Quantitatively probing intra-nucleosomal chromatin variation and function

定量探测核小体内染色质变异和功能

基本信息

批准号：
9904744
负责人：
Alexander Jackson Ruthenburg
金额：
$ 30.82万
依托单位：
UNIVERSITY OF CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-04-06 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9904744
关键词：
Address Architecture Attention Bar Codes Binding Biochemical Biochemistry Biogenesis Biological Process Biophysics Cells Chemicals Chromatin Chromatin Structure Complex DNA Development Developmental Gene Discrimination Elements Enzymes Epigenetic Process Gene Expression Genes Genetic Transcription Genome Genomics Grain Heritability Histone H3 Histones Immunoprecipitation In Vitro Individual Location Maps Mass Spectrum Analysis Measurable Measurement Measures Methods Modification Molecular NURF Nature Nucleosome Core Particle Nucleosomes Organism Outcome Output Partner in relationship Pathway interactions Pattern Positioning Attribute Post-Translational Protein Processing Property Protein Isoforms Proteins Proteomics Protomer Recombinants Resolution Rotation Sampling Signal Transduction Site Structure Surface Tail Technology Time Transcription Initiation Site Transcriptional Regulation Ursidae Family Variant chromatin immunoprecipitation chromatin modification chromosomal location enzyme activity epigenetic regulation exhaustion experimental study genome-wide histone modification insight programs public health relevance reconstitution recruit spatial relationship stem cells tool trimethyllysine

项目摘要

DESCRIPTION (provided by applicant): Much epigenetic information is thought to be encoded in the identity and localization of potentially heritable chemical modifications to histone protein that package the genome, to which modification-contingent binding partners bind, thereby transducing downstream functional consequences. The fundamental repeating unit of chromatin, the nucleosome core particle, is a two-fold symmetric octamer of histone proteins enshrouded by two superhelical turns of DNA. This architecture places the two copies of each core histone in defined positions each projecting unstructured "tails" from the core of the structure to that are subject to dense posttranslational modification. For a given modification, is there meaningful information encoded by having two distinct modifiable sites per fundamental repeating unit of chromatin? There are hints that variation at this level is highly regulated, yet little is known about this scale of chromatin modifications owing to lack of tools that can measure these properties. We have developed a breakthrough calibrated ChIP technology that permits us to query this level of nucleosome sub-structure detail for the first time. In Aim 1 we will directly quantify the symmetry of histone modifications within nucleosomes with our calibrated ChIP method, then probe the function of this newly measurable chromatin property. Given that the unit of recognition for binding partners entire nucleosome and flanking DNA, as opposed to merely the tails, precisely how variation at this level spatially manifests is likely tobe an important element of discrimination. To this end, we have recently developed biochemical evidence that nucleosomal binding partners discriminate as a function of mark-symmetry. We seek to understand the mechanistic properties of this unprecedented level of recognition, both in its biophysical details and its functional consequences for cells and organisms. In Aim 2 we will define the molecular nature of bivalent domains-the seeming apposition of canonically activating and repressive histone modifications decorating critical developmental genes in pluripotent cells-- using calibrated sequential ChIP experiments calibrated with an exhaustive set of internal standards. We will then examine their biogenesis and predictive power as barriers to differentiation. We expect that the results of this study will illuminate the general principlesof sub-nucleosomal mark recognition and function, forming a compelling argument that this relatively un-explored level of chromatin modification is important for genome management.

描述（应用程序提供）：许多表观遗传信息被认为是在包装基因组的组蛋白的身份和定位中编码的，并定位了基因组，将修饰的结合伴侣结合到下游功能后果。染色质的基本重复单元，核无生成核心粒子，是由两个DNA的两个超螺旋转弯所包围的组蛋白蛋白的两倍对称八聚体。该体系结构将每个核心组蛋白的两个副本置于定义的位置，每个构造的位置都会投射从结构的核心到结构的核心，从而受到密集的翻译后修改。对于给定的修改，是每个基本重复单元的染色质单位具有两个不同的可修改位点，编码有意义的信息？有一些暗示，在此级别上的变化受到了高度调节，但由于缺乏可以测量这些属性的工具，因此对这种染色质修饰的规模知之甚少。我们已经开发了一种突破性的校准芯片技术，使我们首次查询了这种核小体子结构细节。在AIM 1中，我们将使用校准的芯片方法直接量化核小体内组蛋白修饰的对称性，然后探测该新可测量的染色质特性的功能。鉴于结合伴侣的识别单位，而不是仅尾巴的核体和侧翼DNA，因此，该水平的变异在空间上表现如何可能是歧视的重要元素。为此，我们最近开发了生化证据，表明核渗透构件伴侣会区分与标记对称的函数。我们试图了解这种前所未有的识别水平的机械性能，包括其生物物理细节及其对细胞和组织的功能后果。在AIM 2中，我们将使用校准的顺序芯片实验进行了用详尽的内部标准组校准的校准顺序芯片实验，定义了二价结构域的分子性质 - 在多能细胞中装饰关键的发育基因的典型激活和反射性组蛋白的修饰。然后，我们将研究它们的生物发生和预测能力作为分化的障碍。我们预计这项研究的结果将阐明亚核标记识别和功能的一般原则，这形成了令人信服的论点，即这种相对未探索的染色质修饰水平对于基因组管理很重要。