Analysis of CLCNKB mutations at dimer-interface, calcium-binding site, and pore reveals a variety of functional alterations in ClC-Kb channel leading to Bartter syndrome
Analysis of CLCNKB mutations at dimer-interface, calcium-binding site, and pore reveals a variety of functional alterations in ClC-Kb channel leading to Bartter syndrome
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DOI：
10.1002/humu.23962
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发表时间：
2019-12-24
期刊：
影响因子：
3.9
通讯作者：
Andrini，Olga
中科院分区：
文献类型：
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作者：
Bignon，Yohan;Sakhi，Imene;Andrini，Olga
CLCNKB基因的病理性错义突变可导致Bartter综合征III型患者的广泛临床表型。突变的ClC-Kb通道的分子分析可以有助于根据其功能改变对突变进行分类。我们研究了导致Bartter综合征的CLCNKB基因中9个突变的功能后果。我们首先确定所有测试的突变都会导致ClC-Kb电流降低。结合电生理和生化方法在非洲爪蟾细胞和MDCKII细胞，我们确定了三类突变。一类的特征是改变渠道贩运。p.A210V、p.P216L、p.G424R和p.G437R全部或部分保留在内质网中。p.S218N的特征在于质膜上的通道插入减少和pH敏感性改变;因此，它福尔斯第二类突变。最后，我们发现了一类新的功能失活突变体通常存在于质膜。事实上，我们发现p.A204T改变pH敏感性，p.A254V消除钙敏感性。p.G219C和p.G465R可能在质膜上部分失活。总之，大多数致病性突变体部分或全部积累在细胞内室，但一些突变体通常存在于膜表面，同时显示出大范围的改变通道门控特性。
Pathological missense mutations inCLCNKBgene give a wide spectrum of clinical phenotypes in Bartter syndrome type III patients. Molecular analysis of the mutated ClC‐Kb channels can be helpful to classify the mutations according to their functional alteration. We investigated the functional consequences of nine mutations in theCLCNKBgene causing Bartter syndrome. We first established that all tested mutations lead to decreased ClC‐Kb currents. Combining electrophysiological and biochemical methods inXenopus laevisoocytes and in MDCKII cells, we identified three classes of mutations. One class is characterized by altered channel trafficking. p.A210V, p.P216L, p.G424R, and p.G437R are totally or partially retained in the endoplasmic reticulum. p.S218N is characterized by reduced channel insertion at the plasma membrane and altered pH‐sensitivity; thus, it falls in the second class of mutations. Finally, we found a novel class of functionally inactivated mutants normally present at the plasma membrane. Indeed, we found that p.A204T alters the pH‐sensitivity, p.A254V abolishes the calcium‐sensitivity. p.G219C and p.G465R are probably partially inactive at the plasma membrane. In conclusion, most pathogenic mutants accumulate partly or totally in intracellular compartments, but some mutants are normally present at the membrane surface and simultaneously show a large range of altered channel gating properties.