DIFERENTIAL ACTIVATION OF THE VITAMIN D RECEPTOR

描述(改编自申请人摘要)：本申请是一份 继续对类似物的机制进行正在进行的研究 1，25-二羟基维生素D3(1，25D)调节细胞的转录反应 维生素D受体(VDR)。工作假设是接触点 激素和类似物在配体结合口袋中使用的是不同的 从而能够不同地影响VDR的功能表面。 因为VDR的配体结合结构域的表面提供了一个界面 用于与二聚化伙伴、转录共活化子和 辅抑制子，这些相互作用中的任何细微变化都可能改变水平和 VDR介导的基因表达谱。在具体目标1中，佩勒博士和她 实验室将完成对VDR的配体结合口袋的分析 通过定点突变。他们将定义荷尔蒙相互作用的部位 通过比较接触点得到其1-α-羟基和25-羟基 由天然激素和三种类型的配体使用：20-epi类似物， 修饰A环的类似物和25-羟基取代的类似物 一群人。 在具体目标2中，他们将确定不同配体的作用 VDR功能表面的相互作用。同样，使用 天然激素和两组类似物将提供有关 超激动剂产生的功能表面的异同 (20-epi类似物)和细胞特异性非钙激动剂(A环修饰的 类比)。三种类型的配体-受体复合体将被检查 它们诱导与二聚化伙伴相互作用的效力和效力， 辅活化子和辅抑制子。利用定点突变， 还将检查由这些配体中的每一个所创建的表面的组成。 在具体目标3中，Peleg博士的实验室将专注于分子和 细胞特异性类似物作用的细胞机制。他们已经确认了一名 在体内具有低钙化活性的环状修饰类似物，以及深刻的 培养中的细胞分离转录图谱。VDR与这些组件的复合体 类似物将被用作探针，以分离增加或限制的因素 受体在特定细胞环境中的作用。他们将研究是否 细胞特有的行为是由于普通细胞的招募而失去功能 辅阻遏子，由于普通辅活化子的过度表达而获得的功能 或者是细胞特异性因子的招募。 这些研究将促进选择性维生素D受体的开发 可能对治疗各种临床情况有用的调节剂， 包括骨质疏松症、继发性甲状旁腺功能亢进症和癌症。

DESCRIPTION (Adapted from the Applicant's Abstract): This application is a continuation of an ongoing investigation of the mechanisms by which analogs of 1,25-dihydroxyvitamin D3 (1,25D) modulate the transcriptional responses of the vitamin D receptor (VDR). The working hypothesis is that the contact points used by the hormone and analogs in the ligand-binding pocket are different and thereby are able to affect differentially the functional surface of the VDR. Because the surface of the ligand-binding domain of VDR provides an interface for interaction with dimerization partners, transcription coactivators, and corepressors, any subtle change in these interactions may alter the level and spectrum of VDR-mediated gene expression. In Specific Aim 1, Dr. Peleg and her laboratory will complete the analysis of the ligand-binding pocket of the VDR by site-directed mutagenesis. They will define the site of hormone interaction through its 1-alpha-hydroxyl and 25-hydroxyl groups by comparing contact points used by the natural hormone and three types of ligands: 20-epi analogs, analogs with modified A ring and analogs with substitution of their 25-hydroxyl group. In Specific Aim 2, they will determine the effect of differential ligand interaction on the functional surface of the VDR. Again, the use of the natural hormone and two groups of analogs will provide information on the differences and similarities of functional surfaces generated by superagonists (20-epi analogs) and by cell-specific noncalcemic agonists (the A ring-modified analogs). The three types of ligand-receptor complexes will be examined for their potency and efficacy to induce interaction with dimerization partners, coactivators, and corepressors. Using site-directed mutagenesis, the composition of surfaces created by each of these ligands will also be examined. In Specific Aim 3, Dr. Peleg's laboratory will focus on the molecular and cellular mechanism of action of cell-specific analogs. They have identified A ring-modified analogs that have low calcemic activity in vivo, and a profound cell-segregated transcriptional profile in culture. VDR complexes with these analogs will be used as probes to isolate factors that augment or restrict receptor action in a given cellular environment. They will examine whether cell-specific action is due to loss of function by recruitment of a common corepressor, a gain of function due to overexpression of a common coactivator or recruitment of cell-specific factors. These studies will facilitate the development of selective vitamin D receptor modulators that may be useful for treatment of various clinical conditions, including osteoporosis, secondary hyperparathyroidism, and cancer.