Abstract Dr. B. Gehm
Title of Project Signaling by Non-Classical Ligands of Estrogen Receptor: Novel Approaches to Detection and Mechanism
Abstract Estrogens affect a large number of physiological functions and disease states. In addition to endogenous hormones, humans are exposed to a wide variety of estrogenic compounds in the diet, environment, and pharmaceuticals. These non-classical ligands for the estrogen receptor (ER) can have major effects on human health. Efficient methods of testing for estrogenic activity are important for drug development, environmental testing, and research on receptor functions and mechanisms. Yeast-based assays using exogenously expressed ERs and reporter genes are inexpensive and widely used, especially for environmental testing, but have the defect that the response to some SERMS (selective estrogen receptor modulators) and phytoestrogens is very different from that seen in mammalian cells. In Aim 1, we will develop and test a reporter system in the nematode C. elegans, based on the hypothesis that the closer evolutionary relationship of humans and C. elegans will enable this system to produce more relevant results than yeast-based assays but retain the advantages of simplicity and low cost. Although there is substantial human exposure to non-classical ER ligands, their modes of action are incompletely understood. SERMs, such as tamoxifen, often have biphasic dose-response curves, with ER agonism at low concentrations and antagonism at high concentrations. It has been proposed that, in addition to occupying the hormone-binding pocket of the ER, these compounds bind (with lower affinity) to a 2nd site that mediates antagonism. Some phytoestrogens have a similar biphasic effect, suggesting that they are natural ligands for this site. In Aim 2, we will determine if phytoestrogens and ANGELS (activators of non-genotropic estrogen-like signaling, a new class of ER-targeted drugs) compete with SERMS for binding to the 2nd site; identify and ablate the site via targeted mutagenesis; and characterize the significance of the site in ER function by reporter gene assays, microarray analysis, and real-time PCR.
Updated 07/31/2006
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