Development of a simple screening system for endocrine disruptors.
BACKGROUND
An endocrine disruptor is a synthetic chemical, which causes adverse effects in an organism, or its progeny, after causing perturbations in the endocrine system. It is important to know which synthetic chemicals have endocrine-disrupting action. However, an increasing number of synthetic chemicals are being produced by modern synthetic chemistry, and the examination of endocrine disruptor potential has not yet caught up with the advances in synthetic chemistry. In this study, we have developed such a screening system for detecting synthetic chemicals with estrogen-like effects.


METHODS
The system was based on the yeast one-hybrid system. Both HIS3 and lacZ reporter genes connected to three tandem copies of the estrogen response element were prepared. Gal4-estrogen receptor is a fusion protein made from the activation domain (AD) of the yeast GAL4 transactivator gene and then incorporated into a plasmid, which was transfected into the YM4271 yeast cell strain. The estrogen effect was judged by this developed screening system.


RESULTS
A dual reporter assay-system was established by transfection of the both HIS3 and lacZ reporter genes into the yeast cells. This screening system enabled the detection of as little as 10-12 mol of beta-estradiol.


CONCLUSIONS
These results show that this newly developed dual assay is useful for the screening of endocrine-disruptors that have estrogen-like action.
Sugawara
Teruo
T
Department of Biochemistry, Hokkaido University School of Medicine, Sapporo, Hokkaido, Japan.
Nakajima
Ayako
A
Nomura
Eiji
E
D004305:Dose-Response Relationship, Drug; D004703:Endocrine System / Q000473:pathology*; D004958:Estradiol / Q000494:pharmacology; D004967:Estrogens / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry; D017930:Genes, Reporter; D006801:Humans; D010957:Plasmids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D020218:Response Elements; D014157:Transcription Factors / Q000737:chemistry; D015533:Transcriptional Activation; D014162:Transfection; D020798:Two-Hybrid System Techniques; D001616:beta-Galactosidase / Q000378:metabolism
D004967:Estrogens; D005656:Fungal Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D004958:Estradiol; D001616:beta-Galactosidase