Renal Tubule Nedd4-2 Deficiency Stimulates Kir4.1/Kir5.1 and Thiazide-Sensitive NaCl Cotransporter in Distal Convoluted Tubule.
Renal Tubule Nedd4-2 Deficiency Stimulates Kir4.1/Kir5.1 and Thiazide-Sensitive NaCl Cotransporter in Distal Convoluted Tubule.
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DOI：
10.1681/asn.2019090923
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发表时间：
2020-04
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影响因子：
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通讯作者：
Peng Wu;X. Su;Zhong-Xiuzi Gao;Dan-dan Zhang;Xin-Peng Duan;Yu Xiao;O. Staub;Wen‐Hui Wang;Daohong Lin
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文献类型：
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作者：
Peng Wu;X. Su;Zhong-Xiuzi Gao;Dan-dan Zhang;Xin-Peng Duan;Yu Xiao;O. Staub;Wen‐Hui Wang;Daohong Lin
背景钾通道Kir4.1在远曲小管（DCT）基底外侧膜形成Kir4.1/Kir5.1异源四聚体，在噻嗪敏感性NaCl协同转运蛋白（NCC）的调节中起重要作用。肾脏特异性缺失泛素连接酶Nedd 4 -2增加NCC的表达，Nedd 4 -2的共表达抑制Kir4.1/Kir5.1。Nedd 4 -2是否部分通过调节DCT中Kir4.1/Kir5.1通道活性来调节NCC表达尚不清楚。方法我们使用电生理学研究、免疫印迹、免疫染色和肾清除率来检测野生型小鼠和肾脏特异性敲除Nedd 4 -2、Kir4.1或两者的小鼠中DCT中Kir4.1/Kir5.1活性和NCC表达/活性。结果Nedd 4 -2的缺失增加了DCT中Kir4.1的活性/表达，并且使DCT膜超极化。磷酸化NCC/总NCC的表达和噻嗪诱导的尿钠排泄在Nedd 4 -2敲除小鼠中显著增加，但这些小鼠的血钾正常。肾脏中缺乏Kir4.1/Kir5.1和Nedd 4 -2的双敲除小鼠表现出上皮钠通道α亚基表达增加，基底外侧钾离子电导基本消失（与肾脏特异性Kir4.1敲除小鼠的程度相似），DCT膜去极化。与野生型小鼠相比，双基因敲除小鼠显示磷酸化NCC和总NCC的表达受到抑制，噻嗪诱导的尿钠排泄以及肾钾消耗和低钾血症显著减弱。然而，NCC表达/活性在双敲除小鼠中高于Kir4.1敲除小鼠。结论Nedd 4 -2通过Kir4.1依赖性和Kir4.1非依赖性机制调节DCT中Kir4.1/Kir5.1的表达/活性和NCC的表达。DCT中的基底外侧Kir4.1/Kir5.1活性部分地解释了由Nedd 4 -2缺失诱导的NCC活性/表达的刺激。
BACKGROUND The potassium channel Kir4.1 forms the Kir4.1/Kir5.1 heterotetramer in the basolateral membrane of the distal convoluted tubule (DCT) and plays an important role in the regulation of the thiazide-sensitive NaCl cotransporter (NCC). Kidney-specific deletion of the ubiquitin ligase Nedd4-2 increases expression of NCC, and coexpression of Nedd4-2 inhibits Kir4.1/Kir5.1 in vitro. Whether Nedd4-2 regulates NCC expression in part by regulating Kir4.1/Kir5.1 channel activity in the DCT is unknown. METHODS We used electrophysiology studies, immunoblotting, immunostaining, and renal clearance to examine Kir4.1/Kir5.1 activity in the DCT and NCC expression/activity in wild-type mice and mice with kidney-specific knockout of Nedd4-2, Kir4.1, or both. RESULTS Deletion of Nedd4-2 increased the activity/expression of Kir4.1 in the DCT and also, hyperpolarized the DCT membrane. Expression of phosphorylated NCC/total NCC and thiazide-induced natriuresis were significantly increased in the Nedd4-2 knockout mice, but these mice were normokalemic. Double-knockout mice lacking both Kir4.1/Kir5.1 and Nedd4-2 in the kidney exhibited increased expression of the epithelial sodium channel α-subunit, largely abolished basolateral potassium ion conductance (to a degree similar to that of kidney-specific Kir4.1 knockout mice), and depolarization of the DCT membrane. Compared with wild-type mice, the double-knockout mice displayed inhibited expression of phosphorylated NCC and total NCC and had significantly blunted thiazide-induced natriuresis as well as renal potassium wasting and hypokalemia. However, NCC expression/activity was higher in the double-knockout mice than in Kir4.1 knockout mice. CONCLUSIONS Nedd4-2 regulates Kir4.1/Kir5.1 expression/activity in the DCT and modulates NCC expression by Kir4.1-dependent and Kir4.1-independent mechanisms. Basolateral Kir4.1/Kir5.1 activity in the DCT partially accounts for the stimulation of NCC activity/expression induced by deletion of Nedd4-2.