The effect of nucleosomes on the earliest stages of RNA polymerase II transcription

核小体对 RNA 聚合酶 II 转录最早阶段的影响

基本信息

批准号：
9766312
负责人：
Donal Luse
金额：
$ 33.25万
依托单位：
CLEVELAND CLINIC LERNER COM-CWRU
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-01 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9766312
关键词：
Active Sites Address Adopted Affect Architecture Biochemical Biological Assay Cell Nucleus Cell physiology Chromatin Chromatin Structure Complex Correlation Studies DNA DNA Polymerase II DNA-Directed RNA Polymerase Drug Targeting ERCC3 gene Elements Eukaryota Event Gene Expression Gene Expression Regulation Genetic Transcription Genomics Health Histones Human In Vitro Lead Location Modeling Movement Nature Nucleosomes Orthologous Gene Pattern Population Positioning Attribute Process Property Regulation Research Personnel Role Saccharomyces cerevisiae Scanning Shapes Signal Transduction Site-Directed Mutagenesis Surveys System TAF1 gene Testing Transcript Transcription Initiation Transcription Initiation Site Variant Work Yeasts base experimental study human disease in vivo mutant promoter recruit response transcription factor transcription factor TFIIH translocase

项目摘要

Project Summary Appropriate control of gene expression is essential for correct cellular function and is disrupted in many human diseases. RNA polymerase II (Pol II) is the engine of gene expression and the target of regulation for almost every facet of its function. In the nucleus of the cell, Pol II is recruited to promoters, locates transcription start sites, and enters into transcription in a chromatin context. This occurs amidst flanking nucleosomes, which are the primary unit of chromatin. It is clear that control of the earliest stages of transcription is a major aspect of gene regulation in eukaryotes from simple yeast to humans. Genomic studies have suggested that the encounter of Pol II with the first nucleosome immediately downstream of the promoter (the +1 nucleosome) is important in this regulatory process, but the results of those studies are correlations, based on averages of distributed populations. The significance of those correlations is rarely tested by direct biochemical assays. This proposal will provide those tests by adopting biochemical approaches developed in the Luse lab to directly address the functional importance of the promoter proximal nucleosome for assembly of the Pol II transcription complex and the initial transition into transcript elongation. We will also determine the consequences for the +1 nucleosome resulting from the advance of the transcriptional machinery. Our in vitro systems have the unique ability to reveal the fundamental nature of Pol II/nucleosome interactions that additional factors can modulate or regulate. We will employ both the human and yeast transcription systems. These are the best understood eukaryotic transcription systems at the biochemical level and they provide complementary strengths for our experiments. We will incorporate the full complexity of eukaryotic promoters (both containing and lacking the TATA motif) in our work, which allows us to address the real possibility that the response of Pol II to the +1 nucleosome depends on promoter architecture. We will build on the extensive earlier studies by both investigators to begin testing proposed models of RNA polymerase II-nucleosome interactions with approaches that lead to mechanistic conclusions.

项目摘要适当控制基因表达对于正确的细胞功能至关重要，并且被破坏在许多人类疾病中。 RNA聚合酶II（POL II）是基因表达的发动机和几乎所有功能方面的调节目标。在细胞的细胞核中，Pol II为招募到启动子，找到转录起始位点，并进入转录染色质上下文。这发生在核小体侧面，这是染色质。显然，对转录的最早阶段的控制是基因的主要方面从简单酵母到人类的真核生物的调节。基因组研究表明 POL II与启动子下游的第一个核小体的相遇（ +1核小体）在此调节过程中很重要，但是这些研究的结果是相关性，基于分布人群的平均值。那些的意义相关性很少通过直接的生化测定来测试。该建议将提供这些测试通过采用Luse Lab开发的生化方法直接解决功能启动子近端核小体对于组装Pol II转录的重要性复杂和初始过渡到转录本的伸长。我们还将确定转录的前进产生的+1核小体的后果机械。我们的体外系统具有揭示POL的基本性质的独特能力 II/核小体相互作用，其他因素可以调节或调节。我们将雇用两者人和酵母转录系统。这些是最好的真核生物在生化水平上的转录系统，它们为我们提供互补的优势实验。我们将结合真核启动子的全部复杂性（都包含并且缺乏塔塔主题），这使我们能够解决真正的可能性 Pol II对+1核小体的反应取决于启动子结构。我们将建立在两位研究者的大量早期研究开始测试提出的RNA模型聚合酶II核体与导致机械结论的方法的相互作用。