Chromatin mobility in response to DNA damage

DNA 损伤时的染色质迁移率

• 批准号: 10242769
• 负责人: Keith D Bonin
• 金额: $ 60.13万
• 依托单位: WAKE FOREST UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-19 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10242769
• 关键词: 3-Dimensional; Acridines; Address; Affect; Affinity; Automobile Driving; Biogenesis; Biological Assay; Biometry; Biophysics; Cancer Patient; Cell Nucleus; Cell model; Cells; Cellular Assay; Cellular biology; Chemistry; Chemotherapy and/or radiation; Chromatin; Chromosomal Breaks; Chromosomes; Clinical; Communities; DNA; DNA Damage; DNA Repair; DNA Sequence Rearrangement; Data; Diffusion; Dimensions; Disease; Dyes; Dysmyelopoietic Syndromes; Elements; Engineering; Environment; Etiology; Exposure to; Freezing; Frequencies; Genome; Genomic Instability; Genomics; Goals; Hematopoietic stem cells; Image; Image Analysis; Kinetics; Knowledge; Label; Length; Light; Link; Lymphoma; Maintenance; Malignant Neoplasms; Maps; Measurement; Mediating; Methodology; Methods; Modeling; Motion; Movement; Myeloproliferative disease; Nuclear; Nuclear Structure; Oncology; Optical Methods; Optics; Outcome; Patients; Pharmacology; Physical condensation; Polymers; Probability; Protocols documentation; Recurrence; Reporter; Research; Resolution; Resources; Rhodamine; Risk; Role; Sampling; Scanning; Second Primary Cancers; Survivors; System; Testing; Therapeutic; Three-Dimensional Imaging; Time; Tissues; Translations; base; biophysical model; cancer initiation; cancer therapy; clinically relevant; genetic approach; genomic aberrations; genomic predictors; imaging system; improved; in silico; insight; inter-individual variation; leukemia; mechanical properties; mortality; novel; novel strategies; prevent; protein distribution; response; therapy development; translational study; treatment choice

Project Summary Genomic translocations are well-established drivers of therapy-related myeloid neoplasms (t-MN) that affect survivors of primary malignancies. t-MN accounts for 10-20% of all myeloid malignancies and have very poor clinical outcomes. It is not possible to predict which patients treated for a primary cancer will develop t-MN, which constitutes a major clinical challenge. A method to assess the risk of translocations after patient exposures to DNA damaging chemotherapy and radiations would inform therapeutic decisions. Translocations depend on the movements of broken DNA ends on non-homologous chromosomes. We developed a method based on diffractive optical elements (DOE) to track photoactivated chromatin reporters (PACR) and map chromatin motions in the cell nucleus. Our preliminary data show that chromatin movements transiently decrease in response to DNA damage, which led to the hypothesis that chromatin `freezes' to facilitate the initial steps of the DNA damage response and reduce frequencies of genomic rearrangements. This effect may be mediated by reversible chromatin compaction and by chromatin interactions with structural nuclear elements. Inter-individual variability may determine genomic aberration frequencies and t-MN risk. The following specific aims will expand PACR assays and test this hypothesis: In Aim 1, methodology will be developed for 3D measurements of chromatin mobility, in order to improve tracking accuracy and to account for inhomogeneities of the nuclear environment in all three dimensions. Rapid light sheet imaging will be used, and a novel multifocus 3D system based on a rotating point spread function DOE will be built. New photoactivatable DNA dyes will be developed to expand the PACR approach to difficult-to-transfect cells and tissues. The goals for Aim 2 are to identify the mechanisms controlling chromatin motions after DNA damage and to assess the functional consequences for DNA repair and translocations of chromatin `freezing'. We will feed PACR measurements with high time resolution (before/during/after DNA damage) into biophysical polymer models to predict physical changes of chromatin (intramolecular forces, persistence length, etc.), then test the predictions using multidimensional maps of chromatin motions, compaction, and nucleoskeletal organization, as well as functional cell assays. Two approaches to alter chromatin diffusion will assess causality between chromatin motions, DNA repair, and genomic translocations. In Aim 3, we will use hematopoietic stem/progenitor cell samples from t-MN patients and healthy donors to test for clinically relevant associations between chromatin motions and genomic translocations. Characterizing the physical origins of genomic translocations may yield new methods to predict, and new targets to prevent, genomic rearrangements driving cancer initiation. Beyond the proposed research focused on chromatin motions during DNA repair, we anticipate broad applicability of our novel 3D imaging resources to study chromatin in multiple contexts.

项目摘要 基因组易位是治疗相关性骨髓肿瘤（t-MN）的公认驱动因素， 原发性恶性肿瘤的幸存者。t-MN占所有骨髓恶性肿瘤的10-20%，并且具有非常差的 临床结果。不可能预测哪些接受原发性癌症治疗的患者将发展t-MN， 这构成了主要的临床挑战。评估患者术后易位风险的方法 暴露于DNA损伤的化疗和辐射将为治疗决策提供信息。易位 依赖于非同源染色体上断裂DNA末端的运动。我们开发了一种方法 基于衍射光学元件（DOE）跟踪光活化染色质报告子（PACR）并映射 细胞核中染色质的运动。我们的初步数据显示， 对DNA损伤的反应减少，这导致了染色质“冻结”以促进细胞凋亡的假设。 DNA损伤反应的初始步骤，并减少基因组重排的频率。这种作用可能 由可逆染色质压缩和染色质与结构核的相互作用介导 元素个体间变异性可能决定基因组畸变频率和t-MN风险。的 以下具体目标将扩展PACR测定并检验这一假设：在目标1中，方法学将是 开发用于染色质迁移率的3D测量，以提高跟踪精度并考虑 核环境在所有三个方面的不均匀性。将使用快速光片成像， 建立了一种基于旋转点扩散函数DOE的多聚焦三维光学系统。新 将开发可光活化的DNA染料，以将PACR方法扩展到难以转染的细胞， 组织中目的2的目标是确定DNA损伤后控制染色质运动的机制 并评估DNA修复和染色质“冻结”易位的功能后果。我们将 将具有高时间分辨率的PACR测量（DNA损伤之前/期间/之后）馈送到生物物理 聚合物模型预测染色质的物理变化（分子内力、持续长度等），然后 使用染色质运动、压实和核骨架的多维图来测试预测 组织，以及功能性细胞测定。两种改变染色质扩散的方法将评估 染色质运动、DNA修复和基因组易位之间的因果关系。在目标3中，我们将使用 来自t-MN患者和健康供体的造血干/祖细胞样品，以测试临床相关的 染色质运动和基因组易位之间的关联。描述了 基因组易位可能会产生新的方法来预测，和新的目标，以防止基因组， 基因重排驱动癌症的发生除了拟议的研究集中在染色质运动过程中 DNA修复，我们预计我们的新的3D成像资源的广泛适用性，以研究染色质在多个 contexts.