The kidney plays a critical role in regulating potassium homeostasis by adjusting secretion in the distal nephron to match daily dietary intake. Distal K+ secretion involves K+ efflux through apical ROMK K+ channels, a process that requires Na+ reabsorption via ENaC to provide the electrical driving force for K+ secretion. The activity of the Na+- Cl- cotransporter, NCC, impacts distal K+ secretion by altering Na+ delivery to ENaC. Thus, besides upregulating ROMK, high dietary K+ intake enhances distal K+ secretion by downregulating NCC. WNK1 and WNK4 are protein kinases in which gain-of-function mutations cause an autosomal-dominant hypertension and hyperkalemia syndrome, pseudohypoaldosteronism type II, which occurs at least in part by increasing Na+ reabsorption via NCC and decreasing K+ secretion via ROMK. How dietary K+ intake regulates NCC and how WNK1 and 4 activate NCC and inhibit ROMK remain unresolved. As a part of long-term goal to understand the mechanism of renal Na+ and K+ transport in health and disease, we propose the following studies.
Aim -1 will test the hypothesis that WNK1 and WNK4 both activate NCC in vivo and that it occurs via the OSR1/SPAK kinase cascade. We will examine the role of WNK1 and 4 on NCC in vivo using kidney-specific conditional Wnk1-knockout mice (to circumvent embryonic lethality of global Wnk1-deleted mice) and global Wnk4-deleted mice, respectively. Double knockout mice will be studied to investigate whether WNK1 and 4 are additive or antagonistic. The role of OSR1/SPAK in mediating WNK1/4 regulation of NCC will be investigated based on the ability of expression of catalytically constitutive-active OSR1/SPAK to rescue loss-of-function of NCC in mice lacking Wnk1 and/or Wnk4. The activity and expression of NCC in kidney will be assessed by analyzing the increase of urinary Na+ excretion in response to hydrochlorothiazide and western blot analysis of the total and phosphorylated NCC.
Aim -2 will examine the hypothesis that WNK kinase cascade mediates dietary K+ intake-induced regulation of NCC. Abundant in vitro and cell-based evidence implicate that dietary K+ intake regulates NCC by altering intracellular chloride concentration to affect WNK kinase activity. We will test the hypothesis in vivo using mice carry Wnk4-null alleles and kidney- specific conditional Wnk1-knockout mice. Mice will be fed a normal K+ or low K+ diet and studied for NCC activity.
Aim -3 will test hypothesis that WNK kinases can directly inhibit renal K+ secretion independently of the effect via increasing NCC-mediated Na+ reabsorption using whole animal clearance studies and in vitro microperfusion of cortical collecting ducts isolated from wild-type mice versus mice whose WNKs are deleted but NCC activity is normalized by constitutive-active SPAK/OSR1. These studies using state-of-the-art animal models and physiological approaches will provide important in vivo information for our understanding of the mechanism of renal Na+ and K+ handling in physiological and diseased states.
Abnormality in sodium and potassium transport in the kidney causes many diseases such as hypertension (high blood pressure) and hyperkalemia (high blood potassium levels), which lead to premature death by causing stroke, congestive heart failure, and arrhythmia, etc. Our studies to understand the mechanism of sodium and potassium transport in the healthy and diseased state will provide important information that may lead to prevention and/or treatment of hypertension and hyperkalemia.
| Yang, Yih-Sheng; Xie, Jian; Yang, Sung-Sen et al. (2018) Differential roles of WNK4 in regulation of NCC in vivo. Am J Physiol Renal Physiol 314:F999-F1007 |
| Taylor 4th, Clinton A; An, Sung-Wan; Kankanamalage, Sachith Gallolu et al. (2018) OSR1 regulates a subset of inward rectifier potassium channels via a binding motif variant. Proc Natl Acad Sci U S A 115:3840-3845 |
| Boyd-Shiwarski, Cary R; Shiwarski, Daniel J; Roy, Ankita et al. (2018) Potassium-regulated distal tubule WNK bodies are kidney-specific WNK1 dependent. Mol Biol Cell 29:499-509 |
| Dalton, George D; Xie, Jian; An, Sung-Wan et al. (2017) New Insights into the Mechanism of Action of Soluble Klotho. Front Endocrinol (Lausanne) 8:323 |
