Collaborative Research: Elucidating Pre-initiation Complex Assembly and Transcription Initiation by Pol-II Using Advanced Single Molecule and Microfluidic Methods

合作研究：利用先进的单分子和微流体方法阐明 Pol-II 的预启动复合物组装和转录启动

• 批准号: 1244098
• 负责人: Shimon Weiss
• 金额: $ 90万
• 依托单位: University of California-Los Angeles
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-03-15 至 2019-02-28
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1244098&HistoricalAwards=false
• 关键词: Collaborative; Research; Elucidating; Pre; initiation

Intellectual Merit: RNA polymerase II (also called pol II) is an essential enzyme that catalyzes the transcription of DNA into messenger RNA (and most other forms of RNA), an important first step in gene expression. This project will elucidate key structural and functional aspects of the assembly of the Pre-Initiation Complex (PIC) of pol II and the mechanism of transcription initiation, i.e. the mechanistic steps involved in starting the transcription of a new gene. Sophisticated solution-based single-molecule fluorescence techniques, combined with microfluidics, will be applied to individual complexes in order to study how pol II assembles at a promoter (gene start site) and how it performs its catalytic function. Because single-molecule approaches remove ensemble and temporal averaging, information about the order and the kinetics of assembly will be unraveled. Broader Impacts: The "nano-biotechnology revolution" and the emergence of single molecule biophysics and super-resolution imaging have opened up tremendous opportunities in the investigation of processes that define life. One such process, fundamental to the "Central Dogma of Biology," is transcription. The approach for studying this basic process to be taken in this research could break conceptual and practical barriers that have hindered progress, and offers potential to catalyze discovery in many new ways. The tools and methods to be developed, as well as the general approaches pursued, will have greatest impact if distributed to a broad audience of receptive and innovative young scientists. Through educational and outreach programs, the project will expose a young audience (high school and college students), as well as more mature scientists, to novel research tools and methodologies that could be used to study many other biological processes and molecular machines.

智力优势：RNA聚合酶II(也称为PolII)是一种重要的酶，它催化DNA转录成信使RNA(和大多数其他形式的RNA)，这是基因表达的重要第一步。本项目将阐明PolII预起始复合体(PIC)组装的关键结构和功能以及转录起始的机制，即启动新基因转录的机械步骤。复杂的基于溶液的单分子荧光技术，结合微流控技术，将被应用于单个复合体，以研究PolII如何在启动子(基因起始点)组装，以及它如何发挥其催化功能。由于单分子方法去除了系综和时间平均，有关组装的顺序和动力学的信息将被解开。更广泛的影响：“纳米生物技术革命”以及单分子生物物理学和超分辨率成像的出现，为研究定义生命的过程提供了巨大的机会。其中一个过程是转录，它是“生物学的核心法则”的基础。这项研究中将采取的研究这一基本过程的方法可以打破阻碍进展的概念和实践障碍，并提供以许多新方式催化发现的潜力。如果将所开发的工具和方法以及所追求的一般方法分发给接受和创新的青年科学家的广大受众，将产生最大的影响。通过教育和推广计划，该项目将使年轻受众(高中生和大学生)以及更成熟的科学家接触到可用于研究许多其他生物过程和分子机器的新颖研究工具和方法。