Regulation of the Renal NaCl Cotransporter and Its Role in Potassium Homeostasis.

Daily dietary potassium (K+) intake may be as large as the extracellular K+ pool. To avoid acute hyperkalemia, rapid removal of K+ from the extracellular space is essential. This is achieved by translocating K+ into cells and increasing urinary K+ excretion. Emerging data now indicate that the renal thiazide-sensitive NaCl cotransporter (NCC) is critically involved in this homeostatic kaliuretic response. This suggests that the early distal convoluted tubule (DCT) is a K+ sensor that can modify sodium (Na+) delivery to downstream segments to promote or limit K+ secretion. K+ sensing is mediated by the basolateral K+ channels Kir4.1/5.1, a capacity that the DCT likely shares with other nephron segments. Thus, next to K+-induced aldosterone secretion, K+ sensing by renal epithelial cells represents a second feedback mechanism to control K+ balance. NCC's role in K+ homeostasis has both physiological and pathophysiological implications. During hypovolemia, NCC activation by the renin-angiotensin system stimulates Na+ reabsorption while preventing K+ secretion. Conversely, NCC inactivation by high dietary K+ intake maximizes kaliuresis and limits Na+ retention, despite high aldosterone levels. NCC activation by a low-K+ diet contributes to salt-sensitive hypertension. K+-induced natriuresis through NCC offers a novel explanation for the antihypertensive effects of a high-K+ diet. A possible role for K+ in chronic kidney disease is also emerging, as epidemiological data reveal associations between higher urinary K+ excretion and improved renal outcomes. This comprehensive review will embed these novel insights on NCC regulation into existing concepts of K+ homeostasis in health and disease.

A New Phase for WNK Kinase Signaling Complexes as Biomolecular Condensates.

The purpose of this review is to highlight transformative advances that have been made in the field of biomolecular condensates, with special emphasis on condensate material properties, physiology, and kinases, using the With-No-Lysine (WNK) kinases as a prototypical example. To convey how WNK kinases illustrate important concepts for biomolecular condensates, we start with a brief history, focus on defining features of biomolecular condensates, and delve into some examples of how condensates are implicated in cellular physiology (and pathophysiology). We then highlight how WNK kinases, through the action of "WNK droplets" that ubiquitously regulate intracellular volume and kidney-specific "WNK bodies" that are implicated in distal tubule salt reabsorption and potassium homeostasis, exemplify many of the defining features of condensates. Finally, this review addresses the controversies within this emerging field and questions to address.

Deletion of KS-WNK1 promotes NCC activation by increasing WNK1/4 abundance.

Dietary potassium deficiency causes stimulation of sodium reabsorption leading to an increased risk in blood pressure elevation. The distal convoluted tubule (DCT) is the main rheostat linking plasma K+ levels to the activity of the Na-Cl cotransporter (NCC). This occurs through basolateral membrane potential sensing by inwardly rectifying K+ channels (Kir4.1/5.1); decrease in intracellular Cl-; activation of WNK4 and interaction and phosphorylation of STE20/SPS1-related proline/alanine-rich kinase (SPAK); binding of calcium-binding protein 39 (cab39) adaptor protein to SPAK, leading to its trafficking to the apical membrane; and SPAK binding, phosphorylation, and activation of NCC. As kidney-specific with-no-lysine kinase 1 (WNK1) isoform (KS-WNK1) is another participant in this pathway, we examined its function in NCC regulation. We eliminated KS-WNK1 specifically in the DCT and demonstrated increased expression of WNK4 and long WNK1 (L-WNK1) and increased phosphorylation of NCC. As in other KS-WNK1 models, the mice were not hyperkalemic. Although wild-type mice under low-dietary K+ conditions demonstrated increased NCC phosphorylation, the phosphorylation levels of the transporter, already high in KS-WNK1, did not change under the low-K+ diet. Thus, in the absence of KS-WNK1, the transporter lost its sensitivity to low plasma K+. We also show that under low K+ conditions, in the absence of KS-WNK1, there was no formation of WNK bodies. These bodies were observed in adjacent segments, not affected by the targeting of KS-WNK1. As our data are overall consistent with those of the global KS-WNK1 knockout, they indicate that the DCT is the predominant segment affecting the salt transport regulated by KS-WNK1.NEW & NOTEWORTHY In this paper, we show that KS-WNK1 is a critical component of the distal convoluted tubule (DCT) K+ switch pathway. Its deletion results in an inability of the DCT to sense changes in plasma potassium. Absence of KS-WNK1 leads to abnormally high levels of WNK4 and L-WNK1 in the DCT, resulting in increased Na-Cl phosphorylation and function. Our data are consistent with KS-WNK1 targeting WNK4 and L-WNK1 to degradation.

WNK Inhibition Increases Surface Liquid pH and Host Defense in Cystic Fibrosis Airway Epithelia.

In cystic fibrosis (CF), reduced HCO3- secretion acidifies the airway surface liquid (ASL), and the acidic pH disrupts host defenses. Thus, understanding the control of ASL pH (pHASL) in CF may help identify novel targets and facilitate therapeutic development. In diverse epithelia, the WNK (with-no-lysine [K]) kinases coordinate HCO3- and Cl- transport, but their functions in airway epithelia are poorly understood. Here, we tested the hypothesis that WNK kinases regulate CF pHASL. In primary cultures of differentiated human airway epithelia, inhibiting WNK kinases acutely increased both CF and non-CF pHASL. This response was HCO3- dependent and involved downstream SPAK/OSR1 (Ste20/SPS1-related proline-alanine-rich protein kinase/oxidative stress responsive 1 kinase). Importantly, WNK inhibition enhanced key host defenses otherwise impaired in CF. Human airway epithelia expressed two WNK isoforms in secretory cells and ionocytes, and knockdown of either WNK1 or WNK2 increased CF pHASL. WNK inhibition decreased Cl- secretion and the response to bumetanide, an NKCC1 (sodium-potassium-chloride cotransporter 1) inhibitor. Surprisingly, bumetanide alone or basolateral Cl- substitution also alkalinized CF pHASL. These data suggest that WNK kinases influence the balance between transepithelial Cl- versus HCO3- secretion. Moreover, reducing basolateral Cl- entry may increase HCO3- secretion and raise pHASL, thereby improving CF host defenses.

Activation of the kidney sodium chloride cotransporter by the ß2-adrenergic receptor agonist salbutamol increases blood pressure.

The thiazide-sensitive sodium-chloride-cotransporter (NCC) in the kidney distal convoluted tubule (DCT) plays an essential role in sodium and potassium homeostasis. Here, we demonstrate that NCC activity is increased by the ß2-adrenoceptor agonist salbutamol, a drug prevalently used to treat asthma. Relative to ß1-adrenergic receptors, the ß2-adrenergic receptors were greatly enriched in mouse DCT cells. In mice, administration of salbutamol increased NCC phosphorylation (indicating increased activity) within 30 minutes but also caused hypokalemia, which also increases NCC phosphorylation. In ex vivo kidney slices and isolated tubules, salbutamol increased NCC phosphorylation in the pharmacologically relevant range of 0.01-10 µM, an effect observed after 15 minutes and maintained at 60 minutes. Inhibition of the inwardly rectifying potassium channel (Kir) 4.1 or the downstream with-no-lysine kinases (WNKs) and STE20/SPS1-related proline alanine-rich kinase (SPAK) pathway greatly attenuated, but did not prevent, salbutamol-induced NCC phosphorylation. Salbutamol increased cAMP in tubules, kidney slices and mpkDCT cells (model of DCT). Phosphoproteomics indicated that protein phosphatase 1 (PP1) was a key upstream regulator of salbutamol effects. A role for PP1 and the PP1 inhibitor 1 (I1) was confirmed in tubules using inhibitors of PP1 or kidney slices from I1 knockout mice. On normal and high salt diets, salbutamol infusion increased systolic blood pressure, but this increase was normalized by thiazide suggesting a role for NCC. Thus, ß2-adrenergic receptor signaling modulates NCC activity via I1/PP1 and WNK-dependent pathways, and chronic salbutamol administration may be a risk factor for hypertension.

Dysfunction of Cullin 3 RING E3 ubiquitin ligase causes vasoconstriction and increased sodium reabsorption in diabetes.

Impaired endothelium-mediated vasodilation and/or increased sensitivity to vasoconstrictors lead to vascular smooth muscle cell (VSMC) dysfunction in individuals with diabetes. Diabetic nephropathy is associated with a considerably higher risk of cardiovascular disease and death than their nondiabetic counterparts. We studied the activity of Cullin 3 RING ubiquitin ligase (CRL3) and its substrates in mice using an intraperitoneal injection of streptozotocin (STZ) and db/db mice. The levels of CRL3 adaptors, including Kelch-like 2/3 (KLHL2/3) and Rho-related BTB domain-containing protein 1, were significantly decreased in the aortic tissues and heart of the STZ group, whereas the levels of Cullin 3 (CUL3) and its neddylated derivatives were substantially increased. Decreased KLHL3 expression and significantly increased expression of NEDD8 conjugates were observed in the kidneys of db/db mice. The neddylation inhibitor MLN4924 decreased the degradation of KLHL2/KLHL3 under high-glucose conditions with/without insulin, and transfection with KLHL2 promoted the degradation of its substrates with-no-lysine (WNK) kinases. Increased abundance of WNK3, RhoA/ROCK activity and phosphodiesterase 5 enhanced the sensibility to vasoconstrictors and impaired vasodilation. Moreover, WNK3 localized in VSMCs undergoing cell division, and high-glucose medium increased WNK3 signaling in VSMCs undergoing mitosis, which might explain the increased thickness of aortic tissues in subjects with diabetes. Increases in WNK4 abundance resulted in increased sodium reabsorption in the distal renal tubules. Thus, KLHL2/RhoBTB1/KLHL3 inactivation in the aortic tissues and kidney is a result of excessive activation of neddylation in hyperglycemia and hyperinsulinemia, which affects vascular tone and sodium reabsorption.

Severe Arterial Hypertension from Cullin 3 Mutations Is Caused by Both Renal and Vascular Effects.

BACKGROUND: Mutations in four genes, WNK lysine deficient protein kinase 1 and 4 (WNK1 and WNK4), kelch like family member 3 (KLHL3), or Cullin 3 (CUL3), can result in familial hyperkalemic hypertension (FHHt), a rare Mendelian form of human arterial hypertension. Although all mutations result in an increased abundance of WNK1 or WNK4, all FHHt-causing CUL3 mutations, resulting in the skipping of exon 9, lead to a more severe phenotype. METHODS: We created and compared two mouse models, one expressing the mutant Cul3 protein ubiquitously (pgk-Cul3∆9) and the other specifically in vascular smooth muscle cells (SM22-Cul3∆9). We conducted pharmacologic investigations on isolated aortas and generated stable and inducible HEK293 cell lines that overexpress the wild-type Cul3 or mutant Cul3 (Cul3∆9) protein. RESULTS: As expected, pgk-Cul3∆9 mice showed marked hypertension with significant hyperkalemia, hyperchloremia and low renin. BP increased significantly in SM22-Cul3∆9 mice, independent of any measurable effect on renal transport. Only pgk-Cul3∆9 mice displayed increased expression of the sodium chloride cotransporter and phosphorylation by the WNK-SPAK kinases. Both models showed altered reactivity of isolated aortas to phenylephrine and acetylcholine, as well as marked acute BP sensitivity to the calcium channel blocker amlodipine. Aortas from SM22-Cul3∆9 mice showed increased expression of RhoA, a key molecule involved in regulation of vascular tone, compared with aortas from control mice. We also observed increased RhoA abundance and t1/2 in Cul3∆9-expressing cells, caused by decreased ubiquitination. CONCLUSIONS: Mutations in Cul3 cause severe hypertension by affecting both renal and vascular function, the latter being associated with activation of RhoA.

Molecular mechanisms for the regulation of blood pressure by potassium.

It has been well documented that the amount of potassium in the diet is associated with blood pressure levels in the population: the higher the potassium consumption, the lower the blood pressure and the cardiovascular mortality. In the last few years certain mechanisms for potassium regulation of salt reabsorption in the kidney have been elucidated at the molecular level. In this work we discuss the evidence demonstrating the relationship between potassium intake and blood pressure levels in human populations and in animal models, as well as the experimental data that reveal the effects of potassium on transepithelial Na+ reabsorption in different nephron segments. We also discuss the physiological relevance of K+-induced natriuresis, and finally, we focus on the molecular mechanisms by which extracellular potassium modulates the activity of the renal NaCl cotransporter, which is the mechanism that has been best dissected so far.

Renal potassium physiology: integration of the renal response to dietary potassium depletion.

We summarize the current understanding of the physiology of the renal handling of potassium (K+), and present an integrative view of the renal response to K+ depletion caused by dietary K+ restriction. This renal response involves contributions from different nephron segments, and aims to diminish the rate of excretion of K+ as a result of: decreasing the rate of electrogenic (and increasing the rate of electroneutral) reabsorption of sodium in the aldosterone-sensitive distal nephron (ASDN), decreasing the abundance of renal outer medullary K+ channels in the luminal membrane of principal cells in the ASDN, decreasing the flow rate in the ASDN, and increasing the reabsorption of K+ in the cortical and medullary collecting ducts. The implications of this physiology for the association between K+ depletion and hypertension, and K+ depletion and formation of calcium kidney stones are discussed.

WNK signalling pathways in blood pressure regulation.

Hypertension (high blood pressure) is a major public health problem affecting more than a billion people worldwide with complications, including stroke, heart failure and kidney failure. The regulation of blood pressure is multifactorial reflecting genetic susceptibility, in utero environment and external factors such as obesity and salt intake. In keeping with Arthur Guyton's hypothesis, the kidney plays a key role in blood pressure control and data from clinical studies; physiology and genetics have shown that hypertension is driven a failure of the kidney to excrete excess salt at normal levels of blood pressure. There is a number of rare Mendelian blood pressure syndromes, which have shed light on the molecular mechanisms involved in dysregulated ion transport in the distal kidney. One in particular is Familial hyperkalemic hypertension (FHHt), an autosomal dominant monogenic form of hypertension characterised by high blood pressure, hyperkalemia, hyperchloremic metabolic acidosis, and hypercalciuria. The clinical signs of FHHt are treated by low doses of thiazide diuretic, and it mirrors Gitelman syndrome which features the inverse phenotype of hypotension, hypokalemic metabolic alkalosis, and hypocalciuria. Gitelman syndrome is caused by loss of function mutations in the thiazide-sensitive Na/Cl cotransporter (NCC); however, FHHt patients do not have mutations in the SCL12A3 locus encoding NCC. Instead, mutations have been identified in genes that have revealed a key signalling pathway that regulates NCC and several other key transporters and ion channels in the kidney that are critical for BP regulation. This is the WNK kinase signalling pathway that is the subject of this review.

An unexpected journey: conceptual evolution of mechanoregulated potassium transport in the distal nephron.

Flow-induced K secretion (FIKS) in the aldosterone-sensitive distal nephron (ASDN) is mediated by large-conductance, Ca(2+)/stretch-activated BK channels composed of pore-forming α-subunits (BKα) and accessory ß-subunits. This channel also plays a critical role in the renal adaptation to dietary K loading. Within the ASDN, the cortical collecting duct (CCD) is a major site for the final renal regulation of K homeostasis. Principal cells in the ASDN possess a single apical cilium whereas the surfaces of adjacent intercalated cells, devoid of cilia, are decorated with abundant microvilli and microplicae. Increases in tubular (urinary) flow rate, induced by volume expansion, diuretics, or a high K diet, subject CCD cells to hydrodynamic forces (fluid shear stress, circumferential stretch, and drag/torque on apical cilia and presumably microvilli/microplicae) that are transduced into increases in principal (PC) and intercalated (IC) cell cytoplasmic Ca(2+) concentration that activate apical voltage-, stretch- and Ca(2+)-activated BK channels, which mediate FIKS. This review summarizes studies by ourselves and others that have led to the evolving picture that the BK channel is localized in a macromolecular complex at the apical membrane, composed of mechanosensitive apical Ca(2+) channels and a variety of kinases/phosphatases as well as other signaling molecules anchored to the cytoskeleton, and that an increase in tubular fluid flow rate leads to IC- and PC-specific responses determined, in large part, by the cell-specific composition of the BK channels.

Kinase-KCC2 coupling: Cl- rheostasis, disease susceptibility, therapeutic target.

The intracellular concentration of Cl(-) ([Cl(-)]i) in neurons is a highly regulated variable that is established and modulated by the finely tuned activity of the KCC2 cotransporter. Despite the importance of KCC2 for neurophysiology and its role in multiple neuropsychiatric diseases, our knowledge of the transporter's regulatory mechanisms is incomplete. Recent studies suggest that the phosphorylation state of KCC2 at specific residues in its cytoplasmic COOH terminus, such as Ser940 and Thr906/Thr1007, encodes discrete levels of transporter activity that elicit graded changes in neuronal Cl(-) extrusion to modulate the strength of synaptic inhibition via Cl(-)-permeable GABAA receptors. In this review, we propose that the functional and physical coupling of KCC2 to Cl(-)-sensitive kinase(s), such as the WNK1-SPAK kinase complex, constitutes a molecular "rheostat" that regulates [Cl(-)]i and thereby influences the functional plasticity of GABA. The rapid reversibility of (de)phosphorylation facilitates regulatory precision, and multisite phosphorylation allows for the control of KCC2 activity by different inputs via distinct or partially overlapping upstream signaling cascades that may become more or less important depending on the physiological context. While this adaptation mechanism is highly suited to maintaining homeostasis, its adjustable set points may render it vulnerable to perturbation and dysregulation. Finally, we suggest that pharmacological modulation of this kinase-KCC2 rheostat might be a particularly efficacious strategy to enhance Cl(-) extrusion and therapeutically restore GABA inhibition.

Integration of the response to a dietary potassium load: a paleolithic perspective.

Our purpose is to integrate new insights in potassium (K(+)) physiology to understand K(+) homeostasis and illustrate some of their clinical implications. Since control mechanisms that are essential for survival were likely developed in Paleolithic times, we think the physiology of K(+) homeostasis can be better revealed when viewed from what was required to avoid threats and achieve balance in Paleolithic times. Three issues will be highlighted. First, we shall consider the integrative physiology of the gastrointestinal tract and the role of lactic acid released from enterocytes following absorption of sugars (fruit and berries) to cause a shift of this K(+) load into the liver. Second, we shall discuss the integrative physiology of WNK kinases and modulation of delivery of bicarbonate to the distal nephron to switch the aldosterone response from sodium chloride retention to K(+) secretion when faced with a K(+) load. Third, we shall emphasize the role of intra-renal recycling of urea in achieving K(+) homeostasis when the diet contains protein and K(+).

The TSC22D, WNK, and NRBP gene families exhibit functional buffering and evolved with Metazoa for cell volume regulation.

The ability to sense and respond to osmotic fluctuations is critical for the maintenance of cellular integrity. We used gene co-essentiality analysis to identify an unappreciated relationship between TSC22D2, WNK1, and NRBP1 in regulating cell volume homeostasis. All of these genes have paralogs and are functionally buffered for osmo-sensing and cell volume control. Within seconds of hyperosmotic stress, TSC22D, WNK, and NRBP family members physically associate into biomolecular condensates, a process that is dependent on intrinsically disordered regions (IDRs). A close examination of these protein families across metazoans revealed that TSC22D genes evolved alongside a domain in NRBPs that specifically binds to TSC22D proteins, which we have termed NbrT (NRBP binding region with TSC22D), and this co-evolution is accompanied by rapid IDR length expansion in WNK-family kinases. Our study reveals that TSC22D, WNK, and NRBP genes evolved in metazoans to co-regulate rapid cell volume changes in response to osmolarity.

The Integral Role of Chloride & With-No-Lysine Kinases in Cell Volume Regulation & Hypertension.

Chloride anions are the most abundant in humans. For many years, it has been believed that chloride is simply a counterion of all other cations, ensuring the electroneutrality of the extracellular space. Recent data suggests that chloride anions possess a broad spectrum of important activities that regulate vital cellular functions. It is now evident that, apart from its contribution to the electroneutrality of the extracellular space, it acts as an osmole and contributes to extracellular and intracellular volume regulation. Its anionic charge also contributes to the generation of cell membrane potential. The most interesting action of chloride anions is their ability to regulate the activity of with-no-lysine kinases, which in turn regulate the activity of sodium chloride and potassium chloride cotransporters and govern the reabsorption of salt and excretion of potassium by nephron epithelia. Chloride anions seem to play a crucial role in cell functions, such as cell volume regulation, sodium reabsorption in the distal nephron, potassium balance, and sodium sensitivity, which lead to hypertension. All of these functions are accomplished on a molecular level via complicated metabolic pathways, many of which remain poorly defined. We attempted to elucidate some of these pathways in light of recent advances in our knowledge, obtained mainly from experimental studies.

NCC regulation by WNK signal cascade.

With-no-lysine (K) (WNK) kinases have been identified as the causal genes for pseudohypoaldosteronism type II (PHAII), a rare hereditary hypertension condition characterized by hyperkalemia, hyperchloremic metabolic acidosis, and thiazide-hypersensitivity. We thought that clarifying the link between WNK and NaCl cotransporter (NCC) would bring us new mechanism(s) of NCC regulation. For the first time, we were able to produce a knock-in mouse model of PHAII and anti-phosphorylated NCC antibodies against the putative NCC phosphorylation sites and discover that constitutive activation of NCC and increased phosphorylation of NCC are the primary pathogenesis of the disease in vivo. We have since demonstrated that this regulatory mechanism is mediated by the kinases oxidative stress-response protein 1 (OSR1) and STE20/SPS1-related proline/alanine-rich kinase (SPAK) (WNK-OSR1/SPAK-NCC signaling cascade) and that the signaling is not only important in the pathological condition of PHAII but also plays a crucial physiological role in the regulation of NCC.

Modified HEK cells simulate DCT cells in their sensitivity and response to changes in extracellular K.

A potassium (K+ ) rich diet is known to have an antihypertensive effect that has been embodied by the NHLBI in the DASH diet. However, the molecular basis for this blood pressure-lowering effect has been unclear, until a recent study proposed a model in which the DCT cells of the kidney regulate their salt transport in response to variations in intracellular chloride ([Cl- ]i ), which are directly regulated by serum K+ . With the knowledge that WNK proteins are Cl- sensors, and are a part of the WNK/SPAK/NCC signaling cascade which regulates the NCC, the main salt transporter in the distal nephron, we examined the effect of serum K+ on the ([Cl- ]i ) and, in turn its effect on the WNK4 signaling pathway in a "modified HEK 293T" cell line. Using a fluorescence-based approach in this cell line, we have shown that the membrane potential of the cell membrane is sensitive to the small changes in external KCl within the physiological range (2-5 mM), thus functioning as a K+ electrode. When the extracellular K+ was progressively increased (2-5 mM), the membrane depolarization lead to a subsequent increase in [Cl- ]i measured by fluorescence quenching of an intracellular chloride sensor. Increase in extracellular [K] resulted in a decrease in the phosphorylation of the WNK4 protein and its downstream targets, SPAK and NCC. This confirms that small changes in serum K can affect WNK4/SPAK/NCC signaling and transcellular Na+ flux through the DCT and provide a possible mechanism by which a K-rich DASH diet could reduce blood pressure.

ROMK expression remains unaltered in a mouse model of familial hyperkalemic hypertension caused by the CUL3Δ403-459 mutation.

Familial hyperkalemic hypertension (FHHt) is a rare inherited form of salt-dependent hypertension caused by mutations in proteins that regulate the renal Na(+)-Cl(-) cotransporter NCC Mutations in four genes have been reported to cause FHHt including CUL3 (Cullin3) that encodes a component of a RING E3 ligase. Cullin-3 binds to WNK kinase-bound KLHL3 (the substrate recognition subunit of the ubiquitin ligase complex) to promote ubiquitination and proteasomal degradation of WNK kinases. Deletion of exon 9 from CUL3 (affecting residues 403-459, CUL3(Δ403-459)) causes a severe form of FHHt (PHA2E) that is recapitulated closely in a knock-in mouse model. The loss of functionality of CUL3(Δ403-459) and secondary accumulation of WNK kinases causes substantial NCC activation. This accounts for the hypertension in FHHt but the origin of the hyperkalemia is less clear. Hence, we explored the impact of CUL3(Δ403-459) on expression of the distal secretory K channel, ROMK, both in vitro and in vivo. We found that expressing wild-type but not the CUL3(Δ403-459) mutant form of CUL3 prevented the suppression of ROMK currents by WNK4 expressed in Xenopus oocytes. The mutant CUL3 protein was also unable to affect ROMK-EGFP protein expression at the surface of mouse M-1 cortical collecting duct (CCD) cells. The effects of CUL3 on ROMK expression in both oocytes and M-1 CCD cells was reduced by addition of the neddylation inhibitor, MLN4924. This confirms that neddylation is important for CUL3 activity. Nevertheless, in our knock-in mouse model expressing CUL3(Δ403-459) we could not show any alteration in ROMK expression by either western blotting whole kidney lysates or confocal microscopy of kidney sections. This suggests that the hyperkalemia in our knock-in mouse and human PHA2E subjects with the CUL3(Δ403-459) mutation is not caused by reduced ROMK expression in the distal nephron.

American Society of Nephrology Quiz and Questionnaire 2015: Electrolytes and Acid-Base Disorders.

The Nephrology Quiz and Questionnaire remains an extremely popular session for attendees of the annual Kidney Week meeting of the American Society of Nephrology. During the 2015 meeting the conference hall was once again overflowing with eager quiz participants. Topics covered by the experts included electrolyte and acid-base disorders, glomerular disease, end-stage renal disease and dialysis, and kidney transplantation. Complex cases representing each of these categories together with single-best-answer questions were prepared and submitted by the panel of experts. Before the meeting, training program directors of nephrology fellowship programs and nephrology fellows in the United States answered the questions through an internet-based questionnaire. During the live session members of the audience tested their knowledge and judgment on the same series of case-oriented questions in a quiz. The audience compared their answers in real time using a cell-phone app containing the answers of the nephrology fellows and training program directors. The results of the online questionnaire were displayed, and then the quiz answers were discussed. As always, the audience, lecturers, and moderators enjoyed this highly educational session. This article recapitulates the session and reproduces selected content of educational value for theClinical Journal of the American Society of Nephrologyreaders. Enjoy the clinical cases and expert discussions.