Department of Pharmacology, New York Medical College, Valhalla, New York, USA.
Kidney Int. 2011 Feb;79(4):423-31. doi: 10.1038/ki.2010.380. Epub 2010 Oct 6.
ROMK1 channels are located in the apical membrane of the connecting tubule and cortical collecting duct and mediate the potassium secretion during normal dietary intake. We used a perforated whole-cell patch clamp to explore the effect of angiotensin II on these channels in HEK293 cells transfected with green fluorescent protein (GFP)-ROMK1. Angiotensin II inhibited ROMK1 channels in a dose-dependent manner, an effect abolished by losartan or by inhibition of protein kinase C. Furthermore, angiotensin II stimulated a protein kinase C-sensitive phosphorylation of tyrosine 416 within c-Src. Inhibition of protein tyrosine kinase attenuated the effect of angiotensin II. Western blot studies suggested that angiotensin II inhibited ROMK1 channels by enhancing its tyrosine phosphorylation, a notion supported by angiotensin II's failure to inhibit potassium channels in cells transfected with the ROMK1 tyrosine mutant (R1Y337A). However, angiotensin II restored the with-no-lysine kinase-4 (WNK4)-induced inhibition of R1Y337A in the presence of serum-glucocorticoids-induced kinase 1 (SGK1), which reversed the inhibitory effect of WNK4 on ROMK1. Moreover, protein tyrosine kinase inhibition abolished the angiotensin II-induced restoration of WNK4-mediated inhibition of ROMK1. Angiotensin II inhibited ROMK channels in the cortical collecting duct of rats on a low sodium diet, an effect blocked by protein tyrosine kinase inhibition. Thus, angiotensin II inhibits ROMK channels by two mechanisms: increasing tyrosine phosphorylation of the channel and synergizing the WNK4-induced inhibition. Hence, angiotensin II may have an important role in suppressing potassium secretion during volume depletion.
ROMK1 通道位于连接小管和皮质集合管的顶端膜，在正常饮食摄入期间介导钾分泌。我们使用穿孔全细胞膜片钳技术，在转染绿色荧光蛋白（GFP）-ROMK1 的 HEK293 细胞中探索血管紧张素 II 对这些通道的影响。血管紧张素 II 以剂量依赖性方式抑制 ROMK1 通道，该作用被 losartan 或蛋白激酶 C 抑制所消除。此外，血管紧张素 II 刺激 c-Src 内酪氨酸 416 的蛋白激酶 C 敏感磷酸化。蛋白酪氨酸激酶抑制减弱了血管紧张素 II 的作用。Western blot 研究表明，血管紧张素 II 通过增强其酪氨酸磷酸化来抑制 ROMK1 通道，这一观点得到血管紧张素 II 未能抑制转染 ROMK1 酪氨酸突变体（R1Y337A）的细胞中钾通道的支持。然而，血管紧张素 II 在存在血清糖皮质激素诱导激酶 1（SGK1）的情况下恢复了 WNK4 诱导的 R1Y337A 的抑制作用，这逆转了 WNK4 对 ROMK1 的抑制作用。此外，蛋白酪氨酸激酶抑制消除了血管紧张素 II 诱导的 WNK4 介导的 ROMK1 抑制的恢复。血管紧张素 II 抑制低钠饮食大鼠皮质集合管中的 ROMK 通道，该作用被蛋白酪氨酸激酶抑制所阻断。因此，血管紧张素 II 通过两种机制抑制 ROMK 通道：增加通道的酪氨酸磷酸化和协同 WNK4 诱导的抑制。因此，血管紧张素 II 在容量耗竭期间抑制钾分泌可能具有重要作用。
