Causes and consequences of gene copy number change in adapting yeast populations

适应酵母种群时基因拷贝数变化的原因和后果

• 批准号: 8526477
• 负责人: Maitreya J Dunham
• 金额: $ 24.78万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8526477
• 关键词: Address; Affect; Aneuploidy; Animal Model; Antineoplastic Agents; Autistic Disorder; Biological Assay; Cells; Cellular biology; Chromosomes; Chronic; Collection; Complex; Copy Number Polymorphism; DNA Sequence Rearrangement; Data; Disease; Disease susceptibility; Down Syndrome; Drug resistance; Environment; Eukaryota; Evolution; Freezing; Frequencies; Gene Dosage; Gene Rearrangement; Genes; Genetic Variation; Genome; Genomic Segment; Growth; Health; Hereditary Disease; Human; Human Genetics; Human Genome; Individual; Infection; Learning; Malignant Neoplasms; Measures; Methods; Microbial Drug Resistance; Modeling; Molecular; Mutation; Nutrient; Point Mutation; Population; Population Analysis; Process; Recording of previous events; Role; Saccharomyces cerevisiae; Schizophrenia; Series; Specificity; Spontaneous abortion; System; Testing; Time; Variant; Work; Yeasts; comparative genomic hybridization; dosage; driving force; fitness; genome-wide; improved; next generation sequencing; population survey; pressure; research study; tumorigenesis

DESCRIPTION (provided by applicant): Changes in gene and chromosome copy number are widely observed in systems from cancer to drug resistance to genome evolution. Despite the importance of aneuploidy to many important phenomena related to human health, little work has been done to determine how cells adapt to such extreme changes in gene dosage. Prior work from my lab using the model eukaryote yeast has found that aneuploidy can be detrimental to cells at high growth rates, but can be beneficial to cells growing in challenging environments. In nutrient- limited growth in chemostat culture, specific segments of the genome are reproducibly found amplified and deleted. In some cases, the proximal cause is obvious, such as amplification of nutrient transporter genes, but in others, such as those affecting large genome segments, the driving force remains opaque. The chemostat allows precise control over selection and growth conditions, and, importantly, a complete frozen history of each population, making this system ideal to study the role of aneuploidy in adaptation to strong, narrow selection. I propose to leverage this system to accomplish the follow specific aims: Aim 1: Determine the suite of copy number changes present in experimentally evolved cultures. Using a series of previously performed evolution experiments, we will survey populations for copy number changes using array comparative genomic hybridization and next generation sequencing. Aim 2: Determine the fitness consequences of genome rearrangements. Rearrangements found at high frequency in Aim 1 will be reconstructed and tested for fitness in direct competition assays versus matched ancestral strains. Rearrangements will be cross-tested in multiple selective conditions to query specificity. Results will be compared to evolved strains carrying multiple mutations to determine how much of their fitness benefit is due to genome rearrangements. Aim 3. Dissect the fitness contributions of each gene on the aneuploid chromosomes. To test the contribution of each gene on an aneuploid segment, we will take advantage of strain collections consisting of every yeast gene present at dosages ranging from deletion of a single copy to amplification to many copies. By competing these strains against each other and measuring their abundance via barcode sequencing, we can determine the fitness effect associated with every gene simultaneously. These data will be integrated and compared to the fitness effects of strains from Aim 2. We will also compete aneuploid strains in which each gene is returned to wt copy number to determine which genes are necessary for fitness improvements. This combination of approaches will be the first genome-wide attempt to dissect the precise molecular causes of the fitness changes associated with aneuploidy.

描述（由申请人提供）：从癌症到耐药性再到基因组进化的系统中，广泛观察到基因和染色体拷贝数的变化。尽管非整倍性对于许多与人类健康相关的重要现象很重要，但很少有工作来确定细胞如何适应基因剂量的这种极端变化。我的实验室之前使用模型真核酵母进行的研究发现，非整倍体可能对高生长速率的细胞有害，但可能有利于细胞在充满挑战的环境中生长。在恒化培养物的营养限制生长中，基因组的特定片段可重复地被扩增和删除。在某些情况下，近端原因是明显的，例如营养转运基因的扩增，但在其他情况下，例如影响大基因组片段的情况，驱动力仍然不透明。恒化器可以精确控制选择和生长条件，更重要的是，可以控制每个种群的完整冷冻历史，使该系统成为研究非整倍性在适应强、窄选择中的作用的理想选择。我建议利用该系统来实现以下具体目标： 目标 1：确定实验进化培养物中存在的一套拷贝数变化。通过一系列先前进行的进化实验，我们将使用阵列比较基因组杂交和下一代测序来调查群体的拷贝数变化。目标 2：确定基因组重排的适应性后果。在目标 1 中发现的高频率重排将被重建，并在与匹配的祖先菌株的直接竞争测定中测试其适应性。重排将在多个选择性条件下进行交叉测试以查询特异性。结果将与携带多个突变的进化菌株进行比较，以确定它们的适应性益处有多少是由于基因组重排造成的。目标 3. 剖析非整倍体染色体上每个基因的适应度贡献。为了测试每个基因对非整倍体片段的贡献，我们将利用由每个酵母基因组成的菌株集合，其剂量范围从单个拷贝的删除到扩增到多个拷贝。通过将这些菌株相互竞争并通过条形码测序测量它们的丰度，我们可以同时确定与每个基因相关的适应度效应。这些数据将被整合并与目标 2 中菌株的适应度效果进行比较。我们还将对非整倍体菌株进行竞争，其中每个基因都返回到 wt 拷贝数，以确定哪些基因对于适应度改善是必需的。这种方法的组合将是首次在全基因组范围内尝试剖析与非整倍性相关的适应性变化的精确分子原因。