Insulator-mediated chromatin organization during neural lineage commitment

神经谱系定型过程中绝缘体介导的染色质组织

• 批准号: 7870494
• 负责人: Jennifer Elizabeth Phillips-Cremins
• 金额: $ 5.43万
• 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: 2009-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7870494
• 关键词: Adult; Base Sequence; Behavior; Binding; Binding Sites; Biological Models; CCCTC-binding factor; Cell Cycle; Cell Nucleus; Cell division; Cells; Characteristics; Chimeric Proteins; Chromatin; Chromatin Fiber; Chromatin Loop; Chromatin Modeling; Chromatin Structure; Chromosomes; Confocal Microscopy; Cues; Development; Developmental Biology; Diffuse; ES Cell Line; Embryo; Embryonic Development; Epigenetic Process; Figs - dietary; Foundations; Gene Expression; Gene Targeting; Genes; Genome; Genomics; Heterochromatin; Higher Order Chromatin Structure; Histocompatibility Testing; Human; Insulator Elements; Knowledge; Life; Location; Maintenance; Malignant Neoplasms; Malignant neoplasm of brain; Maps; Mediating; Methylation; Modeling; Modification; Molecular; Molecular Conformation; Molecular Profiling; Monitor; Morphology; Mus; Neurodegenerative Disorders; Neurons; Nuclear; Pattern; Phenotype; Play; Pluripotent Stem Cells; Population; Proteins; Regenerative Medicine; Research; Research Personnel; Role; Somatic Cell; Stem cells; Stimulus; Structure; System; Testing; Therapeutic; Time; Work; base; brain cell; cell type; chromatin immunoprecipitation; comparative; demethylation; embryonic stem cell; gene repression; genome wide association study; genome-wide; histone modification; insight; mRNA Differential Displays; next generation; pluripotency; programs; promoter; public health relevance; relating to nervous system; response; self-renewal; stem; stem cell differentiation

DESCRIPTION (provided by applicant): Emerging evidence supports a central role for chromatin insulators in genome-wide organization of higher- order chromosome loop structures. The overall objective of this proposal is to examine the consequences of insulator-mediated chromatin organization during mouse embryonic stem (ES) cell differentiation and development. Our central hypothesis is that the vertebrate insulator CTCF partitions the genome into lineage-specific domains of co-expressed genes by facilitating the formation of higher-order chromatin loop structures in response to developmental cues. This hypothesis will be tested according to three specific aims. In Aim 1, live cell confocal microscopy will be leveraged to investigate CTCF distribution and dynamics in real time during ES cell commitment along the neural lineage. In Aim 2, genome-wide CTCF binding sites will be identified in ES cells using chromatin immunoprecipitation in combination with high- throughput, next generation sequencing (ChlP-Seq). Global CTCF occupancy maps for pluripotent ES cells will be compared to multipotent neuroprogenitors and terminally-differentiated neurons. Finally, in Aim 3, the structural organization of CTCF-based chromatin loops will be characterized using Chromosome- Conformation-Capture (3C) at genomic loci displaying differential insulator occupancy during neural lineage commitment. Correlation of these CTCF-based structures with recent genome-wide analyses of 'traditional' epigenetic modifications and lineage-specific gene expression profiles will provide a more global understanding of how the genome and the epigenome act in concert to regulate the formation of a diverse array of tissue-types during development. Completion of the proposed work will provide significant insight into the mechanisms that govern the commitment of pluripotent stem cells toward neuroectodermal lineages. This knowledge will enable advances in understanding the causes and consequences of higher- order chromatin structure during the onset of cancer and neurodegenerative diseases. Public Health Relevance: Embryonic stem cells have enormous potential for regenerative medicine due to their capacity for indefinite self-renewal while remaining poised for differentiation into all adult cell-types. The proposed research will enhance our understanding of the molecular mechanisms that regulate commitment of stem cells along the neural lineage during embryonic development. This knowledge will provide a foundation for the development of robust strategies which harness the therapeutic potential of stem cells for treatment of neurodegenerative diseases and brain cancer.

描述（由申请人提供）： 新出现的证据支持染色质绝缘体在高阶染色体环结构的全基因组组织中的核心作用。该提案的总体目标是检查小鼠胚胎干（ES）细胞分化和发育过程中绝缘体介导的染色质组织的后果。我们的中心假设是，脊椎动物绝缘体 CTCF 通过促进响应发育线索的高阶染色质环结构的形成，将基因组划分为共表达基因的谱系特异性结构域。该假设将根据三个具体目标进行检验。在目标 1 中，将利用活细胞共聚焦显微镜实时研究 ES 细胞沿神经谱系定型期间的 CTCF 分布和动态。在目标 2 中，将使用染色质免疫沉淀结合高通量下一代测序 (ChlP-Seq) 来鉴定 ES 细胞中的全基因组 CTCF 结合位点。多能 ES 细胞的全局 CTCF 占据图将与多能神经祖细胞和终末分化神经元进行比较。最后，在目标 3 中，将使用染色体构象捕获 (3C) 来表征基于 CTCF 的染色质环的结构组织，这些基因座在神经谱系定型期间显示出不同的绝缘体占据情况。这些基于 CTCF 的结构与最近对“传统”表观遗传修饰和谱系特异性基因表达谱的全基因组分析的关联，将提供对基因组和表观基因组如何协同作用以调节发育过程中多种组织类型的形成的更全面的理解。完成拟议的工作将为控制多能干细胞向神经外胚层谱系的承诺提供重要的见解。这些知识将有助于更好地理解癌症和神经退行性疾病发病过程中高阶染色质结构的原因和后果。公共健康相关性：胚胎干细胞具有无限自我更新的能力，同时保持分化为所有成体细胞类型的能力，因此在再生医学方面具有巨大的潜力。拟议的研究将增强我们对胚胎发育过程中调节干细胞沿神经谱系的分子机制的理解。这些知识将为开发强有力的策略奠定基础，利用干细胞的治疗潜力来治疗神经退行性疾病和脑癌。