Understanding Gene Transcription from First-Principles: A single-molecule study

描述(由申请人提供) 摘要：基因表达的准确调控对于多细胞生物体的正常发育和维持成体组织的细胞内稳态至关重要。基因表达失控是越来越多的人类疾病的标志，许多药物直接针对关键的基因转录因子。然而，目前的治疗方法会影响A基因的转录 泛基因组规模，没有特异性，也没有能力微调基因活性。我们的不足 对转录调控的详细途径的理解排除了设计靶向治疗的可能性。此外，目前的实验工具无法解决涉及人类mRNA转录的生化系统的复杂性，该系统由多组分的核心转录机制和多层额外的调节因子、共激活因子、阻遏物和染色质重塑复合体组成。关键的是，目前还没有一种实验方法可以观察活细胞内这些因子的动态，剖析转录动力学，识别受调控的限速步骤，并最终 揭示机械原理。我的目标是开发能够提供高分辨率动态数据的光学成像和光谱工具，以便于用详细的分子术语描述转录调控。在获奖期间，我预计将朝着这个方向实现以下目标：(1)创造创新技术，使在活细胞内实时、单分子水平上可视化复杂的生化过程成为可能；(2)识别体内转录因子调节mRNA合成周期中限速步骤的机制；(3)为理解信号整合和基因表达的组合控制提供一个框架。这项拟议的研究建立在我在物理和生物科学边缘的独特专业知识的基础上，并寻求创造一种新的想法和方法的综合，以新的战略理解和控制基因表达，以一种与医学相关的方式。 与公共健康相关：转录是基因表达的第一步，也是最严格的调控步骤。转录异常与越来越多的疾病有关，几种药物直接针对转录因子。本项目旨在阐明核心分子转录机制的功能，并为控制其调控治疗应用提供新的策略。

DESCRIPTION (Provided by the applicant) Abstract: The accurate regulation of gene expression is crucial for proper development of multi-cellular organisms and for maintaining cell homeostasis in adult tissues. De-regulation of gene expression is the hallmark of an ever-increasing number of human diseases and many drugs directly target key gene transcription factors. Current therapies however affect transcription on a pan-genomic scale, without specificity and without the ability to fine-tune gene activity. Our lack of understanding of the detailed pathways underlying transcriptional regulation precludes designing targeted therapies. Moreover, current experimental tools cannot address the complexity for the biochemical system involved in human mRNA transcription, comprised by multi-component core transcription machinery and multiple additional layers of regulatory factors, co-activators, repressors and chromatin remodeling complexes. Critically, no current experimental method can observe the dynamics of these factors inside a live cell, dissect transcription kinetics, identify rate-limiting steps that are subject to regulation, and ultimately uncover mechanistic principles. My goal is to develop the enabling optical imaging and spectroscopy tools that can provide high-resolution, dynamic data, to facilitate describing transcription regulation in detailed molecular terms. During the period of this award I envision accomplishing the following towards this direction: (1) create innovating technologies that will make possible to visualize complex biochemical processes at the single-molecule level, in real-time, inside living cells; (2) discern in vivo mechanisms by which transcription factors regulate rate-limiting steps in the cycle of mRNA synthesis; (3) provide a framework for understanding signal integration and combinatorial control of gene expression. The proposed research builds upon my unique expertise at the border of physical and biological sciences, and seeks to create a new synthesis of ideas and methodologies towards novel strategies to understand and control gene expression, in a way relevant to medicine. Public Health Relevance: Transcription is the first and most highly regulated step in gene expression. Aberrant transcription is associated with an ever-growing number of diseases and several drugs directly target transcription factors. This project aims at elucidating the function f the core molecular transcription machinery and provide new strategies for controlling its regulation for therapeutic applications.