Consumption of low dietary potassium, common with ultraprocessed foods, activates the thiazide-sensitive sodium chloride cotransporter (NCC) via the with no (K) lysine kinase/STE20/SPS1-related proline-alanine–rich protein kinase (WNK/SPAK) pathway to induce salt retention and elevate blood pressure (BP). However, it remains unclear how high-potassium “DASH-like” diets (dietary approaches to stop hypertension) inactivate the cotransporter and whether this decreases BP. A transcriptomics screen identified Ppp1Ca, encoding PP1A, as a potassium-upregulated gene, and its negative regulator Ppp1r1a, as a potassium-suppressed gene in the kidney. PP1A directly binds to and dephosphorylates NCC when extracellular potassium is elevated. Using mice genetically engineered to constitutively activate the NCC-regulatory kinase SPAK and thereby eliminate the effects of the WNK/SPAK kinase cascade, we confirmed that PP1A dephosphorylated NCC directly in a potassium-regulated manner. Prior adaptation to a high-potassium diet was required to maximally dephosphorylate NCC and lower BP in constitutively active SPAK mice, and this was associated with potassium-dependent suppression of Ppp1r1a and dephosphorylation of its cognate protein, inhibitory subunit 1 (I1). In conclusion, potassium-dependent activation of PP1A and inhibition of I1 drove NCC dephosphorylation, providing a mechanism to explain how high dietary K⁺ lowers BP. Shifting signaling of PP1A in favor of activation of WNK/SPAK may provide an improved therapeutic approach for treating salt-sensitive hypertension.

The sodium–potassium–chloride transporter NKCC1 of the SLC12 family performs Na⁺‐dependent Cl⁻‐ and K⁺‐ion uptake across plasma membranes. NKCC1 is important for regulating cell volume, hearing, blood pressure, and regulation of hyperpolarizing GABAergic and glycinergic signaling in the central nervous system. Here, we present a 2.6 Å resolution cryo‐electron microscopy structure of human NKCC1 in the substrate‐loaded (Na⁺, K⁺, and 2 Cl⁻) and occluded, inward‐facing state that has also been observed for the SLC6‐type transporters MhsT and LeuT. Cl⁻ binding at the Cl1 site together with the nearby K⁺ ion provides a crucial bridge between the LeuT‐fold scaffold and bundle domains. Cl⁻‐ion binding at the Cl2 site seems to undertake a structural role similar to conserved glutamate of SLC6 transporters and may allow for Cl⁻‐sensitive regulation of transport. Supported by functional studies in mammalian cells and computational simulations, we describe a putative Na⁺ release pathway along transmembrane helix 5 coupled to the Cl2 site. The results provide insight into the structure–function relationship of NKCC1 with broader implications for other SLC12 family members.

Key Points: This study provides the first functional documentation of the ghrelin receptor family member GPR39 in the kidney. GPR39 activation directly reduced the urine concentrating capacity by reducing the AVP-induced water permeability of the collecting duct. In parallel, GPR39 activation increased the K ⁺ excretion through a specific reduction of phosphorylated Na ⁺ -Cl ⁻ cotransporter in the distal convoluted tubule. Background: Low-calorie intake is associated with substantial changes in volume distribution and volume status in the body, resulting in reduced circulatory volume and a reduction in BP. Activation of the orphan receptor GPR39 dampens food intake and causes weight loss in a glucagon like peptide-1–dependent fashion. We speculated that appetite-regulating signaling might also be responsible for the circulatory volume contraction observed in response to anorectic states. Methods: To assess the effect of GPR39 fluid homeostasis, we combined in vivo , ex vivo , and in vitro studies to assess the effect of a selective GPR39 agonist (Cpd1324). Results: Oral gavage of Cpd1324 dose-dependently increased the water intake of wild-type (WT) C57BL/6J mice only and was completely absent in global GPR39 knockout mice. GPR39 is expressed in the distal convoluted tubule and collecting duct of the kidney, and WT mice exclusively showed Cpd1324-induced increase in urine production, increased K ⁺ excretion, and reduced urine concentrating capacity both at baseline and after an 8-hour water restriction compared with vehicle controls. Correspondingly, Cpd1324 reduced AVP-induced cAMP production and directly counteracted the AVP-induced water permeability in perfused cortical collecting ducts. Moreover, specific GPR39 activation reduced the baseline and AVP-stimulated abundance of phosphorylated pS256-aquaporin 2 and pT58-Na ⁺ -Cl ⁻ cotransporter and diminished the AVP-stimulated pS269-aquaporin 2 abundance in renal tubular suspensions. These effects were seen exclusively in GPR39 WT mice and not in knockout mice. Conclusions: These data suggest that Cpd1324 directly targets renal GPR39 to induce increased diuresis and consequently stimulate drinking behavior. We conclude that the activation of GPR39 causes diuresis by opposing AVP-induced Na ⁺ and Cl ⁻ reabsorption in the distal convoluted tubule and water reabsorption in the collecting duct.

Loss of bone mass has a devastating effect on quality of life. Higher potassium (K ⁺ ) intake is positively correlated with bone health. Here, we investigated whether kidney calcium (Ca ²⁺ ) and phosphate (Pi) handling mechanisms mediate dietary K ⁺ effects. Kidney Ca ²⁺ and Pi handling proteins were altered in abundance in mice fed a 0% K ⁺ diet for 2 weeks. In mice fed a 0.1% K ⁺ diet for 4 or 8 weeks, urinary Ca ²⁺ excretion increased, plasma Ca ²⁺ levels were lower and plasma parathyroid hormone (PTH) levels were higher relative to control 1% K ⁺ fed mice. The 0.1% K ⁺ fed mice had greater excretion of the bone resorption marker deoxypyridinoline, increased osteoclast number, and decreased total femoral bone mineral density. During chronic low K ⁺ intake, major changes in renal Ca ²⁺ and Pi transport pathways were absent, except higher abundances of the sodium-potassium-chloride cotransporter (NKCC2) and the sodium-chloride cotransporter (NCC), in line with their role in kidney Ca ²⁺ handling. Low dietary K ⁺ induced hypocalcemia and changes in PTH were absent in mice with constitutively active NCC, supporting its role in mediating low K ⁺ effects on Ca ²⁺ homeostasis. Our study provides insights into the management of bone disorders in conditions of chronic electrolyte imbalance.

Hypertension is a global burden and a major contributor to cardiovascular disease and premature death. Public health guidelines for the management of high BP contain numerous dietary recommendations, of which one is a reduction in salt (NaCl) intake. In addition, the modern diet is also characterized by low potassium (K ⁺ ) content and recent guidelines propose increasing K ⁺ intake as an alternative or complementary measure to reducing salt intake for lowering of BP. Most beneficial effects of K ⁺ supplementation on BP involve a homeostatic response of the kidney to dietary-induced changes in extracellular K ⁺ concentrations, particularly decreased reabsorption of NaCl in the distal convoluted tubule. However, the effects of greater K ⁺ intake on BP are not linear, and the ideal K ⁺ supplementation or intake for management of BP remains unclear. This article covers the mechanisms in the kidney by which changes in K ⁺ translate to alterations in BP, the effects of altered K ⁺ intake in animal models and human populations, and the importance of concurrent salt intake and what constitutes K ⁺ supplementation.

NKCC2, localized to the apical membrane of thick ascending limb epithelial cells, is essential for renal salt handling and systemic electrolyte homeostasis. NKCC2 undergoes extensive ubiquitylation, with the E3 protein ligase Nedd4-2 implicated as a key regulator. However, progress has been limited by challenges expressing NKCC2 in mammalian cell lines, hindering mechanistic studies of NKCC2 ubiquitylation. Therefore, the aims of this study were to develop a mammalian cell model enabling mechanistic investigations of NKCC2 ubiquitylation, including the role of Nedd4-2 and the functional consequences of site-specific modification. A tetracycline-inducible MDCKI cell line was generated expressing human NKCC2 and used to assess Nedd4-2-dependent and site-specific ubiquitylation of NKCC2 using biochemical, imaging, and functional assays. The MDCKI cell line demonstrated stable, inducible expression of full-length human NKCC2. In this cell line, mutating the ubiquitylation site at K871 increased membrane abundance and uptake activity, without altering internalization rates. Nedd4-2 co-immunoprecipitated with NKCC2, and Nedd4-2 deletion increased total, but not membrane NKCC2 abundance. In summary, ubiquitylation on NKCC2 at K871 plays a key role in controlling NKCC2 membrane localization and thus function. Although Nedd4-2 can modulate NKCC2 abundance, it is not involved in NKCC2 trafficking. We conclude that the generated cell line provides a robust model for mechanistic studies of NKCC2 and will aid studies examining posttranslational regulation of NKCC2.