CTCF dependent chromatin looping dynamics

CTCF 依赖性染色质循环动力学

• 批准号: 8490149
• 负责人: Hideo Mabuchi
• 金额: $ 25.72万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-01 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8490149
• 关键词: Affinity; Antibody Diversity; Behavior; Binding; Binding Sites; Biological Process; CCCTC-binding factor; Cell Extracts; Cell Nucleus; Cells; Chromatin; Chromatin Loop; Chromosome Condensation; Chromosome Segregation; Chromosome Structures; Chromosomes; Code; DNA; DNA Binding; DNA Methylation; Development; Dimerization; Distant; Enhancers; Eukaryotic Cell; Fluorescence; Frequencies; Gene Cluster; Gene Expression; Generations; Genes; Genetic Recombination; Genetic Transcription; Genome; Globin; Goals; Histones; Homodimerization; Human; Immunoglobulins; In Vitro; Individual; Length; Measurement; Measures; Mechanics; Mediating; Methods; Mitotic Chromosome; Modeling; Modification; Molecular Conformation; Mutagenesis; Nuclear; Nucleic Acid Regulatory Sequences; Pattern; Plasmids; Play; Process; Property; Proteins; Regulation; Relative (related person); Role; Site; Spectrum Analysis; Techniques; Testing; Text; Transcriptional Activation; Vertebrates; Zinc Fingers; abstracting; chromatin protein; cohesin; cohesion; gene repression; imprint; in vivo; insight; novel; promoter; public health relevance; research study; single molecule; statistics

Enter the text here that is the new abstract information for your application. The organization of the nucleus and the regulated folding of the genome plays essential roles in regulating gene expression, chromosome segregation and chromosome structure. Long range interactions in chromatin are required for activation of gene transcription and repression of genes during the differentiation of eukaryotic cells. Long-range contacts between different chromosomal loci also regulate processes such as antibody diversity and mitotic chromosome condensation. Despite the wide range of processes that involve chromatin loops we know very little about the mechanisms that drive chromatin folding and stabilize long range interactions in the genome. This proposal is focused on developing new methods to measure the formation of looped domains dependent upon the activity of the chromatin protein CTCF. CTCF is known to be required for the stabilization of looped regions in the genome but how it generates or stabilizes looped domains is unknown. We propose to first characterize the dynamics of CTCF dependent looping using defined chromatin substrates in vitro and on chromatinized plasmids in cell extracts. Using mutagenesis and depletion we will alter the binding affinity and dimerization properties of CTCF and its interaction with the loop stabilizing protein cohesin to determine how these activities regulate the frequency of loop generation. Using the insight we gain from these in vitro experiments we will compare the dynamics of loop formation to the statistics of long range interactions at the human globin locus. By depleting CTCF and cohesin we will relate the cellular statistics of loop formation to the in vitro mechanics of loop stabilization. Our studies should provide unique and novel insight into the processes that regulate the formation of long range chromatin interactions and how they relate to essential developmental and cell biological processes.

在此处输入文本，这是您应用程序的新摘要信息。 细胞核的组织和基因组的调节折叠起着重要作用 在调节基因表达，染色体分离和染色体结构中。长的 激活基因转录和抑制需要染色质的范围相互作用 真核细胞分化过程中的基因。不同的远程接触 染色体基因座还调节抗体多样性和有丝分裂染色体等过程 缩合。尽管涉及染色质环的过程范围很广，但我们非常了解 几乎没有关于驱动染色质折叠和稳定远距离相互作用的机制 基因组。该建议的重点是开发新方法来衡量 循环结构域取决于染色质蛋白CTCF的活性。 CTCF已知 基因组中循环区域的稳定所必需，但如何产生或 稳定循环域是未知的。我们建议首先表征CTCF的动力学 使用定义的染色质底物在体外和染色质质粒上使用依赖性循环 在细胞提取物中。使用诱变和耗尽，我们将改变结合亲和力 CTCF的二聚化特性及其与环路稳定蛋白粘着蛋白的相互作用 确定这些活动如何调节循环生成的频率。使用我们的见解 从这些体外实验中获得，我们将比较循环形成的动力学与 人球蛋白基因座的远程相互作用的统计数据。通过耗尽CTCF和粘蛋白 我们将将循环形成的细胞统计数据与循环的体外力学联系起来 稳定。我们的研究应该为过程提供独特而新颖的见解 调节远距离染色质相互作用的形成及其与必不可少的 发育和细胞生物学过程。