Evidence for endocytosis of ROMK potassium channel via clathrin-coated vesicles

Zeng, Wei; Babich, Victor; Ortega, Bernardo; Quigley, Raymond; White, S. J.; Welling, Paul A.; Huang, Chou Long

doi:10.1152/ajprenal.00378.2001

Cited by 67 publications

(81 citation statements)

References 43 publications

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“…To maintain K ϩ homeostasis, the density of ROMK in cortical collecting ducts increases and decreases during high and low dietary K ϩ intake, respectively (25,26). Low dietary K ϩ intake decreases ROMK, likely by stimulating endocytosis via CCVs followed by degradation via lysosomes (27,28). Our results suggest that changes in the ratio of long vs. KS-WNK1 in kidney may, at least partly, mediate the regulation of ROMK by dietary K ϩ intake.…”

Section: Discussionmentioning

confidence: 72%

“…Dynamin II may also be involved in internalization via caveolae (29). ROMK1 contains a canonical NPXY motif for internalization via CCV (30), and mutation of the critical asparagine-375 in the motif abolished endocytosis (28). We found that WNK1 failed to inhibit N375I mutant while inhibiting wild-type ROMK1 (Fig.…”

Section: Long Wnk1 Inhibits Romk1 By Stimulating Endocytosis Of the Cmentioning

confidence: 83%

“…ROMK1 undergoes clathrin-coated vesicle (CCV)-mediated endocytosis, a process believed to be an important mechanism for regulating K ϩ secretion in physiological and͞or pathophysiological states (27,28). To determine whether WNK1 inhibits ROMK1 by stimulating CCV-mediated endocytosis of the channel, we examined the effect of WNK1 on ROMK1 by coexpression with wild-type or a dominant-negative dynamin II.…”

Section: Long Wnk1 Inhibits Romk1 By Stimulating Endocytosis Of the Cmentioning

confidence: 99%

“…To maintain K ϩ homeostasis, the abundance of ROMK in the distal nephron decreases or increases during low or high dietary K ϩ intake, respectively (25,26). Alteration of abundance of ROMK during changes of dietary K ϩ intake involves endocytosis and subsequent degradation of the channel protein (27,28).…”

mentioning

confidence: 99%

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

Lazrak

Liu

Huang³

2006

Proc. Natl. Acad. Sci. U.S.A.

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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...

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Section: Discussionmentioning

confidence: 72%

Section: Long Wnk1 Inhibits Romk1 By Stimulating Endocytosis Of the Cmentioning

confidence: 83%

Section: Long Wnk1 Inhibits Romk1 By Stimulating Endocytosis Of the Cmentioning

confidence: 99%

mentioning

confidence: 99%

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

Lazrak

Liu

Huang³

2006

Proc. Natl. Acad. Sci. U.S.A.

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160

207

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“…Furthermore, we have shown that decreases in ROMK channel activity induced by low K intake are the result of stimulating endocytosis (11). Although the mechanism of ROMK internalization is not completely understood, it is possible that ROMK channels are internalized by a clathrin-dependent mechanism (12,13). Recently, ubiquitination has been shown to be involved in the regulation of endocytosis by attaching one or two ubiquitin molecules to lysine residue of substrate protein (14).…”

mentioning

confidence: 99%

ROMK1 channel activity is regulated by monoubiquitination

Lin

Sterling

Wang

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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The ubiquitination of proteins can signal their degradation, modify their activity or target them to specific membranes or cellular organelles. Here, we show that monoubiquitination regulates the plasma membrane abundance and function of the potassium channel, ROMK. Immunoprecipitation of proteins obtained from renal cortex and outer medulla with ROMK antibody revealed that this channel was monoubiquitinated. To determine the ubiquitin binding site on ROMK1, all intracellular lysine (Lys) residues of ROMK1 were individually mutated to arginine (Arg), and a twoelectrode voltage clamp was used to measure the ROMK1 channel activity in Xenopus oocytes. ROMK1 channel activity increased from 8.1 to 27.2 A only when Lys-22 was mutated to Arg. Furthermore, Western blotting failed to detect the ubiquitinated ROMK1 in oocytes injected with R1K22R. Patch-clamp experiments showed that biophysical properties of R1K22R were identical to those of wild-type ROMK1. Although total protein expression levels of GFP-ROMK1 and GFP-R1K22R in oocytes were similar, confocal microscopy showed that the surface fluorescence intensity in oocytes injected with GFP-R1K22R was higher than that of GFP-ROMK1. In addition, biotin labeling of ROMK1 and R1K22R proteins expressed in HEK293 cells showed increased surface expression of the Lys-22 mutant channel. Finally, expression of R1K22R in COS7 cells significantly stimulated the surface expression of ROMK1. We conclude that ROMK1 can be monoubiquitinated and that Lys-22 is an ubiquitin-binding site. Thus, monoubiquitination of ROMK1 regulates channel activity by reducing the surface expression of channel protein. This finding implicates the linking of a single ubiquitin molecule to channels as an important posttranslational regulatory signal.ubiquitination ͉ inwardly rectifying K channel (Kir.11) ͉ K recycling ͉ collecting duct ͉ thick ascending limb R OMK (KCNJ1) channel is an inwardly rectifying K channel located in the apical membrane of the thick ascending limb (TAL) and of the cortical collecting duct (CCD) (1). ROMK channels in the TAL are responsible for K recycling across the apical membrane and K recycling is essential for maintaining normal function of Na͞Cl͞K cotransporter (2). The importance of ROMK in K recycling in the TAL has been best demonstrated by the finding that loss-function mutations in ROMK cause salt wasting and metabolic alkalosis (Barter's syndrome) (3). ROMK is also responsible for K secretion in the CCD. Although the Ca 2ϩ -dependent maxi K channel has been shown to be involved in K secretion when urinary flow rate in the CCD is high (4, 5), ROMK channels play a principal role in K secretion under normal tubule flow (1, 2, 6).ROMK channel activity is regulated by hormones and dietary K intake by modulating the number of channels in the apical membrane (1,7,8). We and others have demonstrated that stimulation of V2 receptor with vasopressin or an increase in dietary K content increases the apical ROMK channel number in the CCD (9, 10). In contrast, low K intake decreas...

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Potassium

Satlin

2009

Pediatric Nephrology

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Evidence for endocytosis of ROMK potassium channel via clathrin-coated vesicles

Cited by 67 publications

References 43 publications

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

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

ROMK1 channel activity is regulated by monoubiquitination

Potassium

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