Electrophysiological Abnormalities and Pharmacological Corrections of Pathogenic Missense Variants in KCNQ3
Xiaorong Wu1,2 · Jili Gong1,2 · Li Qiu2,3 · Guimei Yang2,3 · Hui Yuan2,3 · Xiangchun Shen1 · Yanwen Shen4,5 · Fuyun Tian2 · Zhaobing Gao1,2,31 School of Pharmaceutical Sciences, Guizhou Medical University, Guiyang 550025, China
2 Zhongshan Institute for Drug Discovery, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Zhongshan 528400, China
3 School of Pharmacy, Zunyi Medical University, Zunyi 563000, China
4 Translational Research Center for the Nervous System, Brain Cognition and Brain Disease Institute, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen 518055, Guangdong, China
5 Faculty of Pediatrics, Chinese PLA General Hospital, Graduate School of the PLA General Hospital, Beijing 100853, China
Abstract
The KCNQ potassium channels play a crucial role in modulating neural excitability, and their dysfunction is closely associated with epileptic disorders. While variants in KCNQ2 have been extensively studied, KCNQ3-related disorders have rarely been reported. With advances in next-generation sequencing technologies, an increasing number of cases of KCNQ3-related disorders have been identified. However, the correlation between genotype and phenotype remains poorly understood. In this study, we established a variant library consisting of 24 missense mutations in KCNQ3 and introduced these mutations into three different template types: KCNQ3, KCNQ3-A315T (Q3*), and KCNQ3-KCNQ2 tandem (Q3-Q2). We then analyzed the effects of these mutations on the KCNQ3 channel function using patch-clamp recording. The most informative parameter across all three backgrounds was the current density of the mutant channels. The current density patterns in the Q3* and Q3-Q2 backgrounds were similar, with most mutations resulting in an almost complete loss of function (LOF), they were concentrated in the pore-forming domain of KCNQ3. In contrast, mutations in the voltage-sensing domain or C-terminus did not show significant differences from the wild-type channel. Interestingly, these LOF mutations were typically associated with self-limited familial neonatal epilepsy, while neurodevelopmental disorders (NDD) were more closely associated with mutations that did not significantly differ from the wild-type. V1/2, another important parameter of the electrophysiological properties, could not be accurately determined in the majority of KCNQ3 mutations due to its nearly complete LOF in the Q3* and Q3-Q2 backgrounds. Intriguingly, the V1/2 of functional mutations were primarily leftward shifted, indicating a gain-of-function (GOF) effect, which was typically associated with NDD. In addition to previously reported mutations, we identified G553R as a novel GOF mutation. In the co-transfection background, parameters such as V1/2 could be determined, but the dysfunctional effects of these mutations were mitigated by the co-expression of wild-type KCNQ3 and KCNQ2 subunits, resulting in no significant differences between most mutations and the wild-type channel. Furthermore, we applied KCNQ modulators to reverse the electrophysiological abnormalities caused by KCNQ3 variants. The LOF mutations were reversed by the application of Pynegabine (HN37), a KCNQ opener, while the GOF mutation responded well to Amitriptyline (AMI), a KCNQ inhibitor. These findings provide essential insights into the pathogenic mechanisms underlying KCNQ3-related disorders and may inform clinical decision-making.
Keywords
KCNQ3; Self-limited familial neonatal epilepsy; Neurodevelopmental disorder; Pynegabine; Amitriptyline; Electrophysiology
