Subbotina Ekaterina, Yang Hua-Qian, Gando Ivan, Williams Nori, Sampson Barbara A, Tang Yingying, Coetzee William A
Departments of Pediatrics, NYU School of Medicine, New York, NY 10016 USA.
Molecular Genetics Laboratory, Office of Chief Medical Examiner, New York, NY USA.
Forensic Sci Int. 2019 May;298:80-87. doi: 10.1016/j.forsciint.2019.02.035. Epub 2019 Feb 27.
Genetic variation in ion channel genes ('channelopathies') are often associated with inherited arrhythmias and sudden death. Genetic testing ('molecular autopsies') of channelopathy genes can be used to assist in determining the likely causes of sudden unexpected death. However, different in silico approaches can yield conflicting pathogenicity predictions and assessing their impact on ion channel function can assist in this regard.
We performed genetic testing of cases of sudden expected death in the New York City metropolitan area and found four rare or novel variants in ABCC9, which codes for the regulatory SUR2 subunit of K channels. All were missense variants, causing amino acid changes in the protein. Three of the variants (A355S, M941V, and K1379Q) were in cases of infants less than six-months old and one (H1305Y) was in an adult. The predicted pathogenicities of the variants were conflicting. We have introduced these variants into a human SUR2A cDNA, which we coexpressed with the Kir6.2 pore-forming subunit in HEK-293 cells and subjected to patch clamp and biochemical assays. Each of the four variants led to gain-of-function phenotypes. The A355S and M941V variants increased in the overall patch current. The sensitivity of the K channels to inhibitory 'cytosolic' ATP was repressed for the M941V, H1305Y and K1379Q variants. None of the variants had any effect on the unitary K channel current or the surface expression of K channels, as determined with biotinylation assays, suggesting that all of the variants led to an enhanced open state.
All four variants caused a gain-of-function phenotype. Given the expression of SUR2-containing K channels in the heart and specialized cardiac conduction, vascular smooth muscle and respiratory neurons, it is conceivable that electrical silencing of these cells may contribute to the vulnerability element, which is a component of the triple risk model of sudden explained death in infants. The gain-of-function phenotype of these ABCC9 variants should be considered when assessing their potential pathogenicity.
离子通道基因的遗传变异（“通道病”）常与遗传性心律失常和猝死相关。通道病基因的基因检测（“分子尸检”）可用于协助确定意外猝死的可能原因。然而，不同的计算机模拟方法可能会产生相互矛盾的致病性预测，评估它们对离子通道功能的影响在这方面会有所帮助。
我们对纽约市大都市区意外猝死病例进行了基因检测，在编码钾通道调节性SUR2亚基的ABCC9基因中发现了四个罕见或新的变异。所有变异均为错义变异，导致蛋白质中的氨基酸发生变化。其中三个变异（A355S、M941V和K1379Q）出现在6个月以下婴儿的病例中，一个变异（H1305Y）出现在一名成年人中。这些变异的预测致病性相互矛盾。我们将这些变异引入人SUR2A cDNA中，该cDNA与Kir6.2孔形成亚基在HEK-293细胞中共表达，并进行膜片钳和生化分析。这四个变异中的每一个都导致了功能增强表型。A355S和M941V变异使整体膜片电流增加。M941V、H1305Y和K1379Q变异使钾通道对抑制性“胞质”ATP的敏感性降低。通过生物素化分析确定，这些变异均未对单个钾通道电流或钾通道的表面表达产生任何影响，这表明所有变异均导致开放状态增强。
所有四个变异均导致功能增强表型。鉴于含SUR2的钾通道在心脏、特殊心脏传导系统、血管平滑肌和呼吸神经元中的表达，可以想象这些细胞的电沉默可能是婴儿不明原因猝死三联风险模型中易损因素的一个组成部分。在评估这些ABCC9变异的潜在致病性时，应考虑其功能增强表型。
