Impact of DNA double-strand breaks on 3D genome organization and genome stability in Alzheimer’s disease

DNA 双链断裂对阿尔茨海默病 3D 基因组组织和基因组稳定性的影响

• 批准号: 10282373
• 负责人: Vishnu Dileep
• 金额: $ 13.67万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-15 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10282373
• 关键词: 3-Dimensional; ATAC-seq; Adopted; Advisory Committees; Aging; Alzheimer' s Disease; Alzheimer' s disease brain; Alzheimer' s disease model; Alzheimer' s disease risk; Autopsy; Biological Assay; Biology; Brain; Cells; ChIP-seq; Chromatin; Chromosomal translocation; Coculture Techniques; Complex; DNA; DNA Binding; DNA Damage; DNA Double Strand Break; Data; Development; Disease; Frequencies; Gene Expression; Genome; Genome Stability; Genomic Instability; Genomics; Goals; Hi-C; Human; Hyperactivity; Immediate-Early Genes; Induced pluripotent stem cell derived neurons; Knockout Mice; Label; Lead; Location; Measures; Mediating; Mentors; Mentorship; Microglia; Mission; Modeling; Molecular; Mosaicism; Mus; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neuroglia; Neuronal Dysfunction; Neurons; Nonhomologous DNA End Joining; Oligodendroglia; Orthologous Gene; Partner in relationship; Pathogenesis; Pathologic; Pathway interactions; Pattern; Pharmacology; Phase; Play; Public Health; Publishing; Recurrence; Reporting; Role; Structure; Tauopathies; Technology; Testing; Therapeutic Intervention; Training; Work; aging brain; cell type; chromosome conformation capture; cohesin; computational pipelines; experience; familial Alzheimer disease; improved; in situ sequencing; induced pluripotent stem cell; induced pluripotent stem cell technology; mouse model; nervous system disorder; novel; novel therapeutics; presenilin-1; prevent; promoter; response; single cell analysis; single-cell RNA sequencing; therapeutic evaluation; tool; transcription factor; transcriptome

PROJECT SUMMARY/ABSTRACT The accumulation of DNA Double-Strand Breaks (DSBs) in neurons is an early hallmark of Alzheimer’s disease (AD). Increased DSBs are also associated with aging, which is the largest risk factor for AD. AD is also a complex disease involving all major brain glial cell types. Thus, there is a critical need to understand the molecular mechanisms of DSB induced changes in both neurons and glia. The structural stability of the genome is paramount in maintaining a functional genome, and recently, the 3D organization of the genome has emerged as a major regulator of genome function. My overall hypothesis is that 3D genome reorganization and structural genome instability mediated by DSBs are principle drivers of AD pathogenesis and brain aging. My objective is to determine how and to what extent DSBs within the neurons impact genome organization and the glial response, with the goal of identifying molecular pathways that can be targeted as novel therapies for preventing or halting the progression of neurodegeneration. I will use mouse models that recapitulate the pre-symptomatic accumulation of DSBs in neurons and human iPSC models to determine the degree of the 3D genome disruption caused due to DSBs and the underlying molecular mechanisms. I will also identify the consequences of neuronal DSBs on the structural stability of the genome by measuring the frequency of DSB mediated chromosomal translocations in human AD neurons. Interestingly, normal neuronal activity causes DSBs at immediate early gene promoters. Also, neuronal hyperactivity has been reported in AD. I will test if IEGs are locations of frequent chromosomal translocations after neuronal hyperactivity induced in neuronal culture. Previous studies have shown that microglia transitions to a reactive state in response to neurodegeneration. To understand the role of the 3D genome organization in this microglia transition, I will use single cell Hi-C to measure the unique chromatin interactions that mediate the reactive microglia state. Subsequently, transcription factor predictions using integrative analysis of single cell Hi-C will be tested in iPSC derived microglia-neuron co-cultures for their potential to modulate the reactive microglia response. This approach will be extended to study oligodendrocyte response in the independent phase. I will work with my mentor Dr. Li-Huei Tsai, my mentorship committee, Dr. Bruce Yankner and Dr. Manolis Kellis, my technical support and advisory committee, Dr. Frederick Alt and Dr. Peter Fraser to carry out my proposed training plan. I will gain experience in iPSC technology and differentiation from the Tsai lab to modulate the disease associated microglia response to neuronal DSBs and work with Dr. Manolis Kellis to implement the computational pipelines for the transcription factor predictions. To bridge the gap in my training in the biology of AD and brain aging, I will audit relevant courses and receive additional mentoring from Dr. Bruce Yankner.

项目概要/摘要 神经元中 DNA 双链断裂 (DSB) 的积累是阿尔茨海默病的早期标志 （广告）。 DSB 增加还与衰老有关，而衰老是 AD 的最大风险因素。 AD也是一个复杂的 涉及所有主要脑神经胶质细胞类型的疾病。因此，迫切需要了解分子 DSB 诱导神经元和神经胶质细胞发生变化的机制。基因组的结构稳定性为 对于维持功能基因组至关重要，最近，基因组的 3D 组织已经出现 作为基因组功能的主要调节因子。我的总体假设是 3D 基因组重组和结构 DSB 介导的基因组不稳定性是 AD 发病机制和大脑衰老的主要驱动因素。我的目标是 确定神经元内的 DSB 如何以及在何种程度上影响基因组组织和神经胶质细胞 反应，目标是确定可作为预防新疗法的分子途径 或阻止神经退行性变的进展。我将使用重现症状前的小鼠模型 DSB 在神经元和人类 iPSC 模型中的积累，以确定 3D 基因组破坏的程度 由 DSB 和潜在的分子机制引起。我还将确定神经元的后果 通过测量 DSB 介导的染色体频率来观察 DSB 对基因组结构稳定性的影响 人类 AD 神经元的易位。有趣的是，正常的神经元活动会在早期立即引起 DSB 基因启动子。此外，AD 中也有神经元过度活跃的报道。我将测试 IEG 是否是频繁出现的位置 神经元培养中诱导神经元过度活跃后的染色体易位。 先前的研究表明，小胶质细胞会转变为反应状态以响应 神经变性。为了了解 3D 基因组组织在小胶质细胞转变中的作用，我将使用 单细胞 Hi-C 来测量介导反应性小胶质细胞状态的独特染色质相互作用。 随后，将在 iPSC 中测试使用单细胞 Hi-C 综合分析的转录因子预测 衍生的小胶质细胞-神经元共培养物具有调节反应性小胶质细胞反应的潜力。这 该方法将扩展到研究独立阶段少突胶质细胞的反应。 我将与我的导师 Li-Huei Tsai 博士、我的导师委员会 Bruce Yankner 博士和 Manolis 博士一起工作 凯利斯 (Kellis)、我的技术支持和咨询委员会、弗雷德里克·阿尔特 (Frederick Alt) 博士和彼得·弗雷泽 (Peter Fraser) 博士来执行我的 拟议的培训计划。我将从 Tsai 实验室获得 iPSC 技术和分化方面的经验来进行调节 与疾病相关的小胶质细胞对神经元 DSB 的反应，并与 Manolis Kellis 博士合作实施 转录因子预测的计算管道。为了弥补我在生物学训练中的差距 AD 和大脑老化，我将旁听相关课程并接受 Bruce Yankner 博士的额外指导。