AMP-activated protein kinase inhibits KCNQ1 channels through regulation of the ubiquitin ligase Nedd4-2 in renal epithelial cells
AMP-activated protein kinase inhibits KCNQ1 channels through regulation of the ubiquitin ligase Nedd4-2 in renal epithelial cells
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DOI：
10.1152/ajprenal.00423.2010
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发表时间：
2010-12-01
影响因子：
4.2
通讯作者：
Hallows, Kenneth R.
中科院分区：
文献类型：
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作者：
Alzamora, Rodrigo;Gong, Fan;Hallows, Kenneth R.
Alzamora R、Gong F、Rondanino C、Lee JK、Smolak C、Pastor-Soler NM、Hallows KR。 AMP 激活的蛋白激酶通过调节肾上皮细胞中的泛素连接酶 Nedd4-2 来抑制 KCNQ1 通道。 Am J Physiol Renal Physiol 299：F1308-F1319，2010。首次发表于 2010 年 9 月 22 日； doi:10.1152/ajprenal.00423.2010.-KCNQ1 K+ 通道在多种生理功能的调节中发挥着关键作用，包括心脏兴奋性、心血管张力和体内电解质稳态。代谢传感器 AMP 激活蛋白激酶 (AMPK) 已被证明可以调节越来越多的离子转运蛋白。为了确定 AMPK 是否调节 KCNQ1，我们研究了 AMPK 激活对非洲爪蟾卵母细胞和集合管上皮细胞中 KCNQ1 电流的影响。 AMPK 激活降低了非洲虎卵母细胞中的 KCNQ1 电流和通道表面表达，但 AMPK 在体外不会磷酸化 KCNQ1，这表明存在间接调节机制。由于最近表明泛素蛋白连接酶 Nedd4-2 抑制 KCNQ1 质膜表达，并且 AMPK 通过 Nedd4-2 调节上皮 Na+ 通道，因此我们检查了 Nedd4-2 在 AMPK 依赖性 KCNQ1 调节中的作用。在共表达显性失活或组成型活性 Nedd4-2 突变体或 Nedd4-2 相互作用缺陷的 KCNQ1 突变体的卵母细胞中，AMPK 的通道抑制被阻断，表明 Nedd4-2 参与 AMPK 对 KCNQ1 的调节。 KCNQ1 在小鼠极化肾皮质集合管 (mpkCCD(c14)) 细胞和大鼠肾脏的基底外侧膜上表达。使用 AMPK 激活剂 AICAR (2 mM) 或二甲双胍 (1 mM) 处理可降低顶部通透极化 mpkCCDc14 细胞的基底外侧 KCNQ1 电流。此外，AICAR 对大鼠肾切片的离体处理诱导 AMPK 激活以及集合管主细胞中基底外侧膜 KCNQ1 的细胞内重新分布。 AICAR 治疗还诱导肾切片匀浆免疫沉淀的 KCNQ1 泛素化增加。这些结果表明 AMPK 通过促进 Nedd4-2 依赖性通道泛素化和质膜修复来抑制 KCNQ1 活性。
Alzamora R, Gong F, Rondanino C, Lee JK, Smolak C, Pastor-Soler NM, Hallows KR. AMP-activated protein kinase inhibits KCNQ1 channels through regulation of the ubiquitin ligase Nedd4-2 in renal epithelial cells. Am J Physiol Renal Physiol 299: F1308-F1319, 2010. First published September 22, 2010; doi:10.1152/ajprenal.00423.2010.-The KCNQ1 K+ channel plays a key role in the regulation of several physiological functions, including cardiac excitability, cardiovascular tone, and body electrolyte homeostasis. The metabolic sensor AMP-activated protein kinase (AMPK) has been shown to regulate a growing number of ion transport proteins. To determine whether AMPK regulates KCNQ1, we studied the effects of AMPK activation on KCNQ1 currents in Xenopus laevis oocytes and collecting duct epithelial cells. AMPK activation decreased KCNQ1 currents and channel surface expression in X. laevis oocytes, but AMPK did not phosphorylate KCNQ1 in vitro, suggesting an indirect regulatory mechanism. As it has been recently shown that the ubiquitin-protein ligase Nedd4-2 inhibits KCNQ1 plasma membrane expression and that AMPK regulates epithelial Na+ channels via Nedd4-2, we examined the role of Nedd4-2 in the AMPK-dependent regulation of KCNQ1. Channel inhibition by AMPK was blocked in oocytes coexpressing either a dominant-negative or constitutively active Nedd4-2 mutant, or a Nedd4-2 interaction-deficient KCNQ1 mutant, suggesting that Nedd4-2 participates in the regulation of KCNQ1 by AMPK. KCNQ1 is expressed at the basolateral membrane in mouse polarized kidney cortical collecting duct (mpkCCD(c14)) cells and in rat kidney. Treatment with the AMPK activators AICAR (2 mM) or metformin (1 mM) reduced basolateral KCNQ1 currents in apically permeabilized polarized mpkCCDc14 cells. Moreover, AICAR treatment of rat kidney slices ex vivo induced AMPK activation and intracellular redistribution of KCNQ1 from the basolateral membrane in collecting duct principal cells. AICAR treatment also induced increased ubiquitination of KCNQ1 immunoprecipitated from kidney slice homogenates. These results indicate that AMPK inhibits KCNQ1 activity by promoting Nedd4-2-dependent channel ubiquitination and retrieval from the plasma membrane.