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.

项目概要/摘要 拷贝数变异（CNV）——基因组区域拷贝数的增加和减少——是 有助于快速适应性进化和人类表型变异的普遍遗传变异 此外，CNV 是基因组进化过程的第一步。 尽管如此，家族扩张和通过修改重复基因产生新功能。 CNV 在人类健康和进化中的重要性，关于它们在进化中的作用的许多基本问题 种群及其功能后果仍未解决。 恒化器是研究 CNV 的进化限制和功能后果的理想系统。 为了定量了解 CNV 在进化人群中的作用，我们开发了一种新型 CNV 使用与 CNV 所在基因座相连的组成型表达荧光蛋白基因的报告系统 通过将该系统耦合到沿袭跟踪系统，可以重复生成和选择。 随机分子条形码我们开发了一个强大的新颖框架来研究多样性和 我们将使用这个新颖的系统来解决复杂群体中 CNV 的动态。 在目标 1 中，我们将解决在波动环境中选择 CNV 的动态问题。 结合使用双色 CNV 生产者和谱系追踪，我们将研究动态和 在目标 2 中，我们将确定可变恒化器环境中 CNV 选择的多样性。 我们将建立数百个分子上独特的 CNV 等位基因的集合。 我们将使用全基因组测序进行表征并通过 DNA 条形码进行独特标记。 在一系列环境条件下进行适应性测定，以量化每个谱系的适应性并使用这些数据 确定条件依赖性和独立健身成本和 CNV 收益的决定因素。 目标 3，我们将使用单细胞 RNA 测试 CNV 对基因表达和表型变异的影响。 测序后，我们将测试 CNV 是否会导致两个基因的表达水平变异增加 我们还将测试 CNV 是否会导致 CNV 和整个基因组中的所有基因。 我们的研究将使用高通量微菌落生长速率测定来增加表型的变异性。 利用芽殖酵母模型系统和 我们的 CNV 检测新方法提供了前所未有的分辨率，可解决以下基本问题： 由于 CNV 在进化和疾病中具有普遍性，因此对于我们理解 CNV 具有广泛的意义。 我们研究的进化和疾病发现将对基因组学、进化和疾病领域产生重大影响 人类健康。