Spatial single-cell analysis of somatic mutation in human brain during aging and neurodegeneration

衰老和神经退行性变过程中人脑体细胞突变的空间单细胞分析

• 批准号: 10687449
• 负责人: Michael Anthony Lodato
• 金额: $ 150.75万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-15 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10687449
• 关键词: Acceleration; Age; Aging; Alzheimer' s Disease; Alzheimer' s disease brain; Alzheimer' s disease related dementia; Amyloid beta-Protein; Award; Brain; Cells; Cockayne Syndrome; DNA Damage; DNA Repair Gene; DNA analysis; DNA sequencing; Disease; Genes; Genetic; Genome; Genomics; Goals; Human; Human Development; Human body; Malignant Neoplasms; Methods; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neurofibrillary Tangles; Neurons; Noise; Parkinson Disease; Pathologic; Pathway interactions; Pattern; Process; Resolution; Skeletal Muscle; Somatic Mutation; Testing; Work; Xeroderma Pigmentosum; age related; age related neurodegeneration; assault; brain cell; disease-causing mutation; early onset; experience; experimental study; genome integrity; genome sequencing; human disease; misfolded protein; neuron loss; normal aging; oxidative DNA damage; postmitotic; presenilin-1; presenilin-2; protein aggregation; repaired; single cell analysis; tau Proteins; tau aggregation; whole genome

Alzheimer’s disease and related dementias display an age-related onset, misfolded protein aggregates such as β-amyloid and tau tangles, increased oxidative DNA damage, and ultimately neuron cell death. Mendelian progeroid diseases caused by mutations in DNA repair genes show early-onset neurodegeneration resembling Alzheimer’s, suggesting that compromised genomic integrity can accelerate aging and directly cause neuron loss. DNA damage can be repaired but may result in permanent changes to the genome called somatic mutations, leading to the hypothesis that increased somatic mutation burden may be common across age-related neurodegenerative disorders. Direct support for this hypothesis has remained elusive in the Alzheimer’s disease brain because standard DNA-sequencing experiments are underpowered to characterize somatic mutations that occur in postmitotic human neurons comprehensively. Such experiments are underpowered because a mutation arising in a postmitotic neuron would be unique to only the single cell in which it occurred and thus it would be indistinguishable from background noise when analyzing DNA isolated from millions of brain cells. We recently developed methods to study somatic mutations in human neurons at single-cell resolution, including methods for single-cell, whole genome sequencing (scWGS). We used scWGS to show that permanent somatic mutations accompany aging in human neurons, with specific mutation signatures nominating discrete pathways generating DNA damage in the human brain. Mutation counts and signatures differ in neurons from donors with genetic early-onset neurodegenerative diseases, specifically Cockayne syndrome and Xeroderma Pigmentosum, and in the late-onset sporadic Alzheimer’s donors, suggesting neurodegeneration is associated with specific patterns of somatic mutation. We found no evidence for mutational hotspots in known Alzheimer’s genes such as APOE, PSEN1, PSEN2, or APP, instead finding that somatic mutations in Alzheimer’s neurons represent a stochastic assault on the genome in each cell. The goal of this New Innovator Award proposal is to develop and apply new scWGS methods to study somatic mutations during neurodegeneration in unprecedented detail. We will focus on neurons from late-stage Alzheimer’s disease donors and from donors with late-stage Parkinson’s disease, another age-associated neurodegenerative disorder characterized by increased DNA damage, aggregates of misfolded protein, and neuron cell death. We will focus on those neurons with known pathological hallmarks of these diseases, for example, β-amyloid and tau aggregates, to test the hypothesis that these neurons experience increased DNA damage and thus increased somatic mutations. This work will have broad impacts in the fields of single-cell genomics, aging, neurodegeneration, and human development.

阿尔茨海默病和相关痴呆症的发病与年龄相关，错误折叠的蛋白质聚集体（例如 β-淀粉样蛋白和 tau 蛋白缠结）、DNA 氧化损伤增加，以及由 DNA 修复基因突变引起的最终神经元细胞死亡，显示出早发性神经变性。重新组装阿尔茨海默氏症，表明基因组完整性受损会加速衰老并直接导致神经元损失，DNA 损伤可以修复，但可能会导致基因组发生永久性变化，称为体细胞突变。导致体细胞突变负担增加可能在与年龄相关的神经退行性疾病中很常见的假设。在阿尔茨海默病大脑中，这一假设的直接支持仍然难以捉摸，因为标准的 DNA 测序实验不足以表征有丝分裂后人类神经元中发生的体细胞突变。综合来看，这样的实验效果不佳，因为有丝分裂后神经元中产生的突变仅对发生该突变的单个细胞来说是独特的，因此在分析从数百万个脑细胞中分离的 DNA 时，它与背景噪音无法区分。 我们最近开发了以单细胞分辨率研究人类神经元体细胞突变的方法，包括单细胞全基因组测序 (scWGS) 方法。我们使用 scWGS 来证明，永久性体细胞突变伴随着人类神经元的衰老，并具有特定的突变特征。提名在人脑中产生 DNA 损伤的离散途径 患有遗传性早发性神经退行性疾病（特别是科凯恩综合征和色素性干皮病）的捐赠者的神经元和晚发性神经退行性疾病的神经元中的突变计数和特征有所不同。散发性阿尔茨海默氏症供体，表明神经变性与特定的体细胞突变模式有关。我们没有发现已知阿尔茨海默氏症基因（如 APOE、PSEN1、PSEN2 或 APP）中存在突变热点的证据，而是发现阿尔茨海默氏症神经元中的体细胞突变代表了对体细胞突变的随机攻击。每个细胞中的基因组。 这项新创新者奖提案的目标是开发和应用新的 scWGS 方法，以前所未有的细节研究神经退行性疾病期间的体细胞突变，我们将重点关注来自晚期阿尔茨海默病供体和晚期帕金森病供体的神经元，这是另一个时代。相关的神经退行性疾病，其特征是 DNA 损伤增加、错误折叠蛋白聚集和神经元细胞死亡。我们将重点关注具有这些疾病已知病理特征的神经元，例如 β-淀粉样蛋白和 tau 蛋白。聚合体，以检验这些神经元经历增加的 DNA 损伤，从而增加体细胞突变的假设，这项工作将对单细胞基因组学、衰老、神经退行性疾病和人类发育领域产生广泛影响。