The Effects of Physical Disruption on Genome Organization and Integrity

描述（由申请人提供）：人类基因组在细胞核中的三维（3D）组织不仅对于在2微米直径的细胞核内包装2米的DNA很重要，而且对于基因调控和其他重要的生物功能也很重要。利用最近开发的Hi-C技术来捕获细胞内整个基因组的染色体折叠和循环，拟议的研究将调查这种基因组组织与细胞和DNA所经历的物理破坏之间的关系。DNA双链断裂（DSB）的剧烈物理破坏可导致癌症和其他疾病，如果由不正确的断裂修复导致的染色体易位破坏基因或基因与调控元件之间的关系。因此，了解影响DSB后可能发生易位的因素是很重要的。测量小鼠B淋巴细胞中基因组组织的Hi-C实验将用于确定基因组区域的预先存在的邻近性与在相同细胞中诱导DSB后观察到的易位位置相关的程度。在物理接近的高概率不能解释复发性易位的情况下，其他因素对易位易感性的贡献将通过计算整合本研究中获得的相互作用和易位数据与先前发表的基因表达，蛋白质-DNA相互作用和染色体常见脆性位点等特征的数据进行检查。3D基因组结构的物理性质对非近端基因组区域之间易位形成的影响将通过将实验观察到的易位与来自聚合物物理学原理和实验Hi-C数据的基因组生物物理模型进行比较来评估。该模型将用于预测基因组结构的物理性质在多大程度上可能限制或增强基因组区域和DNA断裂末端在寻找修复伙伴时的可移动性。在这些模拟之后，将通过对受到物理力和约束的细胞进行Hi-C实验，对基因组的变形性进行直接实验测试。这些力实验的结果将测试生物物理模型的预测，并使物理变形性和易位易感性之间的比较。成功地观察细胞内基因组对物理力的反应也将提供洞察组织和器官中的生物相关力是否可能通过直接改变染色质结构来影响基因表达，细胞行为和细胞命运。这样的洞察力有助于未来 力在用于组织工程目的的细胞定向分化中的应用。通过整合新的实验数据，物理模型和先前发表的基因组数据，这项研究将有助于更全面地了解复杂的3D基因组对健康和疾病的物理破坏的反应所涉及的因素。

DESCRIPTION (provided by applicant): The three-dimensional (3D) organization of the human genome in the nucleus is important not only for packing 2 meters of DNA inside a 2 micron diameter nucleus, but also for gene regulation and other important biological functions. Taking advantage of the Hi-C technology recently developed to capture chromosome folding and looping throughout the genome within a cell, the proposed research will investigate the relationship between this genome organization and physical disruptions experienced by the cell and DNA. The dramatic physical disruption of a DNA double strand break (DSB) can lead to cancer and other diseases if chromosomal translocations resulting from incorrect break repair disrupt genes or the relationship between genes and regulatory elements. Thus, it is important to understand the factors influencing which translocations are likely to occur after a DSB. Hi-C experiments measuring genome organization in mouse B lymphocytes will be used to determine the extent to which pre-existing proximity of genomic regions correlates with the locations of translocations observed after an induced DSB in the same cells. In cases where a high probability of physical proximity does not explain recurrent translocations, the contribution of other factors to translocation susceptibility will be examined by computationally integrating interaction and translocation data obtained in this research with previously published data on features such as gene expression, protein-DNA interactions, and chromosomal common fragile sites. The influence of the physical properties of the 3D genome structure on the formation of translocations between non-proximal genome regions will then be evaluated by comparing experimentally observed translocations with a biophysical model of the genome derived from principles of polymer physics and experimental Hi-C data. This model will be used to predict how much the physical properties of the genome structure might constrain or enhance the movability of genomic regions and broken ends of DNA in their search for repair partners. These simulations will be followed by direct experimental tests of the deformability of the genome by performing Hi-C experiments on cells subjected to physical forces and constraints. The results of these force experiments will test the biophysical model predictions and enable comparisons between physical deformability and translocation susceptibility. Successfully observing the response of the genome inside cells to physical forces will also provide insight into whether biologically relevant forces in tissues and organs might influence gene expression, cell behavior, and cell fate by direct changes to the chromatin structure. Such insight could contribute to future applications of force in the directed differentiation of cells for tissue engineering purposes. By integrating new experimental data, physical models, and previously published genomic data, this research will contribute to a more complete understanding of the factors involved in the response of the complex 3D genome to physical disruptions in health and disease.