Project 3: Mechanics of Nuclei, Chromosomes and Chromatin in Cancer

摘要(研究项目3) 项目3旨在了解癌细胞中染色体和细胞核的组织和功能， 将中心的重点放在染色质的时空动力学和癌症的信息传递上。全 细胞的功能最终由细胞核控制，基因在细胞核中表达，在精细的 监管控制。染色体的分子组织和潜在的染色质是 基因调控。存在多少拷贝的基因，转录因子的模式是什么？ 基因调控元件，哪些区域被异染色质严重抑制，哪些区域 携带表观遗传标记都在控制基因表达方面发挥着核心作用。在癌症中，这些因素通常是 --通常是结合在一起--失调，伴随着细胞核和 染色体。项目3试图分析染色质结构的变化--从纤维水平到 从染色体到整个细胞核-使用物理工具与最先进的细胞生物学相结合 接近了。该项目的目标1检验了一系列初步数据支持的假设，即 在癌细胞中，染色体和核的物理组织和机制发生了变化。这将是 用细胞核和染色体的微观力学研究进行测试，结合主要的荧光成像 染色体和核组织蛋白以及核的分波光谱测量 无序(PWS，一项由拟议的纳米细胞测量核心支持的技术)。将进行实验 在其他项目中正在研究的一系列致癌和非致瘤细胞类型中。实验 使用患者来源的异种移植物(PDX)将在患者疾病背景下测试细胞系中开发的想法 (由拟议的PDX核心提供)。目标2将测试如何去除特定的染色体组织蛋白 (使用siRNA和CRISPR方法)影响染色体和核结构，然后将使用结果 目标1和目标2结合Hi-C数据建立中期数学模型 染色体和间期核组织。目标3将专注于染色质纤维本身 在单染色质纤维组装和力学实验中，使用不同的细胞类型进行物理改变 在AIMS 1和AIMS 2中研究的细胞的核提取物，一个主要的目标是将“开放的”或“软的”染色质关联起来 具有特定组蛋白修饰的州。项目3的结果将是一项关于如何 相对于正常细胞，癌细胞中的染色质和染色体被重塑，其研究将是 通过对相同细胞系的平行研究，使用物理分析，与项目1和项目2紧密相连 在项目3中开发，并通过合作开发siRNA和CRISPR方法。

ABSTRACT (RESEARCH PROJECT 3) Project 3 aims to understand organization and function of chromosomes and the nucleus in cancer cells, engaging the Center's focus on spatio-temporal dynamics of chromatin and information transfer in cancer. All functions of the cell are ultimately controlled by the nucleus, in which genes are expressed, under elaborate regulatory control. The molecular organization of chromosomes and the underlying chromatin is central to gene regulation. How many copies of genes are present, what patterns of transcription factors are bound to gene regulatory elements, which regions are heavily repressed by heterochromatinization, and which regions carry epigenetic marks all play a central role in controlling gene expression. In cancer, these factors are often – and usually in combination – dysregulated, with concomitant aberrant structure of the nucleus and chromosomes. Project 3 seeks to analyze the variation in chromatin structure – from the fiber level to chromosomes to the whole cell nucleus – using physical tools in combination with state-of-the-art cell biological approaches. Aim 1 of the project examines the hypothesis, supported by an array of preliminary data, that the physical organization and mechanics of chromosomes and nuclei are altered in cancer cells. This will be tested using micromechanical studies of nuclei and chromosomes, combined fluorescence imaging of major chromosome- and nucleus-organizing proteins, and partial-wave spectroscopy measurements of nuclear disorder (PWS, a technology supported by the proposed Nanocytometry Core). Experiments will be carried out in a range of tumorigenic and non-tumorigenic cell types being studied in the other Projects. Experiments using patient-derived xenografts (PDX) will test ideas developed in cell lines in a patient disease context (supplied by the proposed PDX Core). Aim 2 will test how removal of specific chromosome-organizing proteins (using siRNA and CRISPR methods) affects chromosome and nuclear structure, and then will use the results of both Aim 1 and Aim 2 in combination with Hi-C data to develop mathematical models of metaphase chromosome and interphase nucleus organization. Aim 3 will focus on how chromatin fibers themselves are physically altered in different cell types, in single-chromatin-fiber assembly and mechanics experiments using nuclear extracts from cells studied in Aims 1 and 2, a primary goal being to correlate “open” or “soft” chromatin states with specific histone modifications. The result of Project 3 will be a comprehensive study of how chromatin and chromosomes are remodeled in cancer cells relative to normal cells, and its research will be tightly linked to Project 1 and Project 2 via parallel studies of the same cell lines, use of the physical assays developed in Project 3, and through cooperation on development of siRNA and CRISPR methodologies.