Modular Platform for Combinatorial Epigenome Manipulation

用于组合表观基因组操作的模块化平台

基本信息

批准号：
10592628
负责人：
Mark D ADAMS
金额：
$ 73.47万
依托单位：
JACKSON LABORATORY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-01-01 至 2023-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10592628
关键词：
Address Adoption Affect Amino Acid Sequence Biological Biological Assay Biological Models Biological Process Cell Line Cell model Cell physiology Cells ChIP-seq Clustered Regularly Interspaced Short Palindromic Repeats Communities Custom DNA Deposition Development Developmental Process Disease Ecosystem Elements Engineering Enhancers Enzymes Epigenetic Process Etiology Gene Expression Gene Expression Regulation Genes Genomics Goals Guide RNA Health Histones In Situ Individual Knock-out Libraries Link Maps Methodology Modification Molecular Open Reading Frames Pathway interactions Phenotype Physiology Plasmids Play Proteins Protocols documentation Publishing RNA library Reagent Registries Regulator Genes Regulatory Element Reporter Research Personnel Resources Scientist Shapes Site Specificity Stretching System Techniques Technology Therapeutic Intervention Transgenes Translating Validation base case-by-case basis cell type combinatorial computational pipelines design developmental disease epigenetic regulation epigenome epigenome editing epigenomics experimental study functional genomics gene regulatory network genome-wide genome-wide analysis genomic locus histone modification human embryonic stem cell in vivo induced pluripotent stem cell innovation insight interest next generation next generation sequencing novel personalized medicine precision medicine programs promoter repository self-renewal side effect tool transcriptome sequencing

Address Adoption Affect Amino Acid Sequence Biological Biological Assay Biological Models Biological Process Cell Line Cell model Cell physiology Cells ChIP-seq Clustered Regularly Interspaced Short Palindromic Repeats Communities Custom DNA Deposition Development Developmental Process Disease Ecosystem Elements Engineering Enhancers Enzymes Epigenetic Process Etiology Gene Expression Gene Expression Regulation Genes Genomics Goals Guide RNA Health Histones In Situ Individual Knock-out Libraries Link Maps Methodology Modification Molecular Open Reading Frames Pathway interactions Phenotype Physiology Plasmids Play Proteins Protocols documentation Publishing RNA library Reagent Registries Regulator Genes Regulatory Element Reporter Research Personnel Resources Scientist Shapes Site Specificity Stretching System Techniques Technology Therapeutic Intervention Transgenes Translating Validation base case-by-case basis cell type combinatorial computational pipelines design developmental disease epigenetic regulation epigenome epigenome editing epigenomics experimental study functional genomics gene regulatory network genome-wide genome-wide analysis genomic locus histone modification human embryonic stem cell in vivo induced pluripotent stem cell innovation insight interest next generation next generation sequencing novel personalized medicine precision medicine programs promoter repository self-renewal side effect tool transcriptome sequencing

展开

项目摘要

Epigenetic modifications of histone and DNA control gene expression and critically shape phenotypes and cell states. These modifications are tightly controlled by interactions with a constellation of trans-acting regulatory factors, and are dysregulated in a plethora of diseases. Next-generation sequencing (NGS) technologies have allowed genome-wide profiling of these modifications in diverse cell types, normal and disease conditions as well as across individuals. However, these strategies have yielded mostly associative and/or correlative insight into relationships between epigenetic state, gene expression and phenotype with limited power to establish the causality of individual epigenetic modifications that is fundamental to understanding normal physiology and diseases. Existing techniques to investigate phenotypic consequences of epigenetic gene regulation irreversibly delete stretches of genomic sequence that can have multiple functions, or perturb protein factors that control thousands of genes, confounding accurate conclusion. A scalable toolbox allowing in situ and in vivo combinatorial modifications of epigenetic states would offer unprecedented opportunities for both mechanistic studies and unbiased discovery of novel functional elements at the genomic scale. This project will respond to this need by developing an expandable molecular toolkit to precisely and reversibly manipulate defined epigenetic modifications (e.g., H3K27ac) at defined genomic addresses based on our innovative CRISPR/Casilio platform. There are three Specific Aims in this project. Aim 1 is focused on the development of a comprehensive set of epigenetic editing modules with which Casilio can achieve multiplexed and combinatorial edits, whereby different epigenetic modifications can be elicited simultaneously at distinct genomic loci while multiple modifications can also be induced in each locus. Aim 2 will deliver Casilio-enabled cell lines with which scientists can achieve epigenome editing with unprecedented ease by delivering short RNA guides. Aim 3 will center on the development of genome-wide guide libraries targeting genomic regulatory elements such as enhancers and insulators with which scientists can perform reverse epigenetic screens to discover novel elements and epigenetic modifications causative to phenotype. This toolkit, once developed, will transform the ways we study epigenetics by providing a fine and scalable technique to directly edit epigenetic states at defined targets, to investigate the underlying causes of gene regulatory changes observed in biological processes and diseases. To truly benefit the field of functional genomics, we will share our methodology and reagents at every stage of platform development with the scientific community. Through the establishment of standards and a module registry as well as reagent sharing via an open repository, we hope to create a sustainable ecosystem of Casilio module users and developers to apply and expand the Casilio toolbox for epigenetic editing. Due to the programmability and precision, epigenetic editing enabled by Casilio may provide new means and tools for powering personalized and precision medicine.

组蛋白和DNA控制基因表达的表观遗传学修饰，并批判性塑造表型和细胞国家。这些修饰是通过与反式调节的星座相互作用而紧密控制的因素，在大量疾病中失调。下一代测序（NGS）技术具有允许在各种细胞类型（正常和疾病状况）中对这些修饰的全基因组分析为以及跨个人。但是，这些策略主要产生的关联和/或相关见解进入表观遗传状态，基因表达和表型之间的关系有限的能力个体表观遗传修饰的因果关系，这是理解正常生理学和疾病。研究表观遗传基因调控的表型后果的现有技术不可逆转地删除可能具有多个功能或扰动蛋白因子的基因组序列的拉伸控制数千个基因，使得准确的结论混淆。可扩展的工具箱，允许原位和体内组合对表观遗传状态的修改将为两者提供前所未有的机会机械研究和基因组量表上新型功能元件的无偏见。这个项目将通过开发可扩展的分子工具包来精确而可逆地操纵这种需求定义的表观遗传修饰（例如，H3K27AC）在我们创新的基因组地址定义的基因组地址 CRISPR/CASILIO平台。该项目有三个具体的目标。 AIM 1专注于发展 Casilio可以实现多重和组合编辑，可以同时在不同的表观遗传修饰下同时引发不同的表观遗传修饰基因组基因座虽然也可以在每个基因座中诱导多个修饰。 AIM 2将提供支持西里奥的科学家可以通过简短的简便性来实现表观基因组编辑的细胞系 RNA指南。 AIM 3将集中于针对基因组的全基因组指南库的发展法规元素（例如增强剂和绝缘子）可以通过这些元素进行反向表观遗传学筛选以发现对表型的新型元素和表观遗传修饰。这个工具包一次开发的，将通过提供一种可直接的精细可扩展技术来改变我们研究表观遗传剂的方式在定义靶标处编辑表观遗传状态，以研究基因调节变化的根本原因在生物过程和疾病中观察到。为了真正受益于功能基因组学领域，我们将分享我们的方法和试剂在平台开发的每个阶段都与科学界一起开发。通过建立标准和模块注册表以及通过公开存储库共享的试剂共享，我们希望创建Casilio模块用户和开发人员的可持续生态系统，以应用和扩展 Casilio工具箱，用于表观遗传编辑。由于可编程性和精度，表观遗传编辑由 Casilio可以提供为个性化和精确医学供电的新手段和工具。