Development of method for assessing species-specific risk of Ah receptor-mediated dioxin toxicity

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We identified two distinct AhR cDNAs (AhR1 and AhR2) from the livers of black-footed albatross (Diomedea nigripes) and common cormorant (Phalacrocorax carbo). The objective of this study is to investigate the functional characterization of the AhRs and their expression levels related to planar halogenated aromatic hydrocarbons (PHAHs) exposure and CYP1A expression in aquatic birds. In velocity sedimentation analysis using AhR proteins expressed by in vitro transcription and translation, both AhRs exhibited specific binding to [^3H]TCDD. Focusing on the amino acid residues corresponding to Ile^<325> and Ser^<381> in chicken AhR1, which may contribute to the differential TCDD-binding affinity, the amino acid residues in albatross and cormorant AhR1 were Ile-Ala and Val-Ala, respectively. The [^3H]TCDD binding affinity measured was in the order of chicken > albatross > cormorant, and agreed with the order that may be expected from the amino acids. In contrast, the binding affinity of avia … More n AhR2 appeared to be unrelated to the corresponding amino acids.To investigate the molecular mechanism of TCDD toxicity in a marine fish species, red seabream (Pagrus major), we identified two distinct AHR isoform cDNAs (rsAHR1 and rsAHR2), which shared only 32% identity in full-length amino acid sequence. Quantitative analyses of both rsAHR mRNAs revealed that their tissue expression profiles were isoform-specific in different tissues of adult fishes ; rsAHR1 mRNA expressed primarily in brain, heart and ovary, while rsAHR2 mRNA was observed in all tissues examined. The expression of rsAHR1,rsAHR2 and CYP1A mRNAs were also determined in different developmental stages treated either with TCDD or solvent control at 10 hpf (hours post fertilization). Both rsAHR transcript levels increased prior to the stage at which heart beat and blood circulation were observable, but expression patterns were slightly different ; maximal expression of rsAHR1 and rsAHR2 mRNA were 96 hpf and 190 hpf, respectively. The rsAHR2 mRNA expression increased following TCDD exposure compared with solvent control, while TCDD did not affect the rsAHR1 mRNA level. Although basal expression of CYP1A mRNA was detectable, TCDD exposure further enhanced CYP1A mRNA level. The maximal expression of CYPIA mRNA was recorded before the onset of the first signs of TCDD toxicities such as yolk sac edema. Less