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: 10061636
• 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 2023-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10061636
• 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; Molecular Structure; 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、分子和结构转化与细胞内重要的生物学过程密切相关。 现象，包括分化，信号传导和发病机制。详细了解分子如何 涉及DNA的多个酶和数百种蛋白质的复合物对细胞中最细微的变化做出反应， 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。总体 这个项目的目标是开发和应用计算工具来研究如何结构和动力学的 染色质决定其功能状态。我们的中心假设是， 染色质纤维和相关蛋白质有助于编码成有效和有用的超粗- UCG（Grained Representations）我们的战略，以达到这一目标是通过桥接在一起的几个计算和 实验方法学我们发起了分子生物系统（MB）的发展，一个计算 UCG模拟平台专门适用于染色质生物学。甲基溴方法是一种混合 基于物理学的机制，如染色质纤维的动态，随机过程包括 蛋白质-蛋白质相互作用和酶促反应。MB研究将得到全原子MD和CG的补充 使用独特的染色质体内测定（CiA）方法进行模拟和实验测试。 具体而言，我们将研究染色质介导的Oct 4的抑制，Oct 4是调控胚胎发育的关键基因， 在哺乳动物发育的特定点，干细胞（ES）多能性。这很重要，因为 逆转Oct 4抑制将简化诱导多能细胞（iPSC）的生产， 再生医学小鼠ES细胞Oct 4位点的CiA技术将用于探索 染色质结构的变化，以及用于测试MB模拟的充分性。实验终点 将产生与计算假设直接可比的结果：（1）Oct 4抑制细胞的分数 在细胞培养物中;（2）Oct 4启动子上的H3 K9甲基化模式;和（3）染色质构象捕获。 我们的研究的三个主要组成部分是：（一）扩展和加强UCG MB方法;（二）多 染色质过程的规模模拟，以阐明Oct 4异染色质的结构和动力学 （iii）利用基因组学方法实时监测异染色质分子特征 染色质体内试验（CiA），用于研究Oct 4去阻遏的机制和时间过程，并提供反馈 用于计算模型。 这项工作是重要的，因为它的重点是基因抑制的物理，其理解将 使我们朝着再生医学的承诺和癌症治疗的新前景前进了一步。