Khare Swati, Nick Jerelyn A, Zhang Yalan, Galeano Kira, Butler Brittany, Khoshbouei Habibeh, Rayaprolu Sruti, Hathorn Tyisha, Ranum Laura P W, Smithson Lisa, Golde Todd E, Paucar Martin, Morse Richard, Raff Michael, Simon Julie, Nordenskjöld Magnus, Wirdefeldt Karin, Rincon-Limas Diego E, Lewis Jada, Kaczmarek Leonard K, Fernandez-Funez Pedro, Nick Harry S, Waters Michael F
Department of Neurology, University of Florida, Gainesville, FL, United States of America.
McKnight Brain Institute, University of Florida, Gainesville, FL, United States of America.
PLoS One. 2017 May 3;12(5):e0173565. doi: 10.1371/journal.pone.0173565. eCollection 2017.
The autosomal dominant spinocerebellar ataxias (SCAs) are a diverse group of neurological disorders anchored by the phenotypes of motor incoordination and cerebellar atrophy. Disease heterogeneity is appreciated through varying comorbidities: dysarthria, dysphagia, oculomotor and/or retinal abnormalities, motor neuron pathology, epilepsy, cognitive impairment, autonomic dysfunction, and psychiatric manifestations. Our study focuses on SCA13, which is caused by several allelic variants in the voltage-gated potassium channel KCNC3 (Kv3.3). We detail the clinical phenotype of four SCA13 kindreds that confirm causation of the KCNC3R423H allele. The heralding features demonstrate congenital onset with non-progressive, neurodevelopmental cerebellar hypoplasia and lifetime improvement in motor and cognitive function that implicate compensatory neural mechanisms. Targeted expression of human KCNC3R423H in Drosophila triggers aberrant wing veins, maldeveloped eyes, and fused ommatidia consistent with the neurodevelopmental presentation of patients. Furthermore, human KCNC3R423H expression in mammalian cells results in altered glycosylation and aberrant retention of the channel in anterograde and/or endosomal vesicles. Confirmation of the absence of plasma membrane targeting was based on the loss of current conductance in cells expressing the mutant channel. Mechanistically, genetic studies in Drosophila, along with cellular and biophysical studies in mammalian systems, demonstrate the dominant negative effect exerted by the mutant on the wild-type (WT) protein, which explains dominant inheritance. We demonstrate that ocular co-expression of KCNC3R423H with Drosophila epidermal growth factor receptor (dEgfr) results in striking rescue of the eye phenotype, whereas KCNC3R423H expression in mammalian cells results in aberrant intracellular retention of human epidermal growth factor receptor (EGFR). Together, these results indicate that the neurodevelopmental consequences of KCNC3R423H may be mediated through indirect effects on EGFR signaling in the developing cerebellum. Our results therefore confirm the KCNC3R423H allele as causative for SCA13, through a dominant negative effect on KCNC3WT and links with EGFR that account for dominant inheritance, congenital onset, and disease pathology.
常染色体显性遗传性脊髓小脑共济失调(SCAs)是一组多样的神经系统疾病,其特征为运动不协调和小脑萎缩。疾病的异质性体现在各种合并症上:构音障碍、吞咽困难、动眼神经和/或视网膜异常、运动神经元病变、癫痫、认知障碍、自主神经功能障碍以及精神症状。我们的研究聚焦于SCA13,它由电压门控钾通道KCNC3(Kv3.3)中的几个等位基因变异引起。我们详细描述了四个SCA13家族的临床表型,证实了KCNC3 R423H等位基因的致病性。首发特征显示为先天性发病,伴有非进行性、神经发育性小脑发育不全,以及运动和认知功能在一生中的改善,这暗示了代偿性神经机制。在果蝇中靶向表达人KCNC3 R423H会引发异常的翅脉、发育不良的眼睛和融合的小眼,这与患者的神经发育表现一致。此外,在哺乳动物细胞中表达人KCNC3 R423H会导致糖基化改变以及通道在前向和/或内体小泡中的异常滞留。基于表达突变通道的细胞中电流传导的丧失,证实了细胞膜靶向缺失。从机制上讲,果蝇的遗传学研究以及哺乳动物系统中的细胞和生物物理学研究表明,突变体对野生型(WT)蛋白施加了显性负效应,这解释了显性遗传。我们证明,在果蝇中KCNC3 R423H与表皮生长因子受体(dEgfr)的眼部共表达显著挽救了眼部表型,而在哺乳动物细胞中KCNC3 R423H的表达导致人表皮生长因子受体(EGFR)在细胞内的异常滞留。总之,这些结果表明,KCNC3 R423H的神经发育后果可能是通过对发育中小脑的EGFR信号传导的间接影响来介导的。因此,我们的结果证实了KCNC3 R423H等位基因是SCA13的致病原因,它通过对KCNC3 WT的显性负效应以及与EGFR的联系来解释显性遗传、先天性发病和疾病病理。