Genome-wide investigation of somatic mutation in the developing and aging brain

发育和衰老大脑体细胞突变的全基因组研究

基本信息

批准号：
8762213
负责人：
Kristin Kay Baldwin
金额：
$ 75.29万
依托单位：
SCRIPPS RESEARCH INSTITUTE, THE
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-08-01 至 2019-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8762213
关键词：
Address Adult Age Aging Autistic Disorder Awareness Bioinformatics Brain Cancer Model Cell Aging Cell Line Cell Lineage Cell Nucleus Cells Clonal Expansion Cloning Complex Copy Number Polymorphism DNA Insertion Elements DNA Sequence Rearrangement DNA amplification Detection Development Disease Equilibrium Event Exhibits Fertilization Fibroblasts Fluorescence Future Genes Genetic Genome Genome Mappings Genomics Germ-Line Mutation Health Hippocampus (Brain)Human Individual Inherited Intellectual functioning disability Investigation Knowledge Link Loss of Heterozygosity Malignant Neoplasms Mammals Maps Methods Micromanipulation Mitotic Morphologic artifacts Mosaicism Mus Mutation Mutation Spectra Neonatal Neurons New York Nucleotides Oncogenic Oocytes Pattern Play Point Mutation Population Prevalence Process Publications Publishing Recurrence Reporting Research Resolution Role Schizophrenia Science Sister Solutions Somatic Cell Somatic Mutation Sorting - Cell Movement Stem cells Surveys Technology Time Tissues Tube Validation Variant Work aged aging brain base cell age cell type cost effective design embryo cell frontier genome sequencing genome-wide human disease innovation insight interest nervous system disorder nuclear transfer olfactory bulb public health relevance rare variant relating to nervous system self renewing cell somatic cell nuclear transfer

项目摘要

DESCRIPTION (provided by applicant): Mutations that arise after fertilization in somatic cell lineages are linked to cancer and aging an have been shown to contribute to an increasing number of human disorders. Similarly, de novo germline mutations in neuronal genes are responsible for cases of autism, schizophrenia and intellectual disability, suggesting that similar types of somatic mutations could contribute to these and other neurological disorders by providing large-effect mutations in specific cell types, that may act alone or in concert with inherited variants. Despite the growing awareness of the importance of genomic mosaicism for human health, our present understanding of somatic mutation in different cellular lineages of the body and brain is minimal. This question has been difficult to address because conventional genome-wide methods cannot detect variants that are rare within a cell population, and most tissues are composed of diverse cell types and intermixed lineages. Single cell sequencing offers one solution, however, current methods suffer from high error-rates and low resolution, and do not allow for independent validation of mutations detected in merely one cell. A second means to amplify genomes from single cells is through clonal expansion. This is feasible for cancer and some self-renewing cell types, but not for many interesting or aged cell types such as post-mitotic neurons. Here, we propose to use two innovative strategies to profile genomes from individual neurons and control fibroblasts from young and aged mice. First, we take advantage of the only known method to amplify neuronal cells without use of oncogenic factors: cloning by somatic cell nuclear transfer. This will enable deep whole genome sequencing and comprehensive mutational profiling of neuron-derived cell lines. Second, we will use nuclear transfer to produce pairs of sister cells derived from single neurons after one division. Single cell sequencing of replicate sister cells will enable sensitive and accurate detection of de novo copy number variation in a context where bona-fide mutations can be clearly distinguished from DNA amplification artifacts. Our application of these complementary technologies will reveal the full spectrum of genome variation that arises in different cell lineages during development and aging, and will help resolve longstanding hypotheses regarding the extent, impact and origins of neuronal genome diversity.

描述（由申请人提供）：体细胞谱系受精后出现的突变与癌症和衰老有关，并已被证明会导致越来越多的人类疾病。同样，神经元基因中的从头种系突变是导致自闭症、精神分裂症和智力障碍的原因，这表明类似类型的体细胞突变可能通过在特定细胞类型中提供大效应突变而导致这些和其他神经系统疾病，这可能会起作用单独或与遗传变异一起。尽管人们越来越认识到基因组嵌合对人类健康的重要性，但我们目前对身体和大脑不同细胞谱系的体细胞突变的了解还很少。这个问题很难解决，因为传统的全基因组方法无法检测细胞群中罕见的变异，而且大多数组织由不同的细胞类型和混合谱系组成。单细胞测序提供了一种解决方案，然而，目前的方法存在高错误率和低分辨率，并且不允许对仅在一个细胞中检测到的突变进行独立验证。从单细胞扩增基因组的第二种方法是通过克隆扩增。这对于癌症和一些自我更新的细胞类型是可行的，但对于许多有趣或老化的细胞类型（例如有丝分裂后神经元）来说并不可行。在这里，我们建议使用两种创新策略来分析单个神经元的基因组并控制年轻和老年小鼠的成纤维细胞。首先，我们利用唯一已知的方法来扩增神经元细胞而不使用致癌因子：通过体细胞核移植进行克隆。这将使深度全基因组测序和神经元衍生细胞系的全面突变分析成为可能。其次，我们将使用核移植来产生来自一次分裂后的单个神经元的姐妹细胞对。复制姐妹细胞的单细胞测序将能够灵敏、准确地检测从头拷贝数变异，在这种情况下，可以清楚地区分真正的突变与 DNA 扩增伪影。我们对这些互补技术的应用将揭示不同细胞谱系在发育和衰老过程中出现的全基因组变异，并将有助于解决有关神经元基因组多样性的范围、影响和起源的长期假设。