Approximately 270,000 women will be diagnosed with breast cancer this year and about 40,000 women will die of the disease, second in mortality only to lung cancer. Steroid hormones such as the female sex steroids, estrogen and progesterone, play critical roles in normal development and homeostasis of many organs, particularly breast and uterus and cancers of these tissues. Our understanding of the signaling mechanisms used by steroids has grown remarkably. Nonetheless, investigation of even the """"""""classic"""""""" mode of action of steroid receptors continues to reveal surprising complexity, particularly in the crosstalk between different hormones. Although the estrogen receptor and progesterone receptor (PR) are thought to bind separate and distinct sets of target sites in the DNA, we have made the surprising observation that PR binds to estrogen response elements (EREs) both in vitro and in vivo. In doing so, PR inhibits estrogen-mediated gene induction at a subset of estrogen targets including pS2, amphiregulin, C3 and TERT. This inhibition does not appear to be primarily due to competition at the ERE but is, instead, because the hormone-PR complex differs functionally when bound at an ERE instead of a PRE. Thus, we propose that DNA is a functional ligand for PR. As a corollary, we propose that the ability of the PR to inhibit estrogen-mediated gene induction is due to the recruitment of the corepressors NCoR and/or SMRT to the gene by PR.
Four aims are proposed to test these hypotheses.
Aim one will systematically investigate the influence of target sequence and hormonal ligands on the function of PR using a combination of chromatin immunoprecipi-tation and reporter gene assays.
Aim two tests the prediction that PR bound to an ERE is conformationally distinct via a combination of proteolytic assays and by x-ray crystallography.
In aim three the global ramifications of these data will be explored. Microarray analyses will be used to define the set of estrogen-inducible genes inhibited by liganded PR and to select candidates for further mechanistic analysis.
Aim four will specifically test whether the inhibition of estrogen-mediated induction by progesterone is through the recruitment of NCoR, SMRT, or associated HDACs by PR. In this application we offer a novel view of progesterone receptor action and propose experiments to define both mechanisms and the broader consequences of these mechanisms on the interactions of progestins and estrogens.
