Identifying Determinants of Chromosomal Domains as Defined by Time of Replication

识别由复制时间定义的染色体结构域的决定因素

• 批准号: 7828093
• 负责人: Mignon A Keaton
• 金额: $ 5.05万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7828093
• 关键词: Architecture; Binding Sites; Biological Assay; Boundary Elements; Cell Nucleus; Chromatin; Chromatin Loop; Chromosome Structures; Chromosomes; Communication; DNA; Disease; Down-Regulation; Enhancers; Fluorescent in Situ Hybridization; Gene Expression; Genes; Genome; Genomics; Hela Cells; Heterochromatin; Human Genome; Interphase; Maps; Mediating; Molecular Conformation; Nuclear; Nuclear Matrix; Nuclear Matrix-Associated Proteins; Nuclear Structure; Phase; Plasmids; Regulation; Reporter Genes; Research; Site; Structure; Techniques; Technology; Time; Transgenes; chromatin modification; insight; prevent; programs; promoter; time use

DESCRIPTION (provided by applicant): In this proposal, we aim to identify DNAelements that dictate the organization of the genome into functional chromosomal domains. Large regions of the genome are coordinately replicated during S phase according to a specific spatial and temporal program. These replication domains are likely determined by the overall organization of the genome into highly-ordered structures. In order to further our understanding of nuclear organization and replication timing, this proposal aims to identify replication boundary elements that are responsible for chromatin domain organization. First, potential boundary elements (PBEs) will be identified by changes in replication timing using fluorescence in situ hybridization techniques. As boundary elements are believed to be tethered to nuclear structures by insulator proteins, nuclear matrix attachment and CTCF insulator binding sites will also be mapped to identify PBEs. Next, identified replication PBEs will be assayed for enhancer-blocking and barrier insulator activity using gene expression assays. Finally, a chromosomal interactions with replication domain PBEs will be identified using chromosomeconformation capture technology. These results will yield a better understanding of how replication domains are organized and insight to the mechanisms that regulate replication domain structures. This research will investigate how DNA is duplicated and organized within the cell nucleus. Regulation of chromosome organization is one mechanism that determines if a gene is turned "on" or "off1. Since alterations in gene expression can result in a variety of diseases, studying how DNA is organized within the nucleus will further our understanding of disease formation.

描述（由申请人提供）： 在此提案中，我们旨在确定将基因组组织到功能性染色体领域的组织。根据特定的空间和时间程序，基因组的大区域在S期进行了协调复制。这些复制域可能由基因组的整体组织确定为高度有序的结构。为了进一步了解核组织和复制时间，该提案旨在确定负责染色质域组织的复制边界要素。首先，使用荧光原位杂交技术，将通过复制时机的变化来确定潜在的边界元素（PBE）。由于据信通过绝缘子蛋白将边界元素束缚在核结构上，因此还将绘制核基质附着和CTCF绝缘子结合位点以识别PBE。接下来，将使用基因表达测定法测定确定的复制PBE，以增强障碍物和屏障绝缘体活性。最后，将使用染色体编制捕获技术确定与复制域PBE的染色体相互作用。这些结果将更好地了解如何组织复制域，并了解调节复制域结构的机制。这项研究将研究如何在细胞核内复制和组织DNA。染色体组织的调节是一种确定基因是“ ON”或“ OFF1”的机制。由于基因表达的改变会导致多种疾病，研究核中如何组织DNA将进一步了解我们对疾病形成的理解。