Alternative titles; symbols
HGNC Approved Gene Symbol: WNK4
Cytogenetic location: 17q21.2 Genomic coordinates (GRCh38) : 17:42,780,610-42,797,066 (from NCBI)
| Location | Phenotype |
Phenotype MIM number |
Inheritance |
Phenotype mapping key |
|---|---|---|---|---|
| 17q21.2 | Pseudohypoaldosteronism, type IIB | 614491 | Autosomal dominant | 3 |
WNK4 is an integrative regulator of renal electrolyte transport whose main target is the thiazide-sensitive Na-Cl cotransporter NCC (SLC12A3; 600968) (Wu and Peng, 2013).
WNK4 is a serine-threonine protein kinase identified by Wilson et al. (2001) because of its homology to WNK1 (605232). Proteins encoded by WNK paralogs show high conservation in the kinase domain and have the distinctive substitution of cysteine for lysine in the active site. The encoded amino acid sequence of WNK4 showed 76% identity to WNK1 across a 370-amino acid segment spanning the kinase domain and the first putative coil domain, 51% identity across an 83-amino acid segment encompassing the C-terminal putative coil domain, and 52% identity across a 102-amino acid segment from residues 604 to 741 of WNK4. The intron-exon boundaries within these domains are conserved between the 2 genes. WNK4 is expressed virtually exclusively in the kidney, where it localizes to the distal convoluted tubule and the cortical collecting duct, adjacent segments of the distal nephron that play a key role in salt, water, potassium, and pH homeostasis. WNK4 is present exclusively in intracellular junctions in the distal convoluted tubule and in both the cytoplasm and intercellular junctions in the cortical collecting duct. WNK4 colocalizes with ZO1 (TJP1; 601009), a tight junction protein, but not with vinculin (193065), an adherens junction protein. Thus, Wilson et al. (2001) concluded that WNK4 is part of the tight junction complex.
By immunohistochemical analysis, Ring et al. (2007) showed Wnk4 expression in mouse distal colon and distal nephron.
The clinical features of type II pseudohypoaldosteronism (PHAII; see 145260), i.e., hypertension, hyperkalemia, hyperchloremia, and metabolic acidosis, all depend on chloride anion. Furthermore, the observation that pseudohypoaldosteronism phenotypes resulting from WNK mutations constitute a 'mirror image' of the phenotypes resulting from loss-of-function mutations in NCC suggests that PHAII may result from increased SLC12A3 activity due to altered WNK signaling. By studies with heterologous expression of wildtype and mutant mouse Wnk4 in Xenopus oocytes, Wilson et al. (2003) demonstrated that Slc12a3 activity was inhibited by wildtype but not by mutant Wnk4. Wildtype Wnk4 inhibited Slc12a3 by reducing its expression on the membrane. Inhibition depended on Wnk4 kinase activity, because missense mutations that abrogated kinase function prevented the inhibition. PHAII-causing missense mutations further from the kinase domain also prevented the inhibition of Slc12a3 activity. Wilson et al. (2003) concluded that WNK4 negatively regulates the surface expression of SLC12A3, and the loss of this regulation may cause an inherited form of hypertension.
Independently, Yang et al. (2003) found that mouse Wnk4 reduced the plasma membrane association of Slc12a3 in injected Xenopus oocytes. However, they found that some Wnk4 mutations that cause PHAII retained the ability to inhibit Slc12a3. In addition, Yang et al. (2003) showed that Wnk1 (605232) did not affect Slc12a3-mediated sodium uptake in oocytes, but coexpression of Wnk1 with both Wnk4 and Slc12a3 restored sodium uptake to levels observed in oocytes expressing Slc12a3 alone.
To investigate the mechanisms by which WNK1 and WNK4 interact to regulate ion transport, Yang et al. (2005) performed experiments in HEK293 cells and Xenopus oocytes which showed that the WNK4 carboxyl terminus mediates SLC12A3 suppression, that the WNK1 kinase domain interacts with the WNK4 kinase domain, and that WNK1 inhibition of WNK4 is dependent on WNK1 catalytic activity and an intact WNK1 protein.
A key question in systems biology is how diverse physiologic processes are integrated to produce global homeostasis. Genetic analysis can contribute by identifying genes that perturb this integration. One system orchestrates renal NaCl and K+ flux to achieve homeostasis of blood pressure and serum K+ concentration. Positional cloning as reported by Wilson et al. (2001) implicated the serine-threonine kinase WNK4 in this process. Wildtype WNK4 inhibits SLC12A3, the renal Na-Cl cotransporter (NCC); WNK4 mutations that cause type IIB pseudohypoaldosteronism relieve this inhibition. This explains the hypertension of that disorder but does not account for the hyperkalemia. By expression in Xenopus oocytes, Kahle et al. (2003) showed that WNK4 also inhibits the renal K+ channel ROMK (KCNJ1; 600359). This inhibition is independent of WNK4 kinase activity and is mediated by clathrin-dependent endocytosis of ROMK, mechanisms distinct from those that characterize WNK4 inhibition of SLC12A3. Notably, the same mutations in WNK4 that relieve SLC12A3 inhibition markedly increase inhibition of ROMK. These findings established WNK4 as a multifunctional regulator of diverse ion transporters; moreover, they explained the pathophysiology of PHAII. The findings also identified WNK4 as a molecular switch that can vary the balance between NaCl reabsorption and K+ secretion to maintain integrated homeostasis.
Kahle et al. (2004) determined that induction of wildtype mammalian Wnk4 reduced transepithelial resistance by increasing the absolute chloride permeability of preformed tight junctions. Wnk4 with a mutation in a residue critical for kinase activity had no effect. Wnk4 did not alter the flux of uncharged solutes, the expression or localization of tight junction proteins, or tight junction structure. Kahle et al. (2004) concluded that WNK4 coordinates tight junction permeability as well as transcellular ion flux to achieve NaCl and K+ homeostasis.
By coexpression in Xenopus oocytes, Ring et al. (2007) found that mouse Wnk4 inhibited the epithelial Na(+) channel (ENaC), the major mediator of aldosterone-sensitive Na(+) absorption, and that inhibition was independent of Wnk4 kinase activity. Inhibition required intact C termini in ENaC beta (SCNN1B; 600760)- and gamma (SCNN1G; 600761)-subunits, which contain PY motifs used to target ENaC for clearance from the plasma membrane. PHAII-causing mutations eliminated the ability of Wnk4 to inhibit ENaC. The colonic epithelium of mice harboring PHAII-mutant Wnk4 showed increased amiloride-sensitive Na(+) flux compared with wildtype littermates. Ring et al. (2007) concluded that ENaC is a downstream WNK4 target and that WNK4 regulates colonic Na(+) absorption.
Ring et al. (2007) showed that mouse Wnk4 is phosphorylated by Sgk1 (602958), a mediator of aldosterone signaling, at a site C-terminal to the second coiled-coil domain. A Wnk4 mutant that mimics phosphorylation at the Sgk1 site alleviated inhibition of both ENaC and ROMK channels, resulting in increased K(+) secretion. Ring et al. (2007) concluded that WNK4 has a role in mediating the aldosterone response to hyperkalemia.
Yang et al. (2007) noted that WNK1, WNK4, and the kidney-specific WNK1 isoform interact to regulate SLC12A3 activity, suggesting that WNKs form a signaling complex. They found that human WNK3 (300358), which is expressed by distal tubule cells, interacted with rodent Wnk1 and Wnk4 to regulate SLC12A3 in cultured kidney cells and Xenopus oocytes. Regulation of SLC12A3 in oocytes resulted from antagonism between WNK3 and Wnk4. A PHAII-associated mutation in Wnk4 exerted a dominant-negative effect on wildtype Wnk4 that mimicked WNK3 excess.
He et al. (2007) showed that mammalian Wnk1 and Wnk4 interacted with the endocytic scaffold protein intersectin-1 (ITSN1; 602442) and that these interactions were crucial for stimulation of Romk1 endocytosis. Stimulation of Romk1 endocytosis by Wnk1 and Wnk4 required their proline-rich motifs, but it did not require their kinase activities. PHAII-causing mutations in Wnk4 enhanced the interactions of Wnk4 with Itsn1 and Romk1, leading to increased endocytosis of Romk1.
Yang et al. (2007) showed that coexpression of rodent Wnk1 and Wnk4 with human CFTR (602421) suppressed CFTR-dependent chloride channel activity in Xenopus oocytes. The effect of Wnk4 was dose dependent, independent of Wnk4 kinase activity, and occurred, at least in part, by reducing CFTR protein abundance at the plasma membrane. In contrast, the effect of Wnk1 on CFTR activity required Wnk1 kinase activity. Moreover, Wnk1 and Wnk4 exhibited additive CFTR inhibition. Mouse Wnk4 with a PHAII-associated mutation was a more potent inhibitor of CFTR activity than wildtype Wnk4.
Using rodent models of salt-induced hypertension and cultured mouse distal collecting tubule cells, Mu et al. (2011) found that beta-2 adrenergic receptor (ADRB2; 109690) stimulation led to decreased transcription of Wnk4. Adrb2 stimulation resulted in cyclic AMP-dependent inhibition of histone deacetylase-8 (HDAC8; 300269), which permitted increased occupancy of the Wnk4 promoter region by acetylated histones H3 and H4 and the binding of glucocorticoid receptor (GCCR; 138040) to negative glucocorticoid-responsive elements in the Wnk4 promoter. Downregulation of Wnk4 caused Ncc and ENaC activation, sodium retention, and salt-induced hypertension.
KLHL3 (605775) and CUL3 (603136) are part of a cullin-RING ubiquitin E3 ligase complex that targets renal electrolyte transporters or their regulators. By injecting constructs encoding human KLHL3, WNK4, and CUL3 and mouse Ncc into Xenopus oocytes, Wu and Peng (2013) found that KLHL3 inhibited the positive effect of WNK4 on Ncc by decreasing WNK4 protein abundance via WNK4 ubiquitination and degradation. KLHL3 had no effect on Ncc or on the coexpressed downstream kinase OSR1 (OXSR1; 604046). Wu and Peng (2013) concluded that KLHL3 is an important adaptor for ubiquitination and proteasome-mediated downregulation of WNK4.
Wilson et al. (2001) determined that the WNK4 gene comprises 19 exons contained within 16 kb of genomic DNA.
Wilson et al. (2001) localized the WNK4 gene to the interval on chromosome 17 between loci D17S250 and D17S579, both of which lie within the minimum genetic interval containing the PHA2B (614491) locus on chromosome 17.
Wilson et al. (2001) identified 4 missense mutations in the WNK4 gene (601844.0001-601844.0004) in patients with pseudohypoaldosteronism type IIB (PHA2B). Three of these are charge-changing substitutions that cluster in a span of 4 amino acids just distal to the first putative coil domain, within a negatively charged 10-amino acid segment that is highly conserved among all members of the WNK family in human as well as orthologs in mouse and rat.
The causes of hypertension have been obscure owing to the physiologic complexity of the trait. Investigation of mendelian forms of high and low blood pressure has implicated variation in renal salt handling in the pathogenesis of hypertension in humans. Pseudohypoaldosteronism type II (see 145260) can be caused by mutations in WNK1 or WNK4. Renal expression of both genes is confined to epithelia of the distal nephron--the distal convoluted tubule (DCT), the connecting tubule (CNT), and collecting duct. These observations suggested and in vitro studies supported the idea that the Wnks might regulate Na+, Cl-, and K+ flux pathways in these nephron segments. Lalioti et al. (2006) showed that renal physiology in mice transgenic for genomic segments harboring wildtype or PHAII mutant Wnk4 is changed in opposite directions: the mice with the PHAII genomic segment had higher blood pressure, hyperkalemia, hypercalciuria, and marked hyperplasia of the DCT, whereas the opposite was true in transgenic mice with the wildtype genomic segment. Genetic deficiency of the Na-Cl cotransporter (NCC) of the DCT reversed phenotypes seen in the mice carrying the PHAII genomic segment, demonstrating that the effects of the PHAII mutation are due to altered NCC activity. These findings established that Wnk4 is a molecular switch that regulates the balance between NaCl reabsorption and K+ secretion by altering the mass and function of the DCT through its effect on NCC.
Ohta et al. (2009) generated Wnk4-hypomorphic mice by deleting exon 7 of the Wnk4 gene. These mice did not show hypokalemia and metabolic alkalosis but did exhibit low blood pressure and increased sodium and potassium excretion under low-salt diet. Phosphorylation of Osr1 (OXSR1; 604046)/Spak (STK39; 607648) and Ncc was significantly reduced in the mutant mice as compared with their wildtype littermates. Protein levels of Romk and maxi-K (KCNMA1; 600150) were not changed, but the epithelial sodium channel (ENaC) appeared to be activated as a compensatory mechanism for the reduced Ncc function. Ohta et al. (2009) concluded that wildtype WNK4 is a positive regulator for the WNK-OSR1/SPAK-NCC cascade and is a potential target of antihypertensive drugs.
In a family with autosomal dominant pseudohypoaldosteronism type IIB (PHA2B; 614491) previously described by Farfel et al. (1978) and further characterized by Mansfield et al. (1997), Wilson et al. (2001) identified a C-to-G substitution in the WNK4 gene resulting in a glutamine-to-glutamic acid substitution at codon 565 (Q565E), within a highly conserved negatively charged 10-amino acid segment.
Mayan et al. (2004) reported an extension of their previously described kindred (Mayan et al., 2002) that contained 34 subjects, 18 of them affected by the Q565E mutation. Hypertension was diagnosed in 13 affected subjects at the age of 31 +/- 12 years. Five of the affected or obligatory affected subjects had stroke, in 4 between the ages of 50 and 62 years. Seven subjects with FHH were diagnosed 27 years previously. All 4 subjects who were normotensive at diagnosis had become hypertensive by follow-up. The mean time between detection of hyperkalemia and appearance of hypertension was 13 years. In the extended kindred, compared with the unaffected subjects, affected subjects had hyperkalemia, low transtubular potassium gradient, hyperchloremia, low bicarbonate, higher aldosterone, and marked suppression of renin. Urinary calcium levels in affected and unaffected subjects were 0.85 +/- 0.27 and 0.28 +/- 0.12 mmol/mmol creatinine, respectively. Hypercalciuria was accompanied by lower serum calcium levels, supporting a mechanism of renal calcium leak. The 6 affected, currently normotensive subjects had the same degree of hyperkalemia, hypercalciuria, and low renin as the affected hypertensive subjects. The authors concluded that in FHH with WNK4 mutations, with time all affected subjects will apparently develop hypertension.
In a family with pseudohypoaldosteronism type IIB (PHA2B; 614491), Wilson et al. (2001) identified a single basepair substitution in the WNK4 gene resulting in a glutamic acid-to-lysine substitution at codon 562 (E562K), within a highly conserved negatively charged 10-amino acid segment.
In a family with pseudohypoaldosteronism type IIB (PHA2B; 614491), Wilson et al. (2001) identified a single basepair substitution in the WNK4 gene resulting in an aspartic acid-to-alanine substitution at codon 564 (D564A), within a highly conserved negatively charged 10-amino acid segment.
In a family with pseudohypoaldosteronism type IIB (PHA2B; 614491), Wilson et al. (2001) identified an arg-to-cys substitution at codon 1185 (R1185C) of the WNK4 gene, a codon just distal to the second putative coil domain. Arginine-1185 is conserved among WNK family members.
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