Regulation of apical localization of the thiazide-sensitive NaCl cotransporter by WNK4 in polarized epithelial cells.
Regulation of apical localization of the thiazide-sensitive NaCl cotransporter by WNK4 in polarized epithelial cells.
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DOI：
10.1016/j.bbrc.2005.02.172
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发表时间：
2005-05
影响因子：
3.1
通讯作者：
S. Yang;K. Yamauchi;T. Rai;Atsushi Hiyama;E. Sohara;Tatsunori Suzuki;T. Itoh;Shin Suda;S. Sasaki;S. Uchida
中科院分区：
文献类型：
--
作者：
S. Yang;K. Yamauchi;T. Rai;Atsushi Hiyama;E. Sohara;Tatsunori Suzuki;T. Itoh;Shin Suda;S. Sasaki;S. Uchida
WNK4基因的错义突变被认为会导致假性醛固酮减少症II型（PHAII），这是一种常染色体显性遗传疾病，其特征是高钾血症和高血压。先前使用非洲爪蟾卵母细胞的报道表明，野生型WNK 4表达抑制噻嗪敏感性NaCl协同转运蛋白（NCC）的表面表达，而致病突变体失去了对NCC表面表达的抑制作用。为了确定在卵母细胞中观察到的这些变化是否真的发生在极化的上皮细胞中，我们产生了表达单独的NCC或NCC加野生型WNK 4或致病（D564A）WNK 4的稳定的MDCK II细胞系。与没有WNK 4共表达的NCC的顶端定位相反，免疫荧光显微镜和生物素表面标记显示，野生型和突变型WNK 4表达均同样降低了该顶端定位。两个PHAII无关的顶端转运蛋白，钠非依赖性氨基酸转运蛋白，BAT1和胆盐输出泵，Bsep的顶端定位也被发现减少野生型和突变WNK 4的表达。这些结果表明，NCC的调节与致病突变无关，也不限于PHAII相关的特异性转运蛋白。WNK 4对NCC细胞内定位的调节可能不参与PHAII的发病机制。
Missense mutations in the WNK4 gene have been postulated to cause pseudohypoaldosteronism type II (PHAII), an autosomal-dominant disorder characterized by hyperkalemia and hypertension. Previous reports using Xenopus oocytes showed that wild-type WNK4 expression inhibited surface expression of the thiazide-sensitive NaCl cotransporter (NCC), while a disease-causing mutant lost the inhibitory effect on NCC surface expression. To determine if these changes observed in oocytes really occur in polarized epithelial cells, we generated stable MDCK II cell lines expressing NCC alone or NCC plus wild-type WNK4 or a disease-causing (D564A) WNK4. In contrast to the apical localization of NCC without co-expression of WNK4, immunofluorescence microscopy and biotin surface labeling revealed that this apical localization was equally decreased by both the wild-type and the mutant WNK4 expression. Apical localizations of two PHAII-unrelated apical transporters, sodium-independent amino acid transporter, BAT1 and bile salt export pump, Bsep, were also found to be decreased by both wild-type and mutant WNK4 expression. These results indicate that the regulation of NCC was not related to the disease-causing mutation and not restricted to the PHAII-related specific transporters. The regulation of intracellular localization of NCC by WNK4 might not be involved in the pathogenesis of PHAII.