoclonal and polyclonal antibodies to various regions of the Ah receptor, c) to prepare an immunoaffinity column to aid in rapid purification, and d) to examine murine tissues for the cellular localization of the Ah receptor using immunohistochemistry.

National Institute of Health (NIH)
National Institute of Environmental Health Sciences (NIEHS)
Method to Extend Research in Time (MERIT) Award (R37)
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Chemical Pathology Study Section (CPA)
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University of Wisconsin Madison
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United States
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Poland, A; Palen, D; Glover, E (1994) Analysis of the four alleles of the murine aryl hydrocarbon receptor. Mol Pharmacol 46:915-21
Pollenz, R S; Sattler, C A; Poland, A (1994) The aryl hydrocarbon receptor and aryl hydrocarbon receptor nuclear translocator protein show distinct subcellular localizations in Hepa 1c1c7 cells by immunofluorescence microscopy. Mol Pharmacol 45:428-38
Burbach, K M; Poland, A; Bradfield, C A (1992) Cloning of the Ah-receptor cDNA reveals a distinctive ligand-activated transcription factor. Proc Natl Acad Sci U S A 89:8185-9
Poland, A; Glover, E; Bradfield, C A (1991) Characterization of polyclonal antibodies to the Ah receptor prepared by immunization with a synthetic peptide hapten. Mol Pharmacol 39:20-6
Poland, A; Glover, E (1990) Characterization and strain distribution pattern of the murine Ah receptor specified by the Ahd and Ahb-3 alleles. Mol Pharmacol 38:306-12
Poland, A; Teitelbaum, P; Glover, E (1989) [125I]2-iodo-3,7,8-trichlorodibenzo-p-dioxin-binding species in mouse liver induced by agonists for the Ah receptor: characterization and identification. Mol Pharmacol 36:113-20
Poland, A; Teitelbaum, P; Glover, E et al. (1989) Stimulation of in vivo hepatic uptake and in vitro hepatic binding of [125I]2-lodo-3,7,8-trichlorodibenzo-p-dioxin by the administration of agonist for the Ah receptor. Mol Pharmacol 36:121-7
Bradfield, C A; Poland, A (1988) A competitive binding assay for 2,3,7,8-tetrachlorodibenzo-p-dioxin and related ligands of the Ah receptor. Mol Pharmacol 34:682-8
Perdew, G H; Poland, A (1988) Purification of the Ah receptor from C57BL/6J mouse liver. J Biol Chem 263:9848-52
Poland, A; Glover, E (1988) Ca2+-dependent proteolysis of the Ah receptor. Arch Biochem Biophys 261:103-11

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