Comprehensive Identification of Active Functional Elements in Human Chromatin

人类染色质活性功能元件的综合鉴定

• 批准号: 7925978
• 负责人: GREGORY E CRAWFORD
• 金额: $ 33万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7925978
• 关键词: Atlases; Binding; Biochemical; Biological; Biological Assay; Biological Process; Chromatin; Data; Enhancers; Formaldehyde; Genes; Genome; Goals; Human; Human Genome; Hypersensitivity; Indium; Location; Locus Control Region; Mammalian Cell; Maps; Methods; Nucleosomes; Pilot Projects; Proteins; Regulatory Element; Research Personnel; Resolution; Site; Transcription Initiation Site; cell type; chromatin immunoprecipitation; human DNA; human tissue; promoter; success

DESCRIPTION (provided by applicant): The goal of this proposal is to identify at high resolution all active gene regulatory elements in the human genome among cell types representative of most human tissues. We will accomplish this goal by identifying regions of open chromatin with two independent and complementary methods: DNasel hypersensitivity and Formaldehyde-Assisted Isolation of Regulatory Elements (FAIRE), combined with single-nucleosome mapping and chromatin immunoprecipitation (ChIP) for selected regulatory factors. The immediate benefit of success will be a high-quality public atlas of the human DNA regulatory elements that are likely to be active in each cell type. Identification of open chromatin regions has been one of the most accurate and robust methods to identify functional promoters, enhancers, silencers, insulators, and locus control regions in mammalian cells. A principal advantage of an open chromatin approach is that all potential sites in the genome are simultaneously assayed in an unbiased manner. DNasel hypersensitivity and FAIRE interrogate chromatin by entirely different underlying mechanisms, and therefore represent independent, cross-validating assessments of chromatin state. In addition, we will perform single-nucleosome mapping in selected cell types as an independent direct biochemical verification of open chromatin regions. For a selected subset of cell-types, we will further annotate open chromatin regions with respect to their biological activity by determining the binding location of proteins that mark transcription start sites, transcriptional units, insulators, or have broad regulatory function. The Aims of this proposal are 1) to identify all regions of open chromatin in 40 cell types by DNasel and FAIRE analyses, and 2) determine the biological function of open chromatin regions by ChIP and nucleosome mapping. To accomplish these Aims, we have assembled a team of five researchers who are leaders in their respective fields and who have contributed significantly to the ENCODE pilot project. Furthermore, we have developed a streamlined pipeline, which has been used to generate high quality whole-genome data from a number of cell types.

描述（由申请人提供）： 该提案的目标是在代表大多数人类组织的细胞类型中以高分辨率鉴定人类基因组中的所有活性基因调控元件。我们将通过两种独立和互补的方法识别开放染色质区域来实现这一目标：DNasel超敏反应和甲醛辅助分离调控元件（FAIRE），结合单核小体作图和染色质免疫沉淀（ChIP）选择的调控因子。成功的直接好处将是一个高质量的人类DNA调控元件的公共图谱，这些调控元件可能在每种细胞类型中起作用。 开放染色质区域的鉴定是鉴定哺乳动物细胞中功能性启动子、增强子、沉默子、绝缘子和基因座控制区域的最准确和最稳健的方法之一。开放染色质方法的主要优点是，基因组中的所有潜在位点都以无偏的方式同时测定。DNA酶1超敏反应和FAIRE通过完全不同的潜在机制询问染色质，因此代表了染色质状态的独立、交叉验证评估。此外，我们将在选定的细胞类型中进行单核小体作图，作为开放染色质区域的独立直接生化验证。对于选定的细胞类型的子集，我们将进一步注释开放染色质区域相对于其生物活性，通过确定标记转录起始位点，转录单位，绝缘体，或具有广泛的调节功能的蛋白质的结合位置。 本提案的目的是1）通过DNaseI和FAIRE分析鉴定40种细胞类型中开放染色质的所有区域，以及2）通过ChIP和核小体作图确定开放染色质区域的生物学功能。为了实现这些目标，我们组建了一个由五名研究人员组成的团队，他们是各自领域的领导者，并为ENCODE试点项目做出了重大贡献。此外，我们还开发了一个精简的管道，用于从多种细胞类型中生成高质量的全基因组数据。