Miceli Francesco, Carotenuto Lidia, Barrese Vincenzo, Soldovieri Maria Virginia, Heinzen Erin L, Mandel Arthur M, Lippa Natalie, Bier Louise, Goldstein David B, Cooper Edward C, Cilio Maria Roberta, Taglialatela Maurizio, Sands Tristan T
Department of Neuroscience, University of Naples "Federico II", Naples, Italy.
Department of Medicine and Health Science, University of Molise, Campobasso, Italy.
Front Physiol. 2020 Sep 4;11:1040. doi: 10.3389/fphys.2020.01040. eCollection 2020.
Pathogenic variants in and , paralogous genes encoding Kv7.2 and Kv7.3 voltage-gated K channel subunits, are responsible for early-onset developmental/epileptic disorders characterized by heterogeneous clinical phenotypes ranging from benign familial neonatal epilepsy (BFNE) to early-onset developmental and epileptic encephalopathy (DEE). variants account for the majority of pedigrees with BFNE and variants are responsible for a much smaller subgroup, but the reasons for this imbalance remain unclear. Analysis of additional pedigrees is needed to further clarify the nature of this genetic heterogeneity and to improve prediction of pathogenicity for novel variants. We identified a BFNE family with two siblings and a parent affected. Exome sequencing on samples from both parents and siblings revealed a novel variant (c.719T>G; p.M240R), segregating in the three affected individuals. The M240 residue is conserved among human Kv7.2-5 and lies between the two arginines (R5 and R6) closest to the intracellular side of the voltage-sensing S transmembrane segment. Whole cell patch-clamp recordings in Chinese hamster ovary (CHO) cells revealed that homomeric Kv7.3 M240R channels were not functional, whereas heteromeric channels incorporating Kv7.3 M240R mutant subunits with Kv7.2 and Kv7.3 displayed a depolarizing shift of about 10 mV in activation gating. Molecular modeling results suggested that the M240R substitution preferentially stabilized the resting state and possibly destabilized the activated state of the Kv7.3 subunits, a result consistent with functional data. Exposure to β-hydroxybutyrate (BHB), a ketone body generated during the ketogenic diet (KD), reversed channel dysfunction induced by the M240R variant. In conclusion, we describe the first missense loss-of-function (LoF) pathogenic variant within the S segment of Kv7.3 identified in patients with BFNE. Studied under conditions mimicking heterozygosity, the M240R variant mainly affects the voltage sensitivity, in contrast to previously analyzed BFNE Kv7.3 variants that reduce current density. Our pharmacological results provide a rationale for the use of KD in patients carrying LoF variants in Kv7.2 or Kv7.3 subunits.
KCNQ2和KCNQ3是编码Kv7.2和Kv7.3电压门控钾通道亚基的旁系同源基因，其致病性变异与早发性发育/癫痫性疾病相关，这些疾病具有从良性家族性新生儿癫痫（BFNE）到早发性发育性和癫痫性脑病（DEE）等异质性临床表型。KCNQ2变异占BFNE家系的大多数，而KCNQ3变异仅导致一小部分家系发病，但其原因尚不清楚。需要分析更多家系以进一步阐明这种遗传异质性的本质，并提高对新变异致病性的预测。我们鉴定了一个有两名患病兄弟姐妹和一名患病父母的BFNE家系。对父母和兄弟姐妹的样本进行外显子组测序，发现了一个新的KCNQ3变异（c.719T>G；p.M240R），在三名患病个体中分离。M240残基在人类Kv7.2 - 5中保守，位于最靠近电压感应S跨膜段细胞内侧的两个精氨酸（R5和R6）之间。在中国仓鼠卵巢（CHO）细胞中进行的全细胞膜片钳记录显示，同聚体Kv7.3 M240R通道无功能，而包含Kv7.3 M240R突变亚基与Kv7.2和Kv7.3的异聚体通道在激活门控方面表现出约10 mV的去极化偏移。分子模拟结果表明，M240R替代优先稳定了Kv7.3亚基的静息状态，并可能使激活状态不稳定，这一结果与功能数据一致。暴露于β-羟基丁酸（BHB），即生酮饮食（KD）期间产生的一种酮体，可逆转由M240R变异诱导的通道功能障碍。总之，我们描述了在BFNE患者中鉴定出的首个位于Kv7.3的S段内的错义功能丧失（LoF）致病性变异。在模拟杂合性的条件下研究发现，与之前分析的降低电流密度的BFNE Kv7.3变异不同，M240R变异主要影响电压敏感性。我们的药理学结果为在携带Kv7.2或Kv7.3亚基LoF变异的患者中使用KD提供了理论依据。
