Dynamic regulation of initiation and pausing of transcription by human RNA polymerase II

人RNA聚合酶II对转录起始和暂停的动态调节

• 批准号: 1818147
• 负责人: Dylan Taatjes
• 金额: $ 120万
• 依托单位: University of Colorado at Boulder
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1818147&HistoricalAwards=false
• 关键词: Dynamic; regulation; initiation; pausing; transcription

The human genome contains about 3 billion base pairs of DNA that provide essential instructions for cells to respond to developmental or environmental cues. A key step in the response is transcribing the information in DNA to intermediary RNA molecules that supply another layer of information. Transcribing DNA to RNA is a highly regulated process, but exactly how it is controlled remains an enigma. The goal of this project is to decipher this problem by bringing together an experienced team of investigators to take a multi-faceted approach to catalyze discovery in new ways. The research plan will integrate educational and outreach programs at University of Colorado-Boulder and University of California-Los Angeles to expose high school, college students, and postdoctoral scholars to high-impact collaborative research. The tools, strategies, and methodologies to be implemented will be applicable to the study of other biological processes and molecular machines.The questions addressed in this project are fundamentally important for understanding the molecular control of eukaryotic transcription. The human RNA polymerase II transcription machinery will be studied in detail using cutting-edge methods developed initially through analysis of bacterial RNA polymerase. Highly regulated stages in gene expression, including transcription initiation and pausing, will be the focus. This collaborative project will merge solution-based single-molecule fluorescence techniques with detailed functional assays enabled by biochemical reconstitution of the entire 4.5 MDa human transcription apparatus. Two key questions will be addressed: How is RNA polymerase II transcription dynamically regulated by specific protein factors? What role does RNA polymerase II backtracking play in regulating initiation and pausing? An important aspect of the work will be to test and validate mechanistic findings with in vitro and cell-based approaches.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

人类基因组包含约30亿个DNA碱基对，为细胞对发育或环境线索做出反应提供必要的指令。 反应的关键步骤是将DNA中的信息转录到中间RNA分子，后者提供另一层信息。 将DNA转录为RNA是一个高度调控的过程，但它究竟是如何被控制的仍然是一个谜。 该项目的目标是通过汇集经验丰富的调查人员团队，采取多方面的方法以新的方式促进发现，来破解这个问题。该研究计划将整合科罗拉多大学博尔德分校和加州大学洛杉矶分校的教育和推广计划，使高中生、大学生和博士后学者参与高影响力的合作研究。本研究所采用的工具、策略和方法将适用于其他生物过程和分子机器的研究，本研究所解决的问题对于理解真核生物转录的分子控制具有重要意义。人类RNA聚合酶II的转录机制将详细研究使用尖端的方法，最初通过分析细菌RNA聚合酶。 基因表达的高度调控阶段，包括转录起始和暂停，将是重点。 该合作项目将合并基于溶液的单分子荧光技术与详细的功能测定，通过整个4.5 MDa人类转录装置的生化重建实现。两个关键问题将得到解决：如何是RNA聚合酶II转录动态调节特定的蛋白质因子？ RNA聚合酶II回溯在调节起始和暂停中起什么作用？这项工作的一个重要方面将是测试和验证与体外和细胞为基础的方法mechanistic发现。这个奖项反映了NSF的法定使命，并已被认为是值得通过使用基金会的智力价值和更广泛的影响审查标准进行评估的支持。

项目成果

High-throughput smFRET analysis of freely diffusing nucleic acid molecules and associated proteins.

• DOI: 10.1016/j.ymeth.2019.07.021
• 发表时间: 2019-10-01
• 期刊: Methods (San Diego, Calif.)
• 作者: Segal M;Ingargiola A;Lerner E;Chung S;White JA;Streets A;Weiss S;Michalet X
• 通讯作者: Michalet X

The Mediator kinase module: an interface between cell signaling and transcription.

• DOI: 10.1016/j.tibs.2022.01.002
• 发表时间: 2022-04
• 期刊: Trends in biochemical sciences
• 影响因子: 13.8
• 作者: Luyties O;Taatjes DJ
• 通讯作者: Taatjes DJ

An overview of continuous and discrete phasor analysis of binned or time-gated periodic decays

分箱或时间选通周期性衰减的连续和离散相量分析概述

• DOI: 10.1117/12.2577747
• 发表时间: 2021
• 期刊: SPIE
• 作者: Michalet, Xavier

The Role of XPB/Ssl2 dsDNA Translocase Processivity in Transcription Start-site Scanning.

• DOI: 10.1016/j.jmb.2021.166813
• 发表时间: 2021-07-09
• 期刊: Journal of molecular biology
• 影响因子: 5.6
• 作者: Tomko EJ;Luyties O;Rimel JK;Tsai CL;Fuss JO;Fishburn J;Hahn S;Tsutakawa SE;Taatjes DJ;Galburt EA
• 通讯作者: Galburt EA

TFIID Enables RNA Polymerase II Promoter-Proximal Pausing

• DOI: 10.1016/j.molcel.2020.03.008
• 发表时间: 2020-05-21
• 期刊: MOLECULAR CELL
• 影响因子: 16
• 作者: Fant, Charli B.;Levandowski, Cecilia B.;Taatjes, Dylan J.
• 通讯作者: Taatjes, Dylan J.