Functional Understanding of Chromosome Arm Aneuploidies

项目摘要 非整倍体(染色体的获得或丢失)和部分非整倍体(染色体臂的获得或丢失) 长期以来在癌细胞中被观察到，是几种先天性疾病的原因。有趣的是， 下一代测序的进展最近证明了健康人群中存在非整倍体 细胞。这项研究计划的目标是揭示个体非整倍体的下游效应。 我们开发了新的计算算法来从基因组和 转录数据。我们的方法可以从大量的非整倍体细胞中鉴定出小群体 测序数据。此外，健康组织的单细胞测序将允许识别更罕见的 非整倍体事件。通过利用现有的全面努力来表征来自整个 我们将建立一个发生在健康组织中的非整倍体事件的图谱。 为了直接测试人类细胞中非整倍体变化的影响，我开发了一种基因组工程 体外定向缺失染色体臂的方法。我们以前使用这种方法来生成 具有chr3p缺失和不chr3p缺失的同基因细胞系配对，发现chr3p缺失导致细胞周期停滞。 我们的一些细胞系通过复制它们的野生型chr3拷贝来适应chr3p的缺失，转换到一种状态 三个季度的收益。具有chr3q增益的细胞对参与DNA复制的基因表现出不同的依赖性。这里, 我们将使用这些细胞来确定这些细胞周期表型的机制，以及确定哪些 Chr3基因与这些表型有关。此外，我们还将为每条染色体创建一个细胞系文库 手臂切除。这些细胞系对于比较和对比不同的 人类细胞中的非整倍体事件。通过这项工作，我们将坚定地建立一个致力于 了解人类细胞中特定非整倍体的后果。

Project Summary Aneuploidy (gain or loss of a chromosome) and partial aneuploidy (gain or loss of a chromosome arm) have long been observed in cancer cells and are the cause of several congenital diseases. Interestingly, advances in next-generation sequencing have more recently demonstrated the presence of aneuploidy in healthy cells. The goal of this research proposal is to uncover the downstream effects of individual aneuploidies. We developed new computational algorithms to identify aneuploidy events from genomic and transcriptomic data. Our methods allow for identification of small populations of aneuploid cells from bulk sequencing data. In addition, single-cell sequencing of healthy tissues will allow for identification of even rarer aneuploidy events. By leveraging existing comprehensive efforts to characterize single cells from across the human body, we will build an atlas of aneuploidy events occurring in healthy tissue. To directly test the effects of aneuploidy alterations in human cells, I developed a genome engineering approach for targeted deletion of chromosome arms in vitro. We previously used this approach to generate isogenic cell line pairs with and without chr3p deletion and found that chr3p deletion leads to cell cycle arrest. Some of our cell lines adapted to chr3p deletion by duplicating their wildtype copy of chr3, transitioning to a state of chr3q gain. Cells with chr3q gain showed differential dependency on genes involved in DNA replication. Here, we will use these cells to identify the mechanisms of these cell cycle phenotypes, as well as determine which chr3 genes contribute to these phenotypes. In addition, we will create a library of cell lines for each chromosome arm deletion. These cell lines will be critical to compare and contrast the downstream effects of different aneuploidy events in human cells. With this work, we will firmly establish a research program dedicated to understanding the consequences of specific aneuploidies in human cells.