Antagonistic regulation of ROMK by long and kidney-specific WNK1 isoforms

Abstract

WNK kinases are serine-threonine kinases with an atypical placement of the catalytic lysine. Intronic deletions with increased expression of a ubiquitous long WNK1 transcript cause pseudohypoaldosteronism type 2 (PHA II), characterized by hypertension and hyperkalemia. Here, we report that long WNK1 inhibited ROMK1 by stimulating its endocytosis. Inhibition of ROMK by long WNK1 was synergistic with, but not dependent on, WNK4. A smaller transcript of WNK1 lacking the N-terminal 1-437 amino acids is expressed highly in the kidney. Whether expression of the KS-WNK1 (kidney-specific, KS) is altered in PHA II is not known. We found that KS-WNK1 did not inhibit ROMK1 but reversed the inhibition of ROMK1 caused by long WNK1. Consistent with the lack of inhibition by KS-WNK1, we found that amino acids 1-491 of the long WNK1 were sufficient for inhibiting ROMK. Dietary K ؉ restriction decreases ROMK abundance in the renal cortical-collecting ducts by stimulating endocytosis, an adaptative response important for conservation of K ؉ during K ؉ deficiency. We found that K ؉ restriction in rats increased whole-kidney transcript of long WNK1 while decreasing that of KS-WNK1. Thus, KS-WNK1 is a physiological antagonist of long WNK1. Hyperkalemia in PHA II patients with PHA II mutations may be caused, at least partially, by increased expression of long WNK1 with or without decreased expression of KS-WNK1. There are four mammalian WNK family members (1). WNK1, the first member identified, is Ͼ2,100 amino acids long (2). It contains an Ϸ270-aa kinase domain located near the amino terminus (e.g., amino acids 218-491 of the rat WNK1). WNK2, 3, and 4 are products of different genes and Ϸ1,200 to 1,600 amino acids in length (1, 2). The kinase domain of the four WNKs that share 85-90% sequence identity are unique in having the catalytic lysine located in the subdomain I instead of the conserved subdomain II of most protein kinases (1-3). Other conserved domains of WNK kinases include an autoinhibitory domain, 1-2 coiled-coil domains, and multiple PXXP prolinerich motifs for potential protein-protein interaction (1-4). Beyond the aforementioned conserved domains͞motifs, sequence identity among the four WNKs is much lower and few homologous regions exist.Pseudohypoaldosteronism type II (PHA II) is an autosomaldominant disease characterized by hypertension and hyperkalemia (5). Recently, Wilson et al. (6) reported that mutations of WNK1 and WNK4 cause PHA II. Mutations in the WNK1 gene are large deletions of the first intron leading to increased expression. Mutations in the WNK4 gene are missense mutations in the coding sequence outside the protein kinase domain. Several recent studies have examined the mechanisms for hypertension and hyperkalemia in PHA II patients. WNK4 inhibits the activity of the sodium chloride cotransporter. WNK4 mutants that cause disease fail to inhibit the sodium chloride cotransporter, suggesting that an increase in sodium chloride cotransporter activity in the distal convoluted tubule is a cause of hypertens...

Similar effects of all WNK3 variants on SLC12 cotransporters

“…WNKs are complex kinases that are still poorly understood, but it seems that minimal disruption of their primary structures can prevent or switch their mode of action toward a particular target (13,23,29,32). For instance, although the effect of WNK3-18a on NCC is clearly dependent on its kinase activity, Yang et al (32) have shown that a similar effect can be obtained by the WNK3 carboxyl terminal domain in the complete absence of the kinase domain.…”

Variations in the WNK1 gene modulates the effect of dietary intake of sodium and potassium on blood pressure determination

“…Recent studies have indicated that the expression of the mRNA and protein of KS-WNK1 is significantly increased by a high potassium diet compared with a low potassium diet in rats. 19,20 These findings indicate the possibility that variations in dietary potassium intake cause reciprocal changes in the ratio of L-WNK1 to KS-WNK1 in human kidneys, and consequently in the regulation of renal potassium secretion and sodium reabsorption.…”

“…12 Furthermore, it has been proposed that the molecular switch for the splicing of L-WNK1 and KS-WNK1 is affected by the amount of dietary potassium intake. [19][20][21] In this study, we investigated whether the common variations in the WNK1 gene are potential contributors to individual variations in blood pressure. Furthermore, we analyzed the interactions between the common WNK1 variants and dietary potassium and sodium intake for determining inter-individual variations in blood pressure.…”

Src-family protein tyrosine kinase phosphorylates WNK4 and modulates its inhibitory effect on KCNJ1 (ROMK)

“…WNK1, WNK3, and WNK4 have been shown to inhibit ROMK channels (1,19,20) and the inhibitory effect of WNK4 is achieved by stimulating clathrin-dependent endocytosis (1,21). The inhibitory effect of WNK1 is blocked by a kidney-specific splice form of WNK1 (KS-WNK1) (7,8,22), in which an alternative 5′ exon replaces the first four exons of WNK1 (23). Because expression of KS-WNK1 is modulated by a dietary K intake, interaction between WNK1 and KS-WNK1 may play a role in regulating ROMK channels and renal K secretion (22).…”

“…The inhibitory effect of WNK1 is blocked by a kidney-specific splice form of WNK1 (KS-WNK1) (7,8,22), in which an alternative 5′ exon replaces the first four exons of WNK1 (23). Because expression of KS-WNK1 is modulated by a dietary K intake, interaction between WNK1 and KS-WNK1 may play a role in regulating ROMK channels and renal K secretion (22). The inhibitory effect of WNK4 is also blocked by SGK1, which phosphorylates WNK4 at Ser 1169 and Ser 1196 (2,5), thereby switching WNK4 from inhibition to stimulation of ROMK channels.…”