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Antiestrogen-liganded estrogen receptor interaction with estrogen responsive element DNA in vitro
C M Klinge1, R A Bambara, R Hilf
1Department of Biochemistry, University of Rochester Cancer Center, NY.
The Journal of Steroid Biochemistry and Molecular Biology
|October 1, 1992
Summary
Antiestrogens like 4-hydroxytamoxifen alter estrogen receptor (ER) binding to DNA. While 4-OHT-ER shows high affinity, it has reduced cooperativity compared to estradiol-ER, impacting gene transcription.
Area of Science:
- Molecular Endocrinology
- Genetics
- Biochemistry
Background:
- Antiestrogens modulate estrogen receptor (ER) function in gene transcription.
- The precise mechanisms by which antiestrogens affect ER binding to DNA remain incompletely understood.
Purpose of the Study:
- To quantify the binding affinity and cooperative binding of ER to estrogen-responsive elements (EREs) when complexed with different ligands.
- To elucidate how antiestrogenic compounds influence ER-DNA interactions.
Main Methods:
- Utilized a microtiter well plate assay to measure the binding of liganded ER to DNA containing EREs.
- Quantified binding affinities (Kd) and cooperativity using Scatchard plots and Hill coefficients for estradiol-ER, 4-hydroxytamoxifen-ER, and ICI 164,384-ER.
Main Results:
- Estradiol-ER and 4-hydroxytamoxifen-ER exhibited high affinity for a single ERE, while ICI 164,384-ER did not bind.
- 4-hydroxytamoxifen-ER showed reduced binding capacity and cooperativity compared to estradiol-ER at saturation.
- Cooperative binding was observed for estradiol-ER but was diminished for 4-hydroxytamoxifen-ER and absent for ICI 164,384-ER, particularly with multiple ERE copies.
Conclusions:
- The conformation of ER induced by 4-hydroxytamoxifen may decrease its DNA-binding capacity to EREs without affecting binding affinity.
- Distinct higher-order protein-protein interactions occur between antiestrogen-liganded ER and DNA compared to estradiol-liganded ER.