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

从第一原理理解基因转录：单分子研究

• 批准号: 8355484
• 负责人: Alexandros Pertsinidis
• 金额: $ 256.95万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-30 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8355484
• 关键词: Address; Adult; Affect; Award; Biochemical; Biochemical Process; Biological Sciences; Cells; Chromatin Remodeling Factor; Complex; Data; Development; Disease; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Genomics; Goals; Homeostasis; Human; Kinetics; Life; Medicine; Messenger RNA; Methodology; Methods; Molecular; Organism; Pathway interactions; Pharmaceutical Preparations; Regulation; Research; Resolution; Signal Transduction; Specificity; Spectrum Analysis; System; Technology; Therapeutic; Time; Tissues; Transcription factor genes; Transcriptional Regulation; abstracting; combinatorial; design; human disease; in vivo; innovation; novel strategies; optical imaging; physical science; public health relevance; single molecule; tool; transcription factor

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.

描述（由申请人提供） 摘要：基因表达的精确调控对于多细胞生物的正常发育和维持成体组织中的细胞稳态至关重要。基因表达失调是人类疾病数量不断增加的标志，许多药物直接针对关键基因转录因子。然而，当前的疗法会影响转录 泛基因组规模，没有特异性，也没有微调基因活性的能力。我们的缺乏 对转录调控的详细途径的了解阻碍了设计靶向疗法。此外，当前的实验工具无法解决人类 mRNA 转录所涉及的生化系统的复杂性，该系统由多组件核心转录机制和多层额外的调节因子、共激活因子、阻遏因子和染色质重塑复合物组成。重要的是，目前没有任何实验方法可以观察活细胞内这些因素的动态，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。我的目标是开发可提供高分辨率动态数据的光学成像和光谱工具，以方便以详细的分子术语描述转录调控。在获得该奖项期间，我设想朝着这个方向实现以下目标：（1）创建创新技术，使活细胞内单分子水平上的复杂生化过程实时可视化成为可能； (2) 辨别转录因子调节 mRNA 合成周期限速步骤的体内机制； （3）提供理解信号整合和基因表达组合控制的框架。拟议的研究建立在我在物理和生物科学领域的独特专业知识的基础上，旨在以与医学相关的方式创建新的思想和方法论综合体，以实现理解和控制基因表达的新策略。 公共卫生相关性：转录是基因表达的第一个也是最受监管的步骤。异常转录与越来越多的疾病有关，并且有几种药物直接针对转录因子。该项目旨在阐明核心分子转录机制的功能，并为控制其治疗应用的调节提供新策略。

项目成果

Volumetric interferometric lattice light-sheet imaging.

体积干涉晶格光片成像。

• 发表时间: 2021
• 影响因子: 46.9
• 作者: Cao, Bin;Coelho, Simao;Li, Jieru;Wang, Guanshi;Pertsinidis, Alexandros
• 通讯作者: Pertsinidis, Alexandros

Simple and versatile imaging of genomic loci in live mammalian cells and early pre-implantation embryos using CAS-LiveFISH.

使用 CAS-LiveFISH 对活哺乳动物细胞和早期植入前胚胎的基因组位点进行简单且多功能的成像。

• 发表时间: 2021-06-09
• 影响因子: 4.6
• 作者: Geng, Yongtao;Pertsinidis, Alexandros
• 通讯作者: Pertsinidis, Alexandros

Single-gene imaging links genome topology, promoter-enhancer communication and transcription control.

单基因成像将基因组拓扑、启动子-增强子通讯和转录控制联系起来。

• 发表时间: 2020
• 影响因子: 16.8
• 作者: Li, Jieru;Hsu, Angela;Hua, Yujing;Wang, Guanshi;Cheng, Lingling;Ochiai, Hiroshi;Yamamoto, Takashi;Pertsinidis, Alexandros
• 通讯作者: Pertsinidis, Alexandros

Single-Molecule Nanoscopy Elucidates RNA Polymerase II Transcription at Single Genes in Live Cells.

单分子纳米显微镜阐明了活细胞中单基因的 RNA 聚合酶 II 转录。

• 发表时间: 2019
• 影响因子: 64.5
• 作者: Li, Jieru;Dong, Ankun;Saydaminova, Kamola;Chang, Hill;Wang, Guanshi;Ochiai, Hiroshi;Yamamoto, Takashi;Pertsinidis, Alexandros
• 通讯作者: Pertsinidis, Alexandros

A Single-Molecule Surface-Based Platform to Detect the Assembly and Function of the Human RNA Polymerase II Transcription Machinery.

用于检测人类 RNA 聚合酶 II 转录机器的组装和功能的单分子表面平台。

• 发表时间: 2020
• 作者: Park, Sang Ryul;Hauver, Jesse;Zhang, Yunxiang;Revyakin, Andrey;Coleman, Robert A;Tjian, Robert;Chu, Steven;Pertsinidis, Alexandros
• 通讯作者: Pertsinidis, Alexandros