WNK3 enhances TRPV5‐mediated calcium transport via a kinase‐dependent pathway

Abstract

WNK3, a member of the With No Lysine (K) family of protein serine/threonine kinases, was shown to regulate members of the SLC12 family of cation‐chloride cotransporters and the renal outer medullary K+ channel ROMK. To evaluate the effect of WNK3 on TRPV5, a renal epithelial Ca2+ channel that serves as a gatekeeper for active Ca2+ reabsorption, WNK3 with TRPV5 were co‐expressed in X. laevis oocytes and the function and expression of TRPV5 were subsequently examined. An 80.6±11.4% increase in TRPV5‐mediated Ca2+ uptake was observed when WNK3 was co‐expressed. Similar increase in TRPV5‐mediated Na+ current was observed with voltage clamp technique. WNK3 also enhanced Ca2+ influx mediated by TRPV6, which is the closest homologue of TRPV5 that mediated active intestinal Ca2+ absorption. The positive regulatory effect was abolished by the kinase‐inactivating D294A mutation in WNK3, indicating a kinase‐dependent mechanism. The fully glycosylated TRPV5 that appears at the plasma membrane was increased by WNK3, indicating that WNK3 increases TRPV5 expression at the cell surface. In the presence of brefeldin A, WNK3 slowed the declining of TRPV5‐mediated Ca2+ uptake over a period of 6 hours, suggesting that WNK3 stabilizes TRPV5 at the plasma membrane. In contrast, WNK3 exerted no significant effect on the delivery of TRPV5 to the plasma membrane from its intracellular pool. These results indicate that WNK3 is a positive regulator of transcellular Ca2+ transport pathway. Supported by NIH grant DK072154.