Genetic testing has increased the number of variants identified in disease genes, but the diagnostic utility is limited by lack of understanding variant function. CARD11 encodes an adaptor protein that expresses dominant-negative and gain-of-function variants associated with distinct immunodeficiencies. Here, we used a “cloning-free” saturation genome editing approach in a diploid cell line to simultaneously score 2,542 variants for decreased or increased function in the region of CARD11 associated with immunodeficiency. We also described an exon-skipping mechanism for CARD11 dominant-negative activity. The classification of reported clinical variants was sensitive (94.6%) and specific (88.9%), which rendered the data immediately useful for interpretation of seven coding and splicing variants implicated in immunodeficiency found in our clinic. This approach is generalizable for variant interpretation in many other clinically actionable genes, in any relevant cell type.

基因检测增加了在疾病基因中鉴定出的变异数量，但由于缺乏对变异功能的理解，诊断效用受到限制。CARD11编码一种接头蛋白，该蛋白表达与不同免疫缺陷相关的显性失活和功能获得变体。在这里，我们在二倍体细胞系中使用“无克隆”饱和基因组编辑方法，同时对CARD11区域功能降低或增加的 2,542 个变体进行评分与免疫缺陷有关。我们还描述了 CARD11 显性负活性的外显子跳跃机制。报告的临床变异的分类是敏感的（94.6%）和特异性的（88.9%），这使得这些数据立即可用于解释我们诊所发现的与免疫缺陷有关的七种编码和剪接变异。这种方法可推广到任何相关细胞类型中许多其他临床可操作基因的变异解释。