Computational and experimental insights into the structure and dynamics of heterochromatin

对异染色质结构和动力学的计算和实验见解

• 批准号: 10300059
• 负责人: Nathaniel A. Hathaway
• 金额: $ 30.42万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-12-01 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10300059
• 关键词: Algorithms; Architecture; Behavior; Biochemical Reaction; Biological Assay; Biology; Calibration; Cell Culture Techniques; Cells; Chemical Structure; Chemicals; Chromatin; Chromatin Fiber; Chromatin Structure; Complement; Complex; Computer Models; DNA; Data; Data Set; Development; Disease; Doxycycline; Enzymes; Epigenetic Process; Equilibrium; Event; Feedback; Fiber; Gene Expression Profiling; Genes; Goals; Grain; Heterochromatin; Individual; Investigation; Knowledge; Language; Lentivirus Vector; Life; Malignant Neoplasms; Measures; Mediating; Methodology; Methods; Modeling; Molecular; Molecular Profiling; Mus; Pathogenesis; Pathway interactions; Phase; Physics; Physiological; Process; Production; Proteins; Regenerative Medicine; Regulatory Element; Repression; Research; Resolution; Role; Running; Signal Transduction; Sirolimus; Stem cell pluripotency; Stochastic Processes; Structure; System; Technology; Testing; Time; Tretinoin; Visualization; Work; base; cancer therapy; chromosome conformation capture; computational platform; computerized tools; embryonic stem cell; experimental study; gene repression; heterochromatin-specific nonhistone chromosomal protein HP-1; improved; in vivo; induced pluripotent stem cell; insight; interest; mechanical properties; methylation pattern; overexpression; particle; physical property; promoter; protein protein interaction; real time monitoring; recruit; response; simulation; small hairpin RNA; temporal measurement

Abstract Chemical, molecular and structural transformations of chromatin are intimately involved in critical cellular phenomena, including differentiation, signaling, and pathogenesis. A detailed knowledge of how molecular complexes involving multiple kilobases of DNA and hundreds of proteins respond to the finest changes in chemical structure is key to elucidating the role of chromatin transformations in life and disease. The overarching goal of this project is to develop and apply computational tools to investigate how the structure and dynamics of chromatin determine its functional states. Our central hypothesis is that physical properties and behavior of the chromatin fiber and associated proteins lend themselves to encoding into efficient and useful ultra-coarse- grained (UCG) representations. Our strategy to reach the goal is by bridging together several computational and experimental methodologies. We initiated the development of Molecular Biosystems (MB), a computational platform for UCG simulations specifically adapted to the chromatin biology. MB methodology represents a blend of physics-based mechanisms, such as dynamics of the chromatin fiber, with stochastic processes encompassing protein-protein interactions and enzymatic reactions. MB studies will be complemented by all-atom MD and CG simulations and experimentally tested using a unique chromatin in vivo assay (CiA) methodology. Specifically, we will investigate the chromatin-mediated repression of Oct4, a key gene regulating embryonic stem (ES) cell pluripotency at defined points in mammalian development. This is important because the ability to reverse the Oct4 repression would streamline production of induced pluripotent cells (iPSC) and advance regenerative medicine. The CiA technology at the Oct4 locus in mouse ES cells will be used for the exploration of changes to chromatin structure, as well as for testing the adequacy of MB simulations. Experimental endpoints that are directly comparable to computational hypotheses will be produced: (1) fraction of Oct4-repressed cells in cell culture; (2) H3K9 methylation patterns on Oct4 promoter; and (3) chromatin conformation capture. Three main components of our research are: (i) Extending and enhancing the UCG MB approach; (ii) Multi- scale simulations of chromatin processes to elucidate the structure and dynamics of heterochromatin of Oct4 regulatory elements; (iii) Experimental real-time monitoring of heterochromatin molecular signatures using Chromatin in vivo Assay (CiA) to study mechanisms and time course of Oct4 de-repression and provide feedback for the computational models. This work is important because of its focus on the physics of the gene repression, whose understanding will bring us one step forward toward the promise of regenerative medicine and new prospects for cancer therapy.

摘要 染色质的化学、分子和结构变化与关键细胞密切相关 现象，包括分化、信号和发病机制。详细了解分子如何 涉及多个千碱基的DNA和数百个蛋白质的复合体对 化学结构是阐明染色质转化在生命和疾病中的作用的关键。最重要的是 这个项目的目标是开发和应用计算工具来研究 染色质决定其功能状态。我们的中心假设是人类的物理特性和行为 染色质纤维和相关蛋白质有助于编码成高效和有用的超粗蛋白质- 粒化(UCG)表示。我们实现这一目标的策略是通过将几个计算和 实验方法。我们发起了分子生物系统(MB)的开发，这是一种计算 专门适用于染色质生物学的UCG模拟平台。MB方法代表了一种混合 基于物理的机制，例如染色质纤维的动力学，其随机过程包括 蛋白质-蛋白质相互作用和酶反应。甲基溴研究将得到全原子MD和CG的补充 使用一种独特的染色质体内分析(CIA)方法进行模拟和实验测试。 具体地说，我们将研究染色质介导的Oct4的抑制，Oct4是调节胚胎发育的关键基因 干细胞(ES)在哺乳动物发育的特定时间点具有多能性。这一点很重要，因为 逆转Oct4抑制将简化诱导多能细胞(IPSC)的生产并促进 再生医学。小鼠胚胎干细胞Oct4基因座的CIA技术将用于探索 染色质结构的变化，以及测试甲基溴模拟的充分性。实验终端 将产生与计算假设直接可比的结果：(1)Oct4抑制细胞的一部分 在细胞培养中；(2)Oct4启动子上的H3K9甲基化模式；(3)染色质构象捕获。 我们研究的三个主要部分是：(I)推广和加强UCG MB方法；(Ii)多种- 染色质过程的尺度模拟研究Oct4异染色质的结构和动力学 调节元件；(3)异染色质分子特征的实验实时监测 染色质体内试验(CIA)研究Oct4抑制的机制和时间进程并提供反馈 用于计算模型。 这项工作之所以重要，是因为它专注于基因抑制的物理学，他们的理解将 让我们朝着再生医学的前景和癌症治疗的新前景迈进一步。