Regulation of Renal WNK Signaling in Intercalated Cells

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The kidneys play a primary role in the maintenance of potassium homeostasis. The aldosterone sensitive distal nephron (ASDN) regulates potassium balance by matching the rate of urinary K+ excretion with changes in extracellular [K+]. Intercalated cells (ICs) play an important role in this process. Within these cells, apical large conductance BK channels open in response to shear stress to facilitate flow-induced K+ secretion (FIKS), a process activated by hyperkalemia. Though ICs mediate the transcellular movement of K+ from the peritubular interstitium into the tubule lumen, they are unique in that their basolateral K+ entry step is not carried out by the Na+/K+-ATPase. Instead, current evidence suggests that this process requires a bumetanide-sensitive Na+-K+-2Cl- cotransporter, NKCC1 (SLC12A2). NKCC1 is activated via direct phosphorylation, a process mediated by the WNK-SPAK/OSR1 pathway. Consistent with a role for this pathway in FIKS, we find that the kinase active forms of WNK1 and SPAK are stimulated in ICs during hyperkalemia, likely to activate basolateral NKCC1 and apical BK channels. This, however, contradicts the current paradigm, which contends that WNK kinases should be switched off when potassium levels are high in the blood. The overall goal of this application is to determine the importance of NKCC1 and the WNK-SPAK/OSR1 pathway in FIKS, and to test a novel mechanism that explains how these proteins are specifically activated in ICs of the ASDN during hyperkalemia. To accomplish this objective, we will use a variety experimental approaches, including whole animal studies, transport measurements in isolated perfused tubules, fluorescent cell sorting and isolation of ICs derived from whole kidney, and in vitro studies in cell culture models. The information gained from these studies will advance our knowledge of the molecular mechanisms underlying the ASDN’s response to potassium stress.