Nuclear organization in stem and differentiated cells

干细胞和分化细胞的核组织

• 批准号: 7939808
• 负责人: Victor G. Corces
• 金额: $ 38.75万
• 依托单位: EMORY 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/7939808
• 关键词: Address; Area; Behavior; Binding; Binding Sites; Biological Models; CCCTC-binding factor; Carbon; Cell Cycle; Cell Differentiation process; Cell Nucleus; Cells; Characteristics; Chimeric Proteins; Chromatin; Chromatin Fiber; Chromatin Loop; Chromosomes; Complex; Computing Methodologies; Confocal Microscopy; DNA; DNA Methylation; DNA Sequence; Data; Data Analyses; Development; Drosophila genus; ES Cell Line; Epigenetic Process; Eukaryotic Cell; Gene Expression; Gene Targeting; Genes; Genetic Transcription; Genome; Genomics; Health; Human; Individual; Internet; Laboratories; Life; Light; Maintenance; Mammals; Maps; Mediating; Modeling; Molecular; Molecular Conformation; Molecular Profiling; Monitor; Morphology; Mus; Neurons; Nuclear; Patients; Pattern; Phenotype; Play; Population; Process; Proteins; Psyche structure; Regulation; Role; Site; Stem cells; Stimulus; Structure; Testing; Time; Work; base; cell type; chromatin immunoprecipitation; comparative; demethylation; dopaminergic neuron; embryonic stem cell; gene repression; genome-wide; histone modification; induced pluripotent stem cell; insight; nerve stem cell; nervous system disorder; next generation; nuclear reprogramming; pluripotency; programs; promoter; protein complex; relating to nervous system; research study; response; self-renewal; stem; stem cell differentiation; three dimensional structure

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (14) Stem Cells and specific Challenge Topic, 14-AG-105 Exploratory studies of induced pluripotent stem (iPS) cells from healthy individuals and patients with mental/nervous system disorders. Chromatin insulators are large DNA protein complexes that play a role in regulating gene expression. The structure, composition and function of these complexes is well understood in Drosophila but much less information is available in mammals. Work in our laboratory has contributed in large part to the current view of the role of insulators in nuclear function i.e. the three-dimensional organization of the chromatin fiber within the nucleus. A critical issue now is to determine whether this organization is important for the establishment and/or maintenance of patterns of gene expression that drive cell differentiation. We propose experimental strategies to understand the molecular mechanisms controlling nuclear organization during the process of stem cell differentiation. Given the possible importance of iPS cells to human health, it is critical also to understand all epigenetic changes that take place when differentiated cells are reprogrammed into iPS cells, including changes in nuclear organization. Preliminary results in our lab suggest that the organization of CTCF insulators in mouse stem cells changes as these cells differentiate. In addition, a wealth of information has become recently available describing the genomic localization of the CTCF protein in a variety of cell types. Interactions between these genomic CTCF sites determine the pattern of organization of the DNA in the nucleus. Here we propose to determine the three-dimensional structure of a region of the genome in cells at different states of differentiation. This will be accomplished using 5C to determine all possible interactions between a subset of CTCF sites and analyzing the data using computational methods derived from those used to analyze NMR protein data. The 5C experiments will be carried out with mouse stem cells, neural progenitor cells, neural-derived iPS cells and dopaminergic neurons. Results will shed light on the epigenetic differences and similarities between stem, differentiated and iPS cells. PUBLIC HEALTH RELEVANCE: The arrangement of the DNA of in the nucleus of eukaryotic cells is important in the regulation of gene transcription. DNA sequences called insulators and associated proteins are important in establishing the pattern of DNA organization in the nucleus. We propose to study how insulators contribute to changes in DNA organization taking place when stem cells become differentiated cells and when these cells are re-programmed to become iPS cells.

描述（由申请人提供）：本申请涉及广泛的挑战领域（14）干细胞和特定的挑战主题，14-AG-105 对来自健康个体和患有精神/神经系统疾病的患者的诱导多能干（iPS）细胞的探索性研究。染色质绝缘子是大型 DNA 蛋白质复合物，在调节基因表达中发挥作用。这些复合物的结构、组成和功能在果蝇中已被充分了解，但在哺乳动物中可获得的信息却少得多。我们实验室的工作在很大程度上促进了目前关于绝缘体在核功能中的作用的观点，即核内染色质纤维的三维组织。现在的一个关键问题是确定该组织对于驱动细胞分化的基因表达模式的建立和/或维持是否重要。我们提出了实验策略来了解干细胞分化过程中控制核组织的分子机制。鉴于 iPS 细胞对人类健康可能具有的重要性，了解分化细胞重编程为 iPS 细胞时发生的所有表观遗传变化（包括核组织的变化）也至关重要。我们实验室的初步结果表明，小鼠干细胞中 CTCF 绝缘体的组织随着这些细胞的分化而发生变化。此外，最近出现了大量描述 CTCF 蛋白在多种细胞类型中基因组定位的信息。这些基因组 CTCF 位点之间的相互作用决定了细胞核中 DNA 的组织模式。在这里，我们建议确定处于不同分化状态的细胞中基因组区域的三维结构。这将通过使用 5C 来确定 CTCF 位点子集之间所有可能的相互作用并使用源自用于分析 NMR 蛋白质数据的计算方法来分析数据来完成。 5C 实验将使用小鼠干细胞、神经祖细胞、神经源性 iPS 细胞和多巴胺能神经元进行。结果将揭示干细胞、分化细胞和 iPS 细胞之间表观遗传的差异和相似之处。 公共卫生相关性：真核细胞核中 DNA 的排列对于基因转录的调节非常重要。称为绝缘体的 DNA 序列和相关蛋白质对于建立细胞核中 DNA 组织模式非常重要。我们建议研究当干细胞变成分化细胞以及当这些细胞被重新编程为 iPS 细胞时，绝缘体如何促进 DNA 组织发生变化。