Molecular mechanism of Androgen Receptor mediated transcription

摘要 转录因子是基因表达的关键决定因素。亲脂性激素配体及其伴生物 共调节分子触发转录因子的活性，包括核激素的成员 受体(NR)家族。AR及其剪接变异体AR-V7是在前列腺癌发生发展中发挥关键作用的NRs， 尤其是CRPC。目前针对AR的治疗主要集中在其配体结合域(LBD)，其中 在AR-V7中不存在。最初对这些疗法有反应的患者在几天内就会对它们产生抵抗力 好几年了。AR和AR-V7都必须招募COR才能在功能上发挥作用。AR-COR接口中断 抑制雄激素依赖细胞和CRPC细胞的AR活性。因此，了解AR如何变化 与特定的CORs相互作用形成转录活性的复合体对于治疗的设计是至关重要的 瞄准AR和AR-V7。我们的初步研究首次提供了对活性NR-COR结构的理解 复杂的组装，并证明了构象变化对NR介导的深远影响 转录激活。 在这个提案中，我们假设AR及其变体有一组共同的COR，但程序集和 这些COR在AR波群中的三维排列是各自独特的，并有助于 转录活动的调控。我们建议利用调查人员的互补专业知识在 NR生物学、冷冻EM和图像处理以确定转录活性AR的结构基础 复合体。我们将通过两个具体目标实现这一目标：1)解决高分辨率DNA-AR结构以 详细识别结构域-结构域相互作用，然后将其与DNA-AR-V7的结构进行比较；以及2)改进 AR-COR络合物结构的分辨以识别详细的相互作用并确定结构 与AR-V7-COR复合波的差异。这两个目标都将利用冷冻EM来可视化功能性AR- COR复合体。拟议的工作具有重要意义，因为这些结构将描述 该系统确定哪些成分应该作为治疗调节的靶点。一种结构 了解AR如何形成功能二聚体并与CORs相互作用以激活基因表达将 提供有关转录生物学的关键信息，并使未来寻找小分子- 分子抑制剂可以影响AR复合体的排列。拟议的多学科工作具有创新性。 因为它使用了先进的成像技术来实现对结构和 AR-COR复合体、AR异源二聚体及其靶向药物的功能。

Abstract Transcription factors are key determinants of gene expression. Lipophilic hormonal ligands and accompanying co-regulatory molecules trigger the activity of transcription factors, including members of the nuclear hormone receptor (NR) family. AR and its splice variant AR-V7 are NRs that play key roles in prostate cancer development, and particularly CRPC. Current therapies targeting AR mainly focus on its ligand-binding domain (LBD), which is not present in AR-V7. Patients that respond initially to those therapies become resistant to them within a few years. Both AR and AR-V7 must recruit CoRs to be functionally active. Disruption of the AR–CoR interface inhibits AR activity in both androgen-dependent cells and CRPC cells. Therefore, knowledge of how AR variants interact with specific CoRs to form a transcriptionally active complex is critical for the design of therapeutics targeting AR and AR-V7. Our preliminary studies provided the first structural understanding of active NR–CoR complex assembly and demonstrated that conformational variability has a profound impact on NR-mediated transcriptional activation. In this proposal, we hypothesize that AR and its variants have a common set of CoRs, but that the assembly and three-dimensional arrangement of those CoRs in AR complexes are unique to each and contribute to the regulation of transcriptional activities. We propose to leverage the complementary expertise of investigators in NR biology, cryoEM, and image processing to determine the structural basis of transcriptionally active AR complexes. We will pursue that goal through two specific aims: 1) Solve a high-resolution DNA–AR structure to identify domain-domain interactions in detail and then compare it to the structure of DNA–AR-V7; and 2) Improve the resolution of AR–CoR complexes structures to identify detailed interactions and determine the structural differences in comparison with AR-V7–CoR complexes. Both aims will utilize cryoEM to visualize functional AR– CoR complexes. The proposed work is significant because the structures will describe the overall interactions in the system to determine which components should be targeted for therapeutic modulation. A structural understanding of how AR forms functional dimers and interacts with CoRs to activates gene expression will provide critical information about the biology of transcription and enable future studies of looking for small- molecular inhibitors can affect the AR complex arrangement. The proposed multidisciplinary work is innovative because it employs advanced imaging techniques to achieve unprecedented insights into the structure and function of AR–CoR complexes, AR heterodimers, and the drugs that target them.