Disease-causing mutant WNK4 increases paracellular chloride permeability and phosphorylates claudins.
Disease-causing mutant WNK4 increases paracellular chloride permeability and phosphorylates claudins.
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DOI：
10.1073/pnas.0306924101
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发表时间：
2004-03
影响因子：
11.1
通讯作者：
K. Yamauchi;T. Rai;Katsuki Kobayashi;E. Sohara;Tatsunori Suzuki;T. Itoh;Shin Suda;Atsushi Hayama;S. Sasaki;S. Uchida
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文献类型：
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作者：
K. Yamauchi;T. Rai;Katsuki Kobayashi;E. Sohara;Tatsunori Suzuki;T. Itoh;Shin Suda;Atsushi Hayama;S. Sasaki;S. Uchida
WNK 4基因突变导致假性醛固酮减少症II型（PHAII），一种常染色体显性高钾血症和高血压疾病。这种推定激酶的靶分子和突变导致表型的分子机制目前尚不清楚。虽然最近的报道发现，在非洲爪蟾卵母细胞中的WNK 4的表达导致抑制噻嗪敏感的NaCl协同转运蛋白和肾K通道ROMK，可能有其他的WNK 4的目标。例如，已经假定细胞旁氯渗透性的增加是PHAII发病机制的介质，这一可能性得到了WNK 4在体内紧密连接处的定位的支持。为了确定这一假设的有效性，我们测量了跨上皮Na和Cl渗透性Madin-Darby犬肾II细胞稳定表达野生型或致病突变体WNK 4。我们发现，在表达致病突变体WNK 4（D564 A）的细胞中，跨上皮细胞旁Cl渗透性增加，但Na渗透性略有下降。此外，WNK 4结合和磷酸化的claudin 1-4，已知参与调节细胞旁离子渗透性的主要紧密连接膜蛋白。表达突变体WNK 4的细胞中封闭蛋白磷酸化的增加大于表达野生型蛋白的细胞。这些结果清楚地表明致病性WNK 4突变体具有功能获得活性，并且密封蛋白可能是WNK 4激酶的重要分子靶点。突变体WNK 4引起的细胞旁“氯分流”增加可能是PHAII的致病机制。
Mutations in the WNK4 gene cause pseudohypoaldosteronism type II (PHAII), an autosomal-dominant disorder of hyperkalemia and hypertension. The target molecules of this putative kinase and the molecular mechanisms by which the mutations cause the phenotypes are currently unknown. Although recent reports found that expression of WNK4 in Xenopus oocytes causes inhibition of the thiazide-sensitive NaCl cotransporter and the renal K channel ROMK, there may be additional targets of WNK4. For example, an increase in paracellular chloride permeability has been postulated to be a mediator of PHAII pathogenesis, a possibility supported by the localization of WNK4 at tight junctions in vivo. To determine the validity of this hypothesis, we measured transepithelial Na and Cl permeability in Madin-Darby canine kidney II cells stably expressing wild-type or a pathogenic mutant of WNK4. We found that transepithelial paracellular Cl permeability was increased in cells expressing a disease-causing mutant WNK4 (D564A) but that Na permeability was decreased slightly. Furthermore, WNK4 bound and phosphorylated claudins 1-4, major tight-junction membrane proteins known to be involved in the regulation of paracellular ion permeability. The increases in phosphorylation of claudins were greater in cells expressing the mutant WNK4 than in cells expressing wild-type protein. These results clearly indicate that the pathogenic WNK4 mutant possesses a gain-of-function activity and that the claudins may be important molecular targets of WNK4 kinase. The increased paracellular "chloride shunt" caused by the mutant WNK4 could be the pathogenic mechanism of PHAII.