1. We created fusion chimeras between GR and other nuclear receptors that manifest ligand dependent cytoplasmic/nuclear translocation for the chimeric receptor in response to the heterologous ligand. The approach has been successfully developed for the glucocorticoid receptor (GR), the retinoic acid receptor (RAR), the estrogen receptor (ER), the aryl hydrocarbon receptor (AhR), the progesterone receptor (PR), and the thyroid hormone receptor (ThR). We developed cell lines which express fluorescently tagged versions of the receptor chimeras. We have utilized this fluorescent protein fusion technology to show that U.S. water supplies are heavily contaminated with chemicals that have glucocorticoid and androgen activity. US patent No. 20150168416 was granted covering licensing of this technology. We have collaborated with multiple intramural and extramural investigators to track hormone contamination in U.S. waterways, and explore the implication of these findings for human health. 2. Transcription factors dynamically bind to chromatin and are essential for the regulation of genes. While a large percentage of these proteins appear to self-associate to form dimers or higher-order oligomers, the stoichiometry of DNAbound transcription factors has been poorly characterized in vivo. The glucocorticoid receptor (GR) is a ligand regulated transcription factor and its oligomerization state has been implicated in clinical outcome of therapy. Although widely believed to act as a dimer, we have shown that GR is predominantly a tetramer when bound to its target DNA. Using a unique set of imaging techniques coupled with a cell line containing an array of DNA binding elements, we found that DNA binding triggers an inter-domain allosteric regulation of the GR, followed by a change in its oligomeric state. These findings demonstrate a new step in steroid receptor activation an open new doors to the rational design of novel GR ligands. We discovered that a mutation in the GR DBD causes the receptor to constitutively adopt a tetrameric form of the receptor. When expressed in GR knock out cells, all nuclear receptor is in the tetrameric state. We have begun an exploration of the genome wide behavior for this receptor state.
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