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：
9256495
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 2021-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9256495
关键词：
Address Architecture Attention Binding Biochemical Biochemistry Biogenesis Biophysics Cells Cereals Chemicals Chromatin Chromatin Structure Complex DNA Development Developmental Gene Discrimination Elements Enzymes Epigenetic Process Gene Expression Gene Expression Regulation Genes Genetic Transcription Genome Genomics Heritability Histone H3 Histones Immunoprecipitation In Vitro Individual Location Maps Mass Spectrum Analysis Measurable Measurement Measures Methods Modification Molecular Mono-S 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 Recruitment Activity Resolution Rotation Sampling Signal Transduction Site Stem cells 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 spatial relationship 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（由申请人提供）：许多表观遗传信息被认为编码在包装基因组的组蛋白的潜在可遗传化学修饰的身份和定位中，修饰相关的结合配偶体与其结合，从而转导下游的功能后果。染色质的重复单元（核小体核心颗粒）是组蛋白的两倍对称八聚体，被两个 DNA 超螺旋圈包围。这种结构将两个超螺旋圈包裹起来。每个核心组蛋白的副本在确定的位置，每个从结构的核心突出非结构化的“尾部”，对于给定的修饰，是。染色质的每个基本重复单元有两个不同的可修饰位点编码了有意义的信息？有迹象表明这个水平的变异受到高度监管，但由于缺乏可以测量这些特性的工具，人们对染色质修饰的这种规模知之甚少。我们开发了一种突破性的校准 ChIP 技术，使我们能够首次查询这种级别的核小体亚结构细节。在目标 1 中，我们将直接量化核小体内组蛋白修饰的对称性。使用我们校准的 ChIP 方法，然后探究这种新可测量的染色质特性的功能。考虑到结合伴侣的识别单位是整个核小体和侧翼 DNA，而不仅仅是尾部，这一水平上的变异很可能在空间上具体表现出来。为此，我们最近开发了生物化学证据，表明核小体结合伙伴的区分是标记对称性的函数，我们试图了解这种前所未有的水平的机制特性。在目标 2 中，我们将定义二价结构域的分子性质（装饰多能细胞中关键发育基因的规范激活和抑制组蛋白修饰的看似并置）。然后，我们将检查它们的生物发生和预测能力作为分化的障碍，我们希望这项研究的结果能够阐明一般原理。亚核小体标记识别和功能，形成一个令人信服的论点，即这种相对未经探索的染色质修饰水平对于基因组管理很重要。