Impaired surface expression and conductance of the KCNQ4 channel lead to sensorineural hearing loss.
Impaired surface expression and conductance of the KCNQ4 channel lead to sensorineural hearing loss.
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DOI：
10.1111/jcmm.12080
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发表时间：
2013-07
影响因子：
5.3
通讯作者：
Nie L
中科院分区：
文献类型：
--
作者：
Gao Y;Yechikov S;Vázquez AE;Chen D;Nie L
KCNQ4 是一种电压门控钾通道，在维持耳蜗离子稳态和调节毛细胞膜电位方面发挥着重要作用，这两者对于正常听觉功能至关重要。 KCNQ4 基因突变导致 DFNA2，这是常染色体显性非综合征性耳聋的一种亚型，其特征是所有频率的进行性感音神经性听力损失。尽管最近在致病性 KCNQ4 突变的鉴定方面取得了进展，但 DFNA2 的分子病因学仍然未知。我们在此报告，细胞表面表达减少和 KCNQ4 通道电导受损是 DFNA2 听力损失的两种机制。在 HEK293T 细胞中，通过免疫荧光显微镜检测到致病性 KCNQ4 突变体 L274H、W276S、L281S、G285C、G285S、G296S 和 G321S 的细胞表面表达显着降低，并通过蛋白质印迹证实，而它们的总体细胞水平保持正常。此外，这些突变均不影响 KCNQ4 通道的四聚体组装。与这些结果一致，所有突变体都对野生型（WT）通道功能表现出强烈的显性负效应。最重要的是，HSP90β（控制 KCNQ4 生物发生的分子伴侣网络的关键组成部分）的过度表达显着增加了 KCNQ4 突变体 L281S、G296S 和 G321S 的细胞表面表达。 HEK293T 细胞中 KCNQ4 表面表达恢复或显着改善，模拟 DFNA2 患者中这些突变的杂合状态。最后，我们的电生理学研究表明，这些突变直接损害 KCNQ4 通道的电导，因为在 KCNQ4 表面表达恢复或改善后，没有观察到 KCNQ4 电流的显着变化。
KCNQ4, a voltage-gated potassium channel, plays an important role in maintaining cochlear ion homoeostasis and regulating hair cell membrane potential, both essential for normal auditory function. Mutations in the KCNQ4 gene lead to DFNA2, a subtype of autosomal dominant non-syndromic deafness that is characterized by progressive sensorineural hearing loss across all frequencies. Despite recent advances in the identification of pathogenic KCNQ4 mutations, the molecular aetiology of DFNA2 remains unknown. We report here that decreased cell surface expression and impaired conductance of the KCNQ4 channel are two mechanisms underlying hearing loss in DFNA2. In HEK293T cells, a dramatic decrease in cell surface expression was detected by immunofluorescent microscopy and confirmed by Western blot for the pathogenic KCNQ4 mutants L274H, W276S, L281S, G285C, G285S, G296S and G321S, while their overall cellular levels remained normal. In addition, none of these mutations affected tetrameric assembly of KCNQ4 channels. Consistent with these results, all mutants showed strong dominant-negative effects on the wild-type (WT) channel function. Most importantly, overexpression of HSP90β, a key component of the molecular chaperone network that controls the KCNQ4 biogenesis, significantly increased cell surface expression of the KCNQ4 mutants L281S, G296S and G321S. KCNQ4 surface expression was restored or considerably improved in HEK293T cells mimicking the heterozygous condition of these mutations in DFNA2 patients. Finally, our electrophysiological studies demonstrated that these mutations directly compromise the conductance of the KCNQ4 channel, since no significant change in KCNQ4 current was observed after KCNQ4 surface expression was restored or improved.
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