Functional effects of ion channel mutations found via exome sequencing

对患有神经疾病的患者进行基因组分析正变得越来越普遍，提供了大量与患者DNA变化与神经疾病相关的基因组数据。然而，在了解这些突变对靶蛋白的功能影响方面的进展，并没有与识别基因突变的速度相匹配。对于大多数基因来说，已知的与疾病相关的错义突变远远超过存在功能数据的突变。因此，在我们所知的突变和我们对单个突变如何影响蛋白质功能的理解之间存在着巨大的鸿沟。随着测序步伐的加快，这一鸿沟正在变得越来越大。虽然存在预测哪些错义突变可能有害的计算方法，但这些算法并不能很好地替代功能数据。因此，迫切需要一种方法，使面向临床的实验室能够获得对突变影响的功能洞察。这项R24提案试图填补配体门控谷氨酸受体基因家族的突变和功能之间的空白，该家族包括分别由GriA、GRIK和GRIN基因编码的AMPA、KAINATE和NMDA受体。将重点放在一个基因家族上的决定是在与NIH协商后做出的，并受到这一资助机会的预算的推动。这个编码NMDA受体亚单位的基因家族的已知突变数量正在快速增长，仅在过去18个月里就描述了100多个与疾病相关的新突变，还有更多已知但未发表的突变。因此，虽然我们将重点关注所有谷氨酸受体突变，但我们预计大多数已发现的突变将属于GRIN家族。对谷氨酸受体突变感兴趣的研究人员将分享氨基酸变化，这些变化将被引入商业cDNA，用于异源表达系统，以获得描述突变影响的功能数据。资源中心将与临床实验室共享这些功能信息，并努力促进数据的发布。这一努力将填补文献中的一个关键空白，促进我们对神经疾病的理解，并为未来的治疗范例提供新的想法。这笔R24资源赠款将支持三项活动。首先，我们将联系至少100个进行外显子组/基因组测序的实验室，向他们通报这一资源。其次，对于已经发现GRIN突变并希望获得功能分析的临床或研究人员，我们将把该突变引入商业化的人类cDNA中，并从功能上评估该突变对激动剂效力、调节剂敏感性、反应时间进程和受体表面表达的影响。第三，我们将在网上开发一个可搜索的数据库，将这个项目的结果存档，并将我们的结果与几个现有的数据库(ClinVar、EGI等)连接起来。

Genomic analysis of patients with neurological conditions is becoming increasingly more common, providing a great deal of genomic data relating changes in patient DNA to neurological disease. However, the rate at which genetic mutations are identified has not been matched by advances in understanding the functional effects of these mutations on target proteins. For most genes, known disease-associated missense mutations far outnumber mutations for which functional data exists. Thus, a large gulf exists between the mutations we know about and our understanding of how individual mutations impact protein function. This gulf is growing wider with the accelerating pace of sequencing. While computational methods exist that predict which missense mutations may be harmful, these algorithms are a poor substitute for functional data. Thus, there is a critical need for a means by which clinically-oriented laboratories can obtain functional insight into the effects of mutations. This R24 proposal seeks to fill the gap between mutations and function for the ligand-gated glutamate receptor gene family, which includes AMPA, kainate, and NMDA receptors encoded by GRIA, GRIK, and GRIN genes, respectively. The decision to focus efforts on one gene family was made in consultation with NIH, and was driven by the budget for this funding opportunity. The number of known mutations in this family of genes encoding NMDA receptor subunits is growing at a fast pace, with well over a hundred novel disease- associated mutations described in just the past 18 months, and many more known but unpublished. Thus, while we will focus on all glutamate receptor mutations, we expect most mutations identified will be in the GRIN family. Interested investigators with glutamate receptor mutations will share the amino acid change, which will be introduced into commercial cDNAs for use in heterologous expressions systems to obtain functional data describing the effects of the mutation. The Resource Center will share this functional information with the clinical lab, and work to facilitate publication of the data. This effort will fill a critical gap in the literature, advance our understanding of neurological disease, and provide new ideas about future treatment paradigms. Three activities will be supported by this R24 resource grant. First, we will contact at least 100 laboratories performing whole exome/genome sequencing to inform them of this resource. Second, for clinics or investigators who have identified GRIN mutations and would like to obtain functional analysis, we will introduce the mutation into commercially available human cDNAs and functionally evaluate the effects of the mutation on agonist potency, modulator sensitivity, response time course, and receptor surface expression. Third, we will develop a searchable database on the web to archive results from this project, and connect our results with several existing databases (ClinVar, EGI, etc.).