Kahle Kristopher T, Macgregor Gordon G, Wilson Frederick H, Van Hoek Alfred N, Brown Dennis, Ardito Thomas, Kashgarian Michael, Giebisch Gerhard, Hebert Steven C, Boulpaep Emile L, Lifton Richard P
Department of Genetics, Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, CT 06510, USA.
Proc Natl Acad Sci U S A. 2004 Oct 12;101(41):14877-82. doi: 10.1073/pnas.0406172101. Epub 2004 Oct 1.
Paracellular ion flux across epithelia occurs through selective and regulated pores in tight junctions; this process is poorly understood. Mutations in the kinase WNK4 cause pseudohypoaldosteronism type II (PHAII), a disease featuring hypertension and hyperkalemia. Whereas WNK4 is known to regulate several transcellular transporters and channels involved in NaCl and K+ homeostasis, its localization to tight junctions suggests it might also regulate paracellular flux. We performed electrophysiology on mammalian kidney epithelia with inducible expression of various WNK4 constructs. Induction of wild-type WNK4 reduced transepithelial resistance by increasing absolute chloride permeability. PHAII-mutant WNK4 produced markedly larger effects, whereas kinase-mutant WNK4 had no effect. The electrochemical and pharmacologic properties of these effects indicate they are attributable to the paracellular pathway. The effects of WNK4 persist when induction is delayed until after tight-junction formation, demonstrating a dynamic effect. WNK4 did not alter the flux of uncharged solutes, or the expression or localization of selected tight-junction proteins. Transmission and freeze-fracture electron microscopy showed no effect of WNK4 on tight-junction structure. These findings implicate WNK signaling in the coordination of transcellular and paracellular flux to achieve NaCl and K+ homeostasis, explain PHAII pathophysiology, and suggest that modifiers of WNK signaling may be potent antihypertensive agents.
上皮细胞间离子通量通过紧密连接中的选择性和调节性孔道发生；这一过程目前了解甚少。激酶WNK4的突变会导致II型假性醛固酮增多症（PHAII），这是一种以高血压和高钾血症为特征的疾病。虽然已知WNK4可调节几种参与氯化钠和钾离子稳态的跨细胞转运体和通道，但其定位于紧密连接表明它可能也调节细胞旁通量。我们对可诱导表达各种WNK4构建体的哺乳动物肾上皮细胞进行了电生理学研究。野生型WNK4的诱导通过增加绝对氯离子通透性降低了跨上皮电阻。PHAII突变型WNK4产生的影响明显更大，而激酶突变型WNK4则没有影响。这些效应的电化学和药理学特性表明它们归因于细胞旁途径。当诱导延迟到紧密连接形成之后时，WNK4的效应仍然存在，这表明其具有动态效应。WNK4没有改变不带电荷溶质的通量，也没有改变所选紧密连接蛋白的表达或定位。透射电子显微镜和冷冻蚀刻电子显微镜显示WNK4对紧密连接结构没有影响。这些发现表明WNK信号在协调跨细胞和细胞旁通量以实现氯化钠和钾离子稳态中起作用，解释了PHAII的病理生理学，并表明WNK信号调节剂可能是有效的抗高血压药物。
