Stanford Center for Connecting DNA Variants to Function and Phenotype

项目总结 全基因组关联研究现已发现数以万计的非编码变异与 人类疾病和特征。事实证明，要解释这些关联是具有挑战性的。大多数因果变异 存在于非编码基因组中，似乎影响控制基因逻辑的DNA顺式调控元件 表达，并可能为我们指明新的细胞类型、基因和疾病的途径。然而，我们一直缺乏 系统地描述这些顺式调控变体和元件如何影响基因组所需的工具 功能和表型。 我们在斯坦福大学的团队现在已经开发出创新的单细胞、CRISPR映射和计算 能够识别数以千计的元素和变体并在功能上表征它们的技术 直接在人类基因组中。这些工具包括用于在单元格中识别候选元素的单元格atac-seq 和组织；连接数千种元素和变种对基因影响的灵敏CRISPR平铺方法 表达和细胞表型；以及预测疾病变异如何调节的ABC和BPNet模型 基因表达。总而言之，这些技术提出了一种系统地连接DNA变体的新策略 以及与功能和表型有关的元素。 在这里，我们将应用这些新技术，与NHGRI的基因组变异对 功能联盟。我们将使用来自人类多能干细胞的四种心血管细胞类型作为 模型系统。首先，我们将利用心脏分化和发育的单细胞图谱来选择 成人和儿童心脏病的因素和风险变量可能控制心血管细胞功能。 其次，我们将应用单细胞CRISPR工具来测量数千个无偏元素和 基因表达的变异，并将优先处理的疾病变异与靶基因、细胞表型和 组织表型。第三，我们将利用这些实验数据集来校准和改进计算 建立涵盖许多人类细胞类型和疾病的变异元素表型目录的模型。第四，我们 将通过共享数据、协议和软件以及进行系统评估来支持未来的研究 将变异与表型联系起来的CRISPR技术和计算模型。总而言之，这些研究将 加深我们对DNA变异和元素如何影响基因组功能的理解，并展示一种新的 利用高通量基因组工具了解人类疾病的生物学机制的战略。

PROJECT SUMMARY Genome-wide association studies have now discovered tens of thousands of noncoding variants associated with human diseases and traits. It has proven challenging to interpret these associations. A majority of causal variants lie in the noncoding genome and appear to affect DNA cis-regulatory elements, which control the logic of gene expression and could point us to new cell types, genes, and pathways for disease. However, we have lacked the tools needed to systematically characterize how these cis-regulatory variants and elements impact genome function and phenotype. Our team at Stanford University has now developed innovative single-cell, CRISPR mapping, and computational technologies that will enable identifying and functionally characterizing many thousands of elements and variants directly in the human genome. These tools include single-cell ATAC-seq to identify candidate elements in cells and tissues; sensitive CRISPR tiling methods to connect thousands of elements and variants to effects on gene expression and cellular phenotypes; and the ABC and BPNet models to predict how disease variants regulate gene expression. Together, these technologies suggest a new strategy to systematically connect DNA variants and elements to function and phenotype. Here we will apply these new technologies in collaboration with the NHGRI Impact of Genomic Variation on Function Consortium. We will use four cardiovascular cell types derived from human pluripotent stem cells as model systems. First, we will leverage single-cell maps of cardiac differentiation and development to select elements and risk variants for adult and children’s heart diseases likely to control cardiovascular cell function. Second, we will apply single-cell CRISPR tools to measure the effects of thousands of unbiased elements and variants on gene expression, and connect prioritized disease variants to target genes, cellular phenotypes, and tissue phenotypes. Third, we will leverage these experimental datasets to calibrate and refine computational models to build a variant-element-phenotype catalog across many human cell types and diseases. Fourth, we will enable future studies by sharing data, protocols, and software, and by conducting systematic evaluations of CRISPR technologies and computational models to connect variants to phenotypes. Together, these studies will advance our understanding of how DNA variants and elements impact genome function and demonstrate a novel strategy to leverage high-throughput genomic tools to understand biological mechanisms of human diseases.