Mechanistic Investigation of RNA-Mediated Gene Regulation and Immunity

项目摘要 这项拟议的研究结合了生物物理学、分子遗传学和基因组学工具，以提供高度的 核糖核酸主导的生物过程的分解机制。我们的研究集中在以下两个中心主题上 核糖核酸生物学。(1)RNA可以作为信息载体来指导蛋白质的作用。这样的系统是 因此，在生物技术和医学领域具有巨大的潜力，例如 例如RNAi和CRISPR-Cas。我的实验室花了十多年的时间剖析了 CRISPR-CAS，以及最近一直在开发基于我们的机制的基因组编辑应用程序 洞察力。(2)RNA可以采用特定的三级结构发挥重要功能。我的实验室已经阐明了 许多核糖开关的结构、功能和构象动力学。高分辨率信息可以 使新抗生素的开发成为可能。我们现在的目标是将遗传筛选和生化纯化相结合，以 系统地确定孤儿核糖开关的配体。这样的研究经常揭示细菌的新方面 生理学，甚至在真核生物中可能是保守的。我们建议在以下四个方面开展工作： 研究：1)I型CRISPR(CRISPR-Cas3)的结构、机制及基因组编辑应用；2) Cas1-Cas2整合酶建立CRISPR免疫的机制：3)机制与基因组编辑 CRISPR-Tn7的应用；4)剖析核糖开关调控机制和细菌生理学。

Project Summary The proposed research combines biophysics, molecular genetics, and genomics tools to provide high- resolution mechanisms for RNA-dominated biological processes. Our study focuses on two central themes in RNA biology. (1) RNA can serve as an information carrier to guide the action of proteins. Such systems are inherently precise and programmable, therefore carry great potential in biotechnology and medicine, as exemplified by RNAi and CRISPR-Cas. My lab spent more than ten years dissecting the mechanisms of CRISPR-Cas, and more recently has been developing genome editing applications based on our mechanistic insights. (2) RNA can adopt defined tertiary structures to exert important functions. My lab has elucidated the structure, function, and conformation dynamics of many riboswitches. The high-resolution information may enable new antibiotics development. We now aim to combine genetic screen with biochemical purification to systematically identify ligands for orphan riboswitches. Such studies often reveal novel aspects of bacterial physiology, which may even be conserved in eukaryotes. We propose to work on the following four areas of research: 1) Structure, mechanism, and genome editing applications of Type I CRISPR (CRISPR-Cas3); 2) Mechanism of CRISPR immunity establishment by Cas1-Cas2 integrase; 3) Mechanism and genome editing application of CRISPR-Tn7; 4) Dissecting riboswitch-controlled mechanism and bacterial physiology.