NUCLEAR AND CHROMOSOME DECONDENSATION DURING G1-S

G1-S 期间的核和染色体解凝结

• 批准号: 3467697
• 负责人: Andrew Steven Belmont
• 金额: $ 10.72万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-07-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3467697
• 关键词: DNA replication; antibody; cell nucleus; chromatin; chromosomes; computer program /software; electron microscopy; gold; image enhancement; image processing; imaging /visualization /scanning; lamins; phase contrast microscopy; physical model; tissue /cell culture

The highest levels of chromatin structure, consisting of the packing of 100 and 300 angstrom chromatin fibers within mitotic and interphase chromosomes, are unknown. Understanding this level of chromatin organization, describing the structure of entire transcription or replication units, is essential to answering fundamental questions related to gene regulation, DNA recombination, cell differentiation, and cell cycle control. This project's long term objectives are to develop a model system allowing dissection of basic structure-function motifs involved in the large-scale chromatin organization of mammalian interphase chromosomes. Specifically, it will focus on the nuclear and chromosomal decondensation occurring during cell cycle progression through Gl into S phase. The strategy will be to develop an in vitro system reproducing decondensation events occurring in vivo, and to exploit this system to analyze their underlying biochemical and structural basis and functional relevance to DNA replication initiation. The grant will lay the foundation for this long term project. A detailed 3-dimensional structural analysis will compare nuclear and chromosome decondensation in vivo during Gl progression with decondensation induced in vitro using cell free extracts. Several 3-dimensional reconstruction methods, including light microscopy optical sectioning, electron microscopy serial sectioning, and electron microscopy axial tomography, will provide overlapping resolution over a complete range of size scales. The specific aims of this grant will include the following: Interphase chromosome decondensation in vivo and in vitro will be described in terms of the intranuclear arrangement of interphase chromosomes, association of chromosomes with nuclear lamins, uncoiling of interphase chromosomes into their component large-scale chromatin structures, and packaging of 300 angstrom chromatin fibers within these individual large- scale domains. The unfolding pathway of early versus late replicating regions will be contrasted, and the architecture of actively replicating chromatin will be determined. Finally, quantitative upper limits on the experimental errors associated with these structural descriptions will be estimated.

染色质结构的最高水平，由100个 和300埃染色质纤维在有丝分裂和间期 染色体是未知的。 了解这一层次的染色质 组织，描述整个转录的结构或 复制单位，是必不可少的回答有关的基本问题， 基因调控、DNA重组、细胞分化和细胞周期 控制 该项目的长期目标是开发一个模型系统， 大规模的结构-功能基序的解剖 哺乳动物间期染色体的染色质组织。 具体地说， 它将集中在细胞核和染色体的解凝聚发生， 在细胞周期进行期间通过G1期进入S期。 该战略将 开发一个体外系统， 发生在体内，并利用这一系统来分析其潜在的 生物化学和结构基础以及与DNA的功能相关性 复制起始。 这笔赠款将为这个长期项目奠定基础。 详细 3-三维结构分析将比较细胞核和染色体 在GI进展期间体内去凝聚， 使用无细胞提取物进行体外培养。 多次三维重建 方法，包括光学显微镜光学切片，电子显微镜 连续切片和电子显微镜轴向断层扫描，将提供 重叠分辨率在一个完整的范围大小尺度。 具体 这笔赠款的目的将包括： 间期染色体去凝聚在体内和体外将被描述 根据间期染色体的核内排列， 染色体与核纤层的联合，间期的解旋 染色体转化为它们的组成大规模染色质结构，和 300埃的染色质纤维包装在这些单独的大， 尺度域 早期复制与晚期复制的展开途径 区域将进行对比，积极复制的架构 将确定染色质。 最后，关于 与这些结构描述相关的实验误差将是 估算