In vitro experiments have demonstrated that members of the steroid receptor coactivator (SRC) family strongly and directly coactivate members of the nuclear receptor (NR) superfamily of ligand-inducible transcription factors. More recently, our in vivo studies have shown that disruption of individual SRC genes in mice results in differential phenotypes, suggesting that issue-specific combinatorial assembly of individual coactivators might support distinct patterns of NR activation. Since NRs are also distributed in a tissue-specific manner and appear to have differential affinities for coactivators, we hypothesize that individual NRs will demonstrate preferences for individual SRC family members. To test our hypothesis, we plan in this Project to determine the in vivo individual and combinatorial contributions of SRC family members to NR transactivation of target genes, under the stimulation of hormones and a novel class of pharmaceutical agents, the selective estrogen receptor modulators (SERMs). Firstly we will generate, in mice, transgenic ligand response systems for individual designer modifications of the estrogen receptor alpha (ERalpha), ERbeta, progesterone receptor, and androgen receptor. These chimeric receptors will be constructed in such a way that they activate only incorporated reporter genes. Furthermore, the receptors will be targeted to the natural hormone responsive cells of the mouse by using large genomic bacterial artificial chromosome DNA constructs bearing regulatory elements and entire chimeric NR genes. This system will allow us to evaluate the in vivo transcriptional activation profile for a given receptor in hormone target tissues of the mammary gland, uterus and prostate. Secondly, these transgenic mice will be crossed with individual and combinatorial SRC null mutant mice, enabling us to quantitate the in vivo effect of each coactivator, alone or in combination, on a specific receptor's transcriptional response to hormone. Finally, we will use these animal models to dissect the effects of SRC family members on the pharmacology of SERMs and to address the question of whether the tissue-selective activity of SERMs is due to the existence of different combinations or amounts of SRC family members in these tissues.
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