Mechanisms of enhancer-promoter communication, genome organization and transcription control

摘要 后生动物基因组通过将调控DNA元件与靶基因解偶联实现复杂的基因控制 启动子，并允许远程调节。因此，基因可以在不同的细胞中差异表达， 类型和不同的环境信号或发展线索下。远端调控元件 （增强子）靶向特定的基因启动子，搜索过程是如何被基因组的拓扑结构塑造的 以及增强子-启动子相互作用如何通过调节复合物促进， RNA聚合酶II的信号和控制转录活性仍然是一个谜。我们的目标是 了解使人类增强子-启动子通讯的分子和生物物理机制 和其他哺乳动物细胞。为了实现这些目标，在这个奖项期间，我们将完成 （i）可视化增强子和靶启动子的动态通讯，同时使用 调节复合物和基因活性的关联，使用新的单分子和超分辨率 用于单个活细胞中基因组的结构和功能的非侵入性4D成像的方法;（ii） 确定不同类型的结构蛋白质塑造基因组折叠的机制，增强子- 启动子通讯和转录动力学;（iii）剖析个体的功能和相互依赖性 在控制细胞身份基因的复杂调控景观中的组成增强子元件。我们的结果 将建立定量框架，了解生物化学的转录调控， 拥挤的环境中的细胞核和解释基因调控和基因组组织使用软- 物质/聚合物物理学和相关的生物物理学概念。这些概念上的飞跃是必要的， 了解亚Mb尺度下的物理染色质组织，该尺度与调控基因组最相关 交互.我们的综合结构-功能方法将为以下方面提供功能验证和关键测试： 基因“远距离调节”模型。拟议的研究不仅将提供大量的新知识， 启动子-增强子通讯的机制，但也将为进一步研究 基因组拓扑/组织和基因表达调控相互作用。

ABSTRACT Metazoan genomes achieve complex gene control by uncoupling regulatory DNA elements from target promoters and allowing regulation at a distance. Thus, a gene can be differentially expressed in different cell types and under different environmental signals or developmental cues. How distal regulatory elements (enhancers) target specific gene promoters, how the search process is shaped by the topology of the genome in the nucleus and how enhancer-promoter interactions are facilitated by regulatory complexes that relay signals to the RNA Polymerase II and control transcription activity remains a mystery. Our goal is to understand molecular and biophysical mechanisms that enable enhancer-promoter communication in human and other mammalian cells. Towards these goals and during the period of this award we will accomplish the following: (i) visualize the dynamic communication of enhancers and target promoters simultaneously with the association of regulatory complexes and gene activity, using novel single-molecule and super-resolution approaches for non-invasive 4D imaging of structure and function of the genome in single live cells; (ii) determine mechanisms by which different classes of architectural proteins shape genome folding, enhancer- promoter communication and transcription kinetics; (iii) dissect the function and interdependencies of individual constituent enhancer elements within complex regulatory landscapes controlling cell identity genes. Our results will establish quantitative frameworks for understanding the biochemistry of transcription regulation in the crowded environment of the nucleus and for interpreting gene regulation and genome organization using soft- matter/polymer physics and related biophysical concepts. These conceptual leaps are needed to ultimately understand physical chromatin organization at sub-Mb scales, the scale most relevant for regulatory genome interactions. Our integrated structure-function approach will provide functional validation and critical tests for gene “regulation-at-a-distance” models. The proposed studies will not only provide substantial new knowledge on the mechanisms of promoter-enhancer communication but will also set the stage for further studies of the interplay of genome topology/organization and gene expression regulation.