The Genome Function Laboratory, The Francis Crick Institute, London, UK.
Renal Division, Department of Medicine, Medical Center-University of Freiburg, Faculty of Medicine, University of Freiburg, Freiburg, Germany.
Nat Genet. 2024 Jul;56(7):1446-1455. doi: 10.1038/s41588-024-01800-z. Epub 2024 Jul 5.
To maximize the impact of precision medicine approaches, it is critical to identify genetic variants underlying disease and to accurately quantify their functional effects. A gene exemplifying the challenge of variant interpretation is the von Hippel-Lindautumor suppressor (VHL). VHL encodes an E3 ubiquitin ligase that regulates the cellular response to hypoxia. Germline pathogenic variants in VHL predispose patients to tumors including clear cell renal cell carcinoma (ccRCC) and pheochromocytoma, and somatic VHL mutations are frequently observed in sporadic renal cancer. Here we optimize and apply saturation genome editing to assay nearly all possible single-nucleotide variants (SNVs) across VHL's coding sequence. To delineate mechanisms, we quantify mRNA dosage effects and compare functional effects in isogenic cell lines. Function scores for 2,268 VHL SNVs identify a core set of pathogenic alleles driving ccRCC with perfect accuracy, inform differential risk across tumor types and reveal new mechanisms by which variants impact function. These results have immediate utility for classifying VHL variants encountered clinically and illustrate how precise functional measurements can resolve pleiotropic and dosage-dependent genotype-phenotype relationships across complete genes.
为了最大限度地发挥精准医学方法的影响，关键是要确定疾病相关的遗传变异，并准确量化它们的功能效应。一个基因范例说明了变异解释的挑战，这个基因就是 von Hippel-Lindau 肿瘤抑制基因（VHL）。VHL 编码一种 E3 泛素连接酶，可调节细胞对缺氧的反应。VHL 的种系致病性变异使患者易患肿瘤，包括透明细胞肾细胞癌（ccRCC）和嗜铬细胞瘤，散发性肾细胞癌中也经常观察到体细胞 VHL 突变。在这里，我们优化并应用饱和基因组编辑来检测 VHL 编码序列中几乎所有可能的单核苷酸变异（SNV）。为了阐明机制，我们量化了 mRNA 剂量效应，并在同基因细胞系中比较了功能效应。2,268 个 VHL SNV 的功能评分准确地确定了一组核心致病性等位基因，驱动 ccRCC 的发生，跨肿瘤类型的差异化风险，并揭示了变异影响功能的新机制。这些结果可立即用于对临床上遇到的 VHL 变异进行分类，并说明了精确的功能测量如何解决完整基因中多效性和剂量依赖性基因型-表型关系。
