Detection of the Ah receptor in rainbow trout: use of 2-azido-3-[125I]iodo-7,8-dibromodibenzo-p-dioxin in cell culture

Area of Science:

• Environmental Toxicology
• Molecular Biology
• Aquatic Toxicology

Background:

• The aryl hydrocarbon receptor (AhR) is a crucial mediator of toxic responses to environmental contaminants.
• Understanding AhR characteristics in non-mammalian species is vital for assessing ecological risks.
• Rainbow trout (Oncorhynchus mykiss) are a model organism for aquatic toxicology studies.

Purpose of the Study:

• To detect and characterize the AhR in rainbow trout embryonic gonad (RTG-2) cells.
• To determine the molecular weight and sedimentation properties of the fish AhR.
• To compare the ligand-binding affinity of the fish AhR with that of mammalian sources.

Main Methods:

• Photoaffinity labeling of RTG-2 cells with [125I]2-azido-3-iodo-7,8-dibromodibenzo-p-dioxin.
• Preparation of cytosolic and nuclear extracts.
• Analysis using SDS-PAGE, sucrose density centrifugation, and competitive ligand binding assays.

Main Results:

• The AhR was successfully detected and photoaffinity labeled in RTG-2 cells.
• The cytosolic AhR sedimented at ~9.8S, and the nuclear form at ~7.5S.
• The molecular weight was determined to be 145 kDa, larger than mammalian AhRs, with similar ligand affinity rankings.

Conclusions:

• The study provides the first characterization of the AhR in rainbow trout cells.
• The fish AhR exhibits distinct molecular properties compared to mammalian counterparts.
• These findings contribute to understanding the toxicological mechanisms of environmental pollutants in fish.