With-No-Lysine Kinase 1 (WNK1) Augments TRPV4 Function in the Aldosterone-Sensitive Distal Nephron

Kidneys play a central role in regulation of potassium homeostasis and maintenance of plasma K⁺ levels within a narrow physiological range. With-no-lysine (WNK) kinases, specifically WNK1 and WNK4, have been recognized to regulate K⁺ balance, in part, by orchestrating maxi K⁺ channel (BK)-dependent K⁺ secretion in the aldosterone-sensitive distal nephron (ASDN), which includes the connecting tubule and collecting duct. We recently demonstrated that the Ca²⁺-permeable TRPV4 channel is essential for BK activation in the ASDN. Furthermore, high K⁺ diet increases TRPV4 activity and expression largely in an aldosterone-dependent manner. In the current study, we aimed to test whether WNK kinases contribute to regulation of TRPV4 activity and its stimulation by aldosterone. Systemic inhibition of WNK with WNK463 (1 mg/kgBW for 3 days) markedly decreased TRPV4-dependent Ca²⁺ influx in freshly isolated split-opened collecting ducts. Aldosterone greatly increased TRPV4 activity and expression in cultured mpkCCD_c14 cells and this effect was abolished in the presence of WNK463. Selective inhibition of WNK1 with WNK-in-11 (400 nM, 24 h) recapitulated the effects of WNK463 on TRPV4-dependent Ca²⁺ influx. Interestingly, WNK-in-11 did not interfere with up-regulation of TRPV4 expression by aldosterone, but prevented translocation of the channel to the apical plasma membrane. Furthermore, co-expression of TRPV4 and WNK1 into Chinese hamster ovary (CHO) cells increased the macroscopic TRPV4-dependent cation currents. In contrast, over-expression of TRPV4 with a dominant negative WNK1 variant (K233M) decreased the whole-cell currents, suggesting both stimulatory and permissive roles of WNK1 in regulation of TRPV4 activity. Overall, we show that WNK1 is essential for setting functional TRPV4 expression in the ASDN at the baseline and in response to aldosterone. We propose that this new mechanism contributes to regulation of K⁺ secretion and, by extension, urinary K⁺ levels to maintain systemic potassium homeostasis.