Epilepsy-associated SCN2A (Na V 1.2) Variants Exhibit Diverse and Complex Functional Properties.
Epilepsy-associated SCN2A (Na V 1.2) Variants Exhibit Diverse and Complex Functional Properties.
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癫痫相关的 SCN2A (Na V 1.2) 变异体表现出多样且复杂的功能特性。
DOI：
10.1101/2023.02.23.529757
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发表时间：
2023
期刊：
影响因子：
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通讯作者：
GeorgeJr,AlfredL
中科院分区：
文献类型：
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作者：
Thompson,ChristopherH;Potet,Franck;Abramova,TatianaV;DeKeyser,Jean-Marc;Ghabra,NoraF;Vanoye,CarlosG;Millichap,John;GeorgeJr,AlfredL
电压门控钠离子通道（NaV）基因的致病性变异包括编码NaV1. 2的SCN 2A，其在伴有或不伴有癫痫的神经发育障碍中频繁发现，SCN 2A也是自闭症谱系障碍（ASD）和非综合征性智力残疾（ID）的高可信度风险基因。以前的工作，以确定功能的后果ofSCN 2A变异产生了一个范例，其中主要是获得的功能的变体导致癫痫发作，而功能丧失的变体与ASD和ID。然而，这个框架是来自有限数量的研究异质性实验条件下进行的，而大多数疾病相关的SCN 2A变异没有功能注释。我们使用自动化膜片钳记录来确定ofSCN 2A变体的功能特性，以证明这种方法的有效性，并检查在统一条件下研究的较大队列中变体功能障碍的二元分类是否明显。我们使用HEK293T细胞中表达的NaV1.2的两种选择性剪接亚型研究了28种疾病相关变体和4种常见变体。自动膜片钳记录提供了一种有效的高通量方法，以确定NaV1.2变体的详细功能特性，与先前使用手动膜片钳研究的变体的一致结果。在我们的研究中，许多癫痫相关的变异表现出复杂的功能获得和丧失模式，难以通过简单的二进制方案进行分类。自动膜片钳可实现的更高通量使研究变异体的记录条件更加标准化，不受操作员偏倚的影响，并增强了实验的严谨性。这种方法提供了一种增强的能力，以辨别通道功能障碍和神经发育障碍之间的关系。
Pathogenic variants in voltage-gated sodium (NaV) channel genes includingSCN2A, encoding NaV1.2, are discovered frequently in neurodevelopmental disorders with or without epilepsy.SCN2Ais also a high-confidence risk gene for autism spectrum disorder (ASD) and nonsyndromic intellectual disability (ID). Previous work to determine the functional consequences ofSCN2Avariants yielded a paradigm in which predominantly gain-of-function variants cause neonatal-onset epilepsy, whereas loss-of-function variants are associated with ASD and ID. However, this framework was derived from a limited number of studies conducted under heterogeneous experimental conditions, whereas most disease-associatedSCN2Avariants have not been functionally annotated. We determined the functional properties ofSCN2Avariants using automated patch-clamp recording to demonstrate the validity of this method and to examine whether a binary classification of variant dysfunction is evident in a larger cohort studied under uniform conditions. We studied 28 disease-associated variants and 4 common variants using two alternatively spliced isoforms of NaV1.2 expressed in HEK293T cells. Automated patch-clamp recording provided a valid high throughput method to ascertain detailed functional properties of NaV1.2 variants with concordant findings for variants that were previously studied using manual patch clamp. Many epilepsy-associated variants in our study exhibited complex patterns of gain- and loss-of-functions that are difficult to classify by a simple binary scheme. The higher throughput achievable with automated patch clamp enables study of variants with greater standardization of recording conditions, freedom from operator bias, and enhanced experimental rigor. This approach offers an enhanced ability to discern relationships between channel dysfunction and neurodevelopmental disorders.