MutSensor System: A Set of Highly Sensitive Mutation Reporters to Dissect Genome Stability in Health and Disease

摘要 我们的细胞经常暴露在导致DNA损伤的诱变剂中，如果不修复，就会导致突变。 在细胞水平上，体细胞组织中突变的逐渐积累推动了癌症的发生和发展。 其他疾病。尽管我们对突变过程的理解取得了重大进展，但许多 根本问题仍然是个谜。人类组织突变的遗传原因是什么？多么 人体内不同类型的细胞是否控制着DNA的损伤和修复？尽管我们有一大堆 在模式生物中控制DNA损伤和修复的途径方面的知识，我们缺乏深入的 对人体组织中这一过程的理解。这对于理解突变是如何导致 包括致癌在内的人类疾病。 在这里，我们将使用一种高度创新的方法来构建突变传感器，一组突变记者来估计 哺乳动物细胞的DNA突变频率比现有的高50倍 方法：研究方法。构建和精确交付这些突变感应器的能力是通过“基因写作和 Boeke实验室开发的“交付”系统。这种新的方法将允许大规模的功能性遗传 以前所未有的规模进行放映。在我们初步数据的基础上，我们的目标是全面识别所有 调节不同人类细胞类型突变频率的基因。为此，我们将利用损失-和 功能增益筛选文库以确定基因扰动对突变频率的影响 单元类型。 这些研究将提供史无前例的控制dna损伤的基因和途径的系统图谱。 以及不同人类细胞类型的突变。此外，我们的新方法将代表一种重要的 为科学界提供的资产，可在各种领域有多种生物医学应用 提供了一种简单的工具来研究在健康和疾病状态下控制突变的遗传因素， 包括(但不限于)致癌。

Summary Our cells are constantly exposed to mutagens that cause DNA damage, which if left unrepaired, cause mutations. At the cellular level, the progressive accumulation of mutations in somatic tissues drives carcinogenesis and other diseases. Despite significant advances in our understanding of mutational processes, the answer to many fundamental questions is still a mystery. What are the genetic causes of mutations in human tissues? How do different cell types in the human body control DNA damage and repair? Despite our large body of knowledge in the pathways that control DNA damage and repair in model organisms, we lack a deep understanding of this process in human tissues. This would be critical to understand how mutations lead to human diseases including carcinogenesis. Here we will use a highly innovative approach to build MutSensor, a set of mutation reporters to estimate DNA mutation frequency in mammalian cells with a sensitivity >50-fold higher than that of existing methods. The ability to build and precision deliver these MutSensors is enabled by “Genetic Writing and Delivery” system developed in the Boeke lab. This novel method will allow large-scale functional genetic screening at an unprecedented scale. Building on our preliminary data, we aim at comprehensively identify all genes that regulate mutation frequency in different human cell types. To this purpose, we will utilize loss- and gain-of-function screening libraries to determine the effect of genetic perturbations on mutation frequency across cell types. These studies will provide an unprecedented systematic map of genes and pathways controlling DNA damage and mutations across different human cell types. In addition, our novel methodology will represent an important asset for the scientific community that can have several biomedical applications in a variety of fields providing an easy tool to study genetic factors that control mutagenesis in health and disease states, including (but not limited to) carcinogenesis.