高通量 PRIME 编辑筛选可鉴定人类基因组中的功能 DNA 变体。
High-throughput PRIME-editing screens identify functional DNA variants in the human genome.
机构信息
Institute for Human Genetics, University of California, San Francisco, San Francisco, CA, USA.
Department of Epidemiology and Biostatistics, University of California, San Francisco, San Francisco, CA, USA.
出版信息
Mol Cell. 2023 Dec 21;83(24):4633-4645.e9. doi: 10.1016/j.molcel.2023.11.021.
Despite tremendous progress in detecting DNA variants associated with human disease, interpreting their functional impact in a high-throughput and single-base resolution manner remains challenging. Here, we develop a pooled prime-editing screen method, PRIME, that can be applied to characterize thousands of coding and non-coding variants in a single experiment with high reproducibility. To showcase its applications, we first identified essential nucleotides for a 716 bp MYC enhancer via PRIME-mediated single-base resolution analysis. Next, we applied PRIME to functionally characterize 1,304 genome-wide association study (GWAS)-identified non-coding variants associated with breast cancer and 3,699 variants from ClinVar. We discovered that 103 non-coding variants and 156 variants of uncertain significance are functional via affecting cell fitness. Collectively, we demonstrate that PRIME is capable of characterizing genetic variants at single-base resolution and scale, advancing accurate genome annotation for disease risk prediction, diagnosis, and therapeutic target identification.
尽管在检测与人类疾病相关的 DNA 变异方面取得了巨大进展，但以高通量和单碱基分辨率的方式解释它们的功能影响仍然具有挑战性。在这里，我们开发了一种 pooled prime-editing 筛选方法 PRIME，它可以在单个实验中以高重复性来表征数千个编码和非编码变体。为了展示其应用，我们首先通过 PRIME 介导的单碱基分辨率分析确定了 716 bp MYC 增强子的必需核苷酸。接下来，我们应用 PRIME 对与乳腺癌相关的 1304 个全基因组关联研究 (GWAS) 鉴定的非编码变体和来自 ClinVar 的 3699 个变体进行功能表征。我们发现，通过影响细胞适应性，103 个非编码变体和 156 个意义不明的变体是有功能的。总的来说，我们证明了 PRIME 能够以单碱基分辨率和规模来表征遗传变异，推进了疾病风险预测、诊断和治疗靶点识别的精确基因组注释。
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