HORMONAL REGULATION OF ANDROGEN RECEPTOR EXPRESSION

雄激素受体(AR)是一种配体激活的转录因子，与雄激素调节基因相关的雄激素反应元件(ARs)结合。AR对雄激素介导的男性性发育是必不可少的，并在前列腺癌的病因和进展中发挥作用。雄激素促进组织特异性上调-AR mRNA下调，提示AR和细胞特异性因子在AR mRNA自我调节中发挥作用。这项建议的主要目标是确定这些非AR因素，并确定它们对AR mRNA上调的贡献。由于雄激素的反应性与AR水平有关，AR mRNA的自动调节可能在调节正常和肿瘤细胞的激素敏感性方面起关键作用。与这一观点一致，我们观察到雄激素介导的AR mRNA和蛋白的上调与细胞内雄激素敏感性的增强有关。AR基因的启动子和上游区域尚未被证明含有AR基因内含的6.5kb雄激素反应区外显子中的ARE和ARE-2。这些RES位于RcDNA350bp的片段上，该片段由外显子D和E组成。在报告基因分析中，6.5kb和350bp的片段都显示出细胞特异性的雄激素调节，反映了天然AR基因的上调。所有四个ARE都是AR与350bp区域结合和AR mRNA转录上调所必需的。此外，与天然基因一样，尽管存在Ares，但6.5kb和350bp的区域不受糖皮质激素和糖皮质激素受体(GR)的调节。与激素反应元件(HRE)的共同序列相似。HRE在其他几个基因上受GR的结合和调节。该方案的目的是利用体内足迹技术来检测AR、GR和非受体因子与6.5kb雄激素反应区域的外显子D和E的结合，这些外显子作为稳定整合的模板存在。我们将确定在雄激素反应区活跃和不活跃的细胞中，受体和核因子结合的模式是否因雄激素(和糖皮质激素)的反应而不同。我们将鉴定和鉴定差异结合的非受体因子(S)，并确定该因子(S)在细胞和雄激素特异性AR基因表达上调中的作用。这些研究的长期目标是提供一个全面的物理和功能特征的R-mRNA自我调节机制，并将这些机制与正常细胞和肿瘤细胞的激素反应联系起来。

The androgen receptor (AR) is a ligand-activated transcription factor that binds to androgen responsive elements (AREs) associated with androgen- regulated genes. AR is essential for androgen mediated male sexual development and plays a role in prostate cancer etiology and progression. Androgen promotes tissue-specific up- an down-regulation of AR mRNA, suggesting roles for both AR and cell-specific factors in AR mRNA autoregulation. Key goals of this proposal are to identify these non-AR factors and to define their contribution to AR mRNA up-regulation. Because androgen responsiveness is related to AR levels, autoregulation of AR mRNA may be critical in modulating hormonal sensitivity in normal and neoplastic cells. Consistent with this idea, we observed that androgen- mediated up-regulation of AR mRNA and protein is associated with enhanced cellular androgen sensitivity. The AR gene promoter and upstream region have not been demonstrated to contain the ARE and ARE-2, in exons of an internal 6.5 kb androgen responsive region of the AR gene. These Res are located on a 350 bp fragment of the R cDNA that consists of exons D and E. Both the 6.5 kb and 350 bp fragments exhibit cell-specific androgen regulation in reporter gene assays that mirrors up-regulation of the native AR gene. All four AREs are required for AR binding to the 350 bp region and for transcriptional up-regulation of AR mRNA. Furthermore, like the native gene, the 6.5 kb and 350 bp regions are not regulated by glucocorticoid and glucocorticoid receptor (GR) despite the presence of AREs. Resembling the hormone response element (HRE) consensus sequence. HREs are bound and regulated by GR in several other genes. The aims of this proposal are to utilize in vivo foot-printing techniques to examine binding of AR, GR and non-receptor factors to exons D and E of the 6.5 kb androgen responsive region present as stably integrated templates. We will determine whether the patterns of receptor and nuclear factor binding differ in response to androgen (and glucocorticoids) in cells in which the androgen responsive region is active versus inactive. We will identify and characterize the non-receptor factor(s) that binds differentially and determine the role of this factor(s) in cell- and androgen-specific AR mRNA up-regulation. The long term goal of these studies is to provide a comprehensive physical and functional characterization of R mRNA autoregulatory mechanisms and to relate these mechanisms to hormonal responsiveness in normal and neoplastic cells.