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）：在从癌症到耐药性到基因组进化的系统中，基因和染色体拷贝数的变化被广泛观察到。尽管非整倍性对许多与人类健康有关的重要现象的重要性很重要，但几乎没有做出的工作来确定细胞如何适应基因剂量的这种极端变化。我的实验室的先前使用真核生物酵母的工作发现，非整倍性可能会以高生长速率对细胞有害，但可以对在有挑战性的环境中生长有益。在化学抑制培养物中的营养有限生长中，基因组的特定片段被重复地发现和删除。在某些情况下，近端原因是显而易见的，例如营养转运蛋白基因的扩增，但在其他情况下，例如影响大基因组段的养分，驱动力仍然不透明。 Chemostat允许精确控制选择和生长条件，并且重要的是，每个人群的完整冻结历史，使该系统理想地研究了非整倍性在适应强，狭窄的选择中的作用。我建议利用该系统来完成以下特定目的：目标1：确定实验演化培养物中存在的拷贝数变化的套件。使用一系列先前执行的进化实验，我们将使用阵列比较基因组杂交和下一代测序对拷贝数更改进行调查。目标2：确定基因组重排的适应性后果。在AIM 1中发现的高频重排将进行重建并测试直接竞争测定中的健身与匹配的祖先菌株。重排将在多个选择性条件下进行跨测试，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，以确定其适应性益处是由于基因组重排。 AIM 3。剖析每个基因在非整倍体染色体上的适应性贡献。为了测试每个基因在非整倍体段上的贡献，我们将利用由剂量收集组成的菌株收集，从单个副本的缺失到放大到许多副本的每个剂量的酵母基因组成。通过相互竞争这些菌株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性效应。这些数据将被整合并与AIM 2的菌株的适应性效应进行比较。我们还将竞争非倍体菌株，其中每个基因都返回WT拷贝数，以确定适应性改善所需的哪些基因。这种方法的组合将是全基因组的第一次尝试，以剖析与非整倍性相关的适应性变化的精确分子原因。