Constraints and Consequences of Copy Number Variation

拷贝数变异的限制和后果

• 批准号: 9973827
• 负责人: David Gresham
• 金额: $ 30.6万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9973827
• 关键词: Address; Alleles; Bar Codes; Biological Assay; Cells; Collection; Color; Complex; Copy Number Polymorphism; Costs and Benefits; Coupling; DNA; DNA Sequence; Data; Disease; Environment; Evolution; Fluorescence; Gene Dosage; Gene Duplication; Gene Expression; Gene Family; Gene Proteins; Generations; Genes; Genetic Variation; Genome; Genomic Segment; Genomics; Goals; Growth; Health; Heterogeneity; Human; Individual; Link; Maintenance; Methods; Modification; Molecular; Phenotype; Ploidies; Population; Process; Property; Proteins; Reporter; Resolution; Role; Saccharomyces cerevisiae; Saccharomycetales; Study models; System; Testing; Variant; Yeast Model System; Yeasts; cost; experience; fitness; genetic variant; genome sequencing; human disease; insight; microbial; molecular phenotype; novel; novel strategies; prevent; response; single-cell RNA sequencing; tumorigenesis; variant detection; whole genome

Project Summary/Abstract Copy number variants (CNVs) – increases and decreases in the number of copies of genomic regions – are a pervasive class of genetic variation that contribute to rapid adaptive evolution and human phenotypic variation and disease. Moreover, CNVs are the first step in processes that result in genome evolution such as gene family expansion and the generation of novel functions through modification of duplicate genes. Despite the importance of CNVs in human health and evolution, many fundamental questions about their role in evolving populations and their functional consequences remain unsolved. Experimental microbial evolution in chemostats is an ideal system for studying the evolutionary constraints and functional consequences of CNVs. To gain quantitative insights into the role of CNVs in evolving populations, we have developed a novel CNV reporter system using a constitutively expressed fluorescent protein gene linked to a locus at which CNVs are known to be repeatedly generated and selected. By coupling this system to a lineage tracking system using random molecular barcodes we have developed a powerful novel framework for studying the diversity and dynamics of CNVs in complex populations. We will use this novel system to address three fundamental questions. In aim 1, we will address the dynamics with which CNVs are selected in fluctuating environments. Using a combination of two-color CNV reporters and lineage tracking we will investigate the dynamics and diversity of CNV selection in variable chemostat environments. In aim 2, we will identify the determinants of CNV fitness effects. We will establish a collection of hundreds of unique CNV alleles that are molecularly characterized using whole genome sequencing and uniquely marked by DNA barcodes. We will use pooled fitness assays in a range of environmental conditions to quantify the fitness of each lineage and use these data to identify determinants of condition dependent and independent fitness fitness costs and benefits of CNVs. In aim 3, we will test the effect of CNVs on variability of gene expression and phenotypes. Using single cell RNA sequencing we will test whether CNVs results in increased variation in expression levels of both the genes contained within a CNV and all genes throughout the genome. We will also test whether CNVs result in increased variability in phenotypes using a high throughput microcolony growth rate assay. Our study will make use of the unparalleled efficiency and rigor afforded by the budding yeast model system and the unprecedented resolution provided by our new method for CNV detection to address fundamental questions of widespread significance for our understanding of CNVs in evolution and disease. As CNVs are universal in evolution and disease findings from our study will have a high impact on the field of genomics, evolution, and human health.

项目摘要/摘要 拷贝数变异体(CNV)-基因组区域拷贝数的增加和减少-是一种 广泛存在的一类有助于快速适应进化和人类表型变异的遗传变异 和疾病。此外，CNV是导致基因组进化的第一步，如基因 家族扩张和通过修改重复基因产生新的功能。尽管 CNV在人类健康和进化中的重要性，以及关于它们在进化中的作用的许多基本问题 人口及其功能后果仍未解决。实验微生物进化在 趋化因子是研究CNV的进化限制和功能后果的理想系统。 为了定量了解CNV在进化人口中的作用，我们开发了一种新的CNV 使用结构性表达的荧光蛋白基因的报告系统，该基因连接到CNV所在的位置 已知是重复生成和选择的。通过将该系统耦合到使用 随机分子条形码我们已经开发了一个强大的新框架来研究多样性和 复杂人群中CNV的动态变化。我们将使用这个新的系统来解决三个基本问题 问题。在目标1中，我们将解决在波动环境中选择CNV的动态。 使用双色CNV记者和血统追踪相结合的方法，我们将调查动态和 在不同的恒化器环境中CNV选择的多样性。在目标2中，我们将确定以下决定因素 CNV健身效果。我们将建立数百个独特的CNV等位基因的集合，这些等位基因是分子上的 使用全基因组测序并由DNA条形码唯一标记的。我们将使用池化 在一系列环境条件下进行适合度分析，以量化每个谱系的适合度，并使用这些数据 确定条件依赖和独立健身的决定因素，以及CNV的健身成本和收益。在……里面 目的：检测CNV对基因表达和表型变异的影响。使用单细胞RNA 我们将测试CNV是否导致这两个基因表达水平的增加 包含在CNV和整个基因组中的所有基因中。我们还将测试CNV是否会导致 使用高通量微集落生长率分析增加表型的可变性。我们的研究将 利用发芽酵母模型系统所提供的无与伦比的效率和严谨性，以及 我们的CNV检测新方法提供了前所未有的分辨率，以解决以下基本问题 对于我们理解CNV在进化和疾病中的作用具有广泛的意义。由于CNV在 我们研究的进化论和疾病发现将对基因组学、进化论和 人类健康。