Estrogen (E), acting through the estrogen receptor (ER), plays a critical role in gene expression in target tissues such as breast, bone, and the pituitary, where it controls hormone production and cell proliferation. Cell-specific responses to E are well-documented, and mechanisms include differential expression of ER isoforms, distinct pathways of ligand- independent ER stimulation, and differential expression of ER coactivator and repressor proteins. We previously identified a pituitary-specific truncated ERalpha in rat pituitary cells (TERP). TERP is transcribed from a unique promoter and dramatically regulated by E, divergent from full-length ER, such that the TERP:ER ratio varies from 0:1 to 3-4:1. We cloned the intronic TERP promoter for the rat and mouse ERalpha genes, and found it is stimulated by E in transfection assays. TERP cannot bind DNA, but modulates ER transcriptional activity biphasic manner, dependent on TERP:ER ratios. When TERP:ER ratios are high (greater than 1:1) ER activity is suppressed by the formation of TERP:ER heterodimers that cannot bind DNA. At low ratios TERP stimulates ER activity, and we hypothesize this occurs by titration of repressor proteins. One such protein is Repressor of Estrogen Receptor Activity (REA), expressed in pituitary cells, and other repressors may be identified. We will determine the biological role of TERP and its physiological regulation in four specific aims. 1) Interactions of TERP with REA and other ER repressors will be evaluated at the level of protein-protein interactions, the ability of TERP to compete with ER binding to repressors, and to rescue suppressed ER transcription. 2) Expression of the mouse TERP promoter will be analyzed in transient transfection assays for basal enhancer and E-regulated regions. Proteins binding to these sites will be identified, and the ability of TERP to autoregulate its own promoter will be determined. 3) Expression of TERP mRNA during development in male and female mice will be evaluated. 4) Transgenic mice in which TERP expression is eliminated by selective TERP promoter disruption will be produced, and physiological responses of lactotropes and gonadotropes to E will be characterized. Cell-specific ER isoforms with varying biological activities has profound physiological implications for cell function and growth. These studies will provide important information on mechanisms for positive and negative E feedback at the cellular level.
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