The Department of Organismic and Evolutionary Biology, Harvard University, 26 Oxford Street, Cambridge, MA 02138, United States.
Broad Institute of MIT and Harvard, 75 Ames Street, Cambridge, MA 02142, United States.
Hum Mol Genet. 2024 Jan 20;33(3):270-283. doi: 10.1093/hmg/ddad177.
While genome-wide association studies (GWAS) and positive selection scans identify genomic loci driving human phenotypic diversity, functional validation is required to discover the variant(s) responsible. We dissected the IVD gene locus-which encodes the isovaleryl-CoA dehydrogenase enzyme-implicated by selection statistics, multiple GWAS, and clinical genetics as important to function and fitness. We combined luciferase assays, CRISPR/Cas9 genome-editing, massively parallel reporter assays (MPRA), and a deletion tiling MPRA strategy across regulatory loci. We identified three regulatory variants, including an indel, that may underpin GWAS signals for pulmonary fibrosis and testosterone, and that are linked on a positively selected haplotype in the Japanese population. These regulatory variants exhibit synergistic and opposing effects on IVD expression experimentally. Alleles at these variants lie on a haplotype tagged by the variant most strongly associated with IVD expression and metabolites, but with no functional evidence itself. This work demonstrates how comprehensive functional investigation and multiple technologies are needed to discover the true genetic drivers of phenotypic diversity.
虽然全基因组关联研究 (GWAS) 和正选择扫描可以确定驱动人类表型多样性的基因组位点，但需要进行功能验证才能发现负责的变异。我们剖析了 IVD 基因座，该基因座编码异戊酰辅酶 A 脱氢酶，选择统计、多次 GWAS 和临床遗传学都表明它对功能和适应性很重要。我们结合了荧光素酶测定、CRISPR/Cas9 基因组编辑、大规模平行报告基因分析 (MPRA) 以及跨越调控基因座的缺失平铺 MPRA 策略。我们鉴定了三个调控变异，包括一个插入缺失，这些变异可能是肺纤维化和睾酮 GWAS 信号的基础，并且在日本人群中与一个正选择的单倍型相关。这些调控变异在实验中表现出协同和相反的作用。在这些变异中，等位基因位于与 IVD 表达和代谢物关联最强的变异标记的单倍型上，但该变异本身没有功能证据。这项工作展示了如何进行全面的功能研究和多种技术来发现表型多样性的真正遗传驱动因素。
