Agonist AMD Antagonist Activity in a GFP Yeast Based Estrogen Receptor Functional Assay

Estrogen receptors (ERα and ER β) are ligand-binding transcription factors activated by the hormone 17-β estradiol. Ligand binding triggers ER dimerization, translocation of the receptor from the cytoplasm into the nucleus and eventually activation of the genes under control of ER. Studies have revealed a role for estrogen receptors in male and female sexual development, reproductive functions, bone metabolism and regulation of neuroendocrine and cardiovascular systems. ER is also known to bind to other non-native ligands known in pharmacology as receptor agonists or antagonists. Agonists provoke a biological response when bound to the receptor; antagonists inhibit a biological response when bound. Our lab is interested in the promiscuous binding of the estrogen receptor and its ability to activate different genes in different tissues. Fluorescence Resonance Energy Transfer (FRET) assays have previously been performed using ER to study ligand binding affinities for the receptor. However, this technique is unable to determine whether these ligands are agonists or antagonists and allow ER dimerization and gene activation. To investigate these phenomena, an activity assay that measures ER controlled gene expression has been developed which provides the opportunity to gain further insight into the functional activity in living systems. Recombinant yeast cells that express ERα use the green fluorescent protein (GFP) reporter to determine whether ER α, in the presence of a particular ligand, has activated gene of expression. We have correlated the binding to agonist and antagonist behavior of several xeno and phyto estrgens.