Regulation of ROMK (Kir1.1) channels: new mechanisms and aspects
http://www.100md.com
《美国生理学杂志》
Department of Pharmacology, New York Medical College, Valhalla, New York
ABSTRACT
This brief review attempts to provide an overview regarding recent developments in the regulation of ROMK channels. Studies performed in ROMK null mice suggest that ROMK cannot only form hometetramers such as the small-conductance (30-pS) K channels but also construct heterotetramers such as the 70-pS K channel in the thick ascending limb (TAL). The expression of ROMK channels in the plasma membrane is regulated by protein tyrosine kinase (PTK), serum and glucorticoid-induced kinase (SGK), and with-no-lysine-kinase 4. PTK is involved in mediating the effect of low K intake on ROMK channel activity. Increases in superoxide anions induced by low dietary K intake are responsible for the stimulation of PTK expression and tyrosine phosphorylation of ROMK channels. Finally, a recent study indicated that ROMK channels can be monoubiquitinated and monoubiquitination regulates the surface expression of ROMK channels.
THE ROMK CHANNEL PLAYS AN important role in K recycling in the thick ascending limb (TAL) and K secretion in the connecting tubule (CT) and cortical collecting duct (CCD) (16, 19, 34, 43, 45, 61). Although maxi-K channels are also possibly involved in the regulation of K secretion in the CCD (66, 67), it is generally agreed that ROMK channels are mainly responsible for K secretion under normal conditions. The regulation of ROMK channels has been extensively studied, and a recent review has provided extensive coverage regarding the regulatory mechanism of ROMK in the past (16). Thus this review is meant to emphasize the newest developments in the field, which may not have been included in previous reviews. Several studies have identified new amino acids that are involved in the regulation of pH sensitivity of ROMK1 channels in addition to lysine residue 80 (14). Also, it has been shown that changes in extracellular K concentrations can affect the sensitivity of ROMK channels to cell pH (9, 49). Although these topics are important to an understanding of the function of ROMK channels, they are beyond the focus of the present review.
ROMK IS A KEY COMPONENT OF THE APICAL K CHANNELS IN THE TAL
Apical K channels play an important role in K recycling, which is essential for maintaining the normal function of the Na-K-Cl cotransporter in the TAL (16, 61). Patch-clamp experiments demonstrated that two types of K channels, a 30- to 40- and a 70- to 80-pS, are expressed in the apical membrane of the TAL (3, 59, 62). It is well established that the 30-pS K channel is related to ROMK because it has similar biophysical properties and regulatory mechanisms to that in native tubules (61). However, it is not clear whether ROMK is also involved in forming the apical 70-pS K channel. The observation that the loss-of-function mutations of ROMK result in severe salt wasting and metabolic alkalosis (Bartter's disease) (34, 51) indicates strongly that ROMK is an important component of both the 30- and 70-pS K channels in the TAL. Because 70-pS K channels could contribute as much as 60–70% of apical K conductance in the TAL of the rat kidney (36), it is hard to imagine that loss-of-function mutations of ROMK could impair the NaCl transport in the TAL if ROMK is only responsible for the construction of the 30-pS K channels. This puzzle is partially resolved by the patch-clamp experiments performed in the TAL from ROMK null mice in which neither 70- nor 30-pS K channels are detected. In addition, ROMK null mice have demonstrated a typical phenotype of Bartter's syndrome: severe salt wasting, metabolic alkalosis, and hypokalemia (34). This suggests strongly that the 70-pS K channel is a heterotetramer including ROMK and an unidentified subunit. Alternatively, deletion of the ROMK gene could indirectly suppress the expression of the 70-pS K channel. However, the first possibility is further suggested by the observation that high K intake doubled the number of the 70-pS K channels while reducing the number of the 30-pS K channels in the TAL from both wild-type and ROMK (+/–) mice (35). It is possible that high K intake facilitates the formation of the 70-pS K channels by decreasing the formation of the 30-pS K channels. This is achieved by increasing reconstruction between ROMK and other unidentified subunits (Fig. 1) to form the heterotetramer (16). The conductance of the heterotetramer K channel could be different from that of the homotetramer. Relevant to this is the acetylcholine-activated K channel, which is composed of GIRK and cardiac inwardly rectifying K (CIR) channels. Expression of CIR alone in oocytes produces a K channel with single-channel conductance of 15 pS, whereas the coexpression of CIR and GIRK forms an acetylcholine-activated K channel with a single-channel conductance of 37 pS (26). It is possible that the 70-pS K channel may be formed by ROMK and another inwardly rectifying K channel. However, the nature of the unidentified subunit of the 70-pS K channel remains to be determined.
The abundance of ROMK channels in the TAL is regulated by hormones and Na diet. It has been shown that infusion of vasopressin into Brattleboro rats for 7 days increased the apical labeling of ROMK in the TAL (13). The role of vasopressin in the regulation of ROMK channel expression is further indicated by the finding that restriction of water intake can also significantly increase ROMK expression in the TAL. Also, ROMK expression is upregulated in the TAL in response to high Na intake (13). This suggests that ROMK channels and Na-K-Cl cotransporters are possibly coordinately regulated in the TAL by vasopressin and Na intake.
NEW ASPECTS OF THE EFFECTS OF PKC AND CFTR ON ROMK
Regulation by PKC
ROMK1 channels have three putative PKC phosphorylation sites: serine residues (Ser) 4, 181, and 201 (19). In vitro phosphorylation has shown that ROMK1 can be phosphorylated by PKC on Ser4 and Ser201 (29). Moreover, stimulation of PKC has been shown to inhibit ROMK channel activity in native tubules (28, 60). However, the K current in oocytes injected with ROMK1, S4D or ROMK1, S4D/S201D in which the serine residue was mutated to aspartate was either unchanged or increased, suggesting that the PKC-induced inhibition of ROMK1 may not be the result of direct phosphorylation. This possibility was further suggested by the finding that stimulation of PKC with phorbol ester reduced phosphatidylinositol 4,5-bisphosphate (PIP2) content in oocyte membrane and inhibition of PKC abolished the phorbol ester-induced decrease in PIP2 (74). Because PIP2 has been shown to maintain ROMK channels in an open state (20, 33), it is possible that decreases in PIP2 concentration induced by stimulating PKC may contribute to the inhibition of ROMK channel activity. In addition, PKC has been shown to regulate the surface expression of ROMK and endocytosis of ROMK induced by stimulation of PTK (29, 54). We demonstrated that mutation of Ser4 and Ser201 to alanine significantly suppressed the surface expression of ROMK1 in oocytes (29). This suggests that direct phosphorylation of ROMK1 by PKC is important for the surface expression of K channels in oocytes. On the other hand, we also observed that inhibition of PKC blocked the internalization of ROMK1 induced by stimulation of PTK (54). This seeming discrepancy may be the result of different PKC targets. It is possible that the role of PKC in mediating the internalization of ROMK channels may be achieved by targeting factors other than ROMK. Relevant to this hypothesis is the observation that PKC is involved in actin reorganization (5). Thus the complexity of PKC's effects on ROMK may be induced because PKC regulates ROMK channel activity by direct and indirect mechanisms.
Regulation by CFTR
The role of CFTR in the regulation of ROMK channel activity has been demonstrated previously. CFTR has been shown to play an important role in mediating sulfonylurea agent-induced inhibition and ATP sensitivity (38, 48), although it has also been reported that ROMK channels have an intrinsic sensitivity to sulfonylurea agents (24). A recent study suggests that CFTR may play a role in mediating the interaction between ROMK and the epithelial Na channel (ENaC). Previous studies have shown the interaction between ENaC and CFTR such that stimulation of CFTR reduced ENaC activity (57), whereas coexpression of ENaC increased CFTR expression (25). Recently, it also has been reported that CFTR may link the activity of ENaC to ROMK. In the presence of CFTR, expression of ROMK channels was augmented in oocytes injected with ENaC (25). Also, the PDZ domain has been shown to facilitate the interaction between ROMK and CFTR (70). In contrast, coexpression of ENaC had no effect on ROMK expression in the absence of CFTR. This suggests that CFTR synchronizes the activity of ENaC and ROMK. However, whether such interaction takes place in the native tubule is not clear and needs to be examined.
SUPEROXIDE IS INVOLVED IN THE REGULATION OF ROMK CHANNEL ACTIVITY
ROMK1 is exclusively located in the CT and CCD (4) and plays an important role in K secretion. A large body of evidence indicates that the number of ROMK channels in the CCD is varied such that high K intake increases (44, 46), whereas low K intake decreases, ROMK channel numbers in the CCD (2). The response of rats to low K is very sensitive because decreases in K content from 1.1 to 0.7% significantly reduced ROMK channel activity in the CCD to an extent not significantly different from that in rats on a K-deficient diet. This could be the reason the previous study failed to detect a significant difference in ROMK channel activity in the CCD between rats on a K-deficient diet and "normal K diet (0.7%)" (64). Several lines of evidence strongly suggest that the PTK-dependent signal transduction pathway plays a key role in mediating the effect of low K intake on ROMK channel activity. First, low K intake significantly increases the expression Src-family PTK such as c-Src and c-Yes in the kidney (64). Second, the Src family PTK is expressed in principal cells of the CCD and CT (32). Third, low K intake enhances whereas high K intake diminishes the tyrosine phosphorylation of ROMK (30). Finally, inhibition of PTK increases whereas blockade of protein tyrosine phosphatase (PTP) decreases the ROMK channel activity in the CCD (63, 64). Studies with confocal microscopy have further indicated that stimulation of PTK-induced phosphorylation of ROMK1 facilitates the internalization (55, 56). These data suggest that low K intake stimulates the activity of Src family PTK which, in turn, enhances the tyrosine phosphorylation of ROMK channels in the CCD. As a consequence, ROMK channels are internalized and the density of ROMK in the apical membrane decreases.
Although the role of PTK in mediating the effect of low K intake on ROMK channel activity has been established, the upstream signaling which regulates PTK activity is not completely understood. A recent study suggests the possible role of superoxide in the regulation of expression of Src family PTK. Using a chemiluminescence technique (68), it was found that low K intake significantly increased the concentration of superoxide anions in the renal cortex and outer medulla (2) and that the superoxide production induced by low K intake was blunted by tempol, a superoxide dismutase mimetic (27). The role of superoxide in mediating the effect of low K intake on PTK expression was further suggested by observations that treatment of M1 cell, a mouse principal cell line, with hydrogen peroxide significantly increased the expression of c-Src. The effect of hydrogen peroxide on c-Src expression was abolished by either cycloheximide or actinomycin A. This indicates that the stimulatory effect of hydrogen peroxide on c-Src expression is the result of increased transcription and translation. Indeed, Western blot analysis revealed that low K intake increased the phosphorylation of c-Jun on Ser73, an indication of activation of c-Jun. The view that increases in superoxide anions may stimulate transcription of PTK was also suggested by experiments in which treatment of M1 cells with hydrogen peroxide significantly increased the serine phosphorylation of c-Jun. Moreover, tempol treatment completely abolished the effect of low K intake on c-Jun activation and c-Src expression in the kidney of rats fed a K-deficient diet.
The possibility that superoxide is involved in the regulation of ROMK channel activity and renal K excretion is supported by several lines of evidence (2). First, lowering superoxide production in the kidney by tempol decreased the tyrosine phosphorylation of ROMK in the renal cortex and outer medulla and increased the ROMK channel activity in the CCD. Second, metabolic cage studies showed that the renal K excretion in the tempol-treated rats was 10 times higher than those in rats on a K-deficient diet alone. Third, animals receiving tempol developed a more severe hypokalemia than those without tempol treatment. Figure 2 is a model of principal cell illustrating the mechanism by which low K intake inhibits ROMK1 channel activity. Low K intake increases the production of superoxide, which, in turn, stimulates PTK expression. As a consequence, tyrosine phosphorylation of ROMK channels in the CCD increases and the ROMK channel is endocytosed. In addition to stimulation of PTK expression, superoxide and the related product have also been shown to stimulate PTK (6, 42) or inhibit the activity of PTP (10, 37). Thus it is possible that superoxide may be able to acutely regulate ROMK channel activity through modulation of PTK and PTP interaction. Indeed, we have observed that acute application of hydrogen peroxide inhibited ROMK channel activity in the rat CCD and that the effect was blocked by herbimycin A (unpublished observations), indicating that the effect of hydrogen peroxide on ROMK channel activity is due to stimulation of PTK activity. Although the role of superoxide in mediating the effect of low K intake on renal K secretion has been suggested, the mechanism by which low K intake stimulates superoxide production is still not clear. Because the major source of superoxide is from activation of NAD(p)H oxidase (12, 15, 39, 40), it would be interesting to determine the role of NAD(p)H oxidase in the regulation of renal K secretion.
REGULATION OF ROMK CHANNEL TRAFFICKING
A large body of evidence indicates that ROMK channel activity is regulated by endocytosis and exocytosis. ROMK channels have been shown to interact with clathrin via AP-2 protein (73). Recently, it has been demonstrated that syntaxin 1A interacts with the COOH terminus of ROMK and that coexpression of syntaxin 1A inhibits ROMK channel activity in oocytes (58). Also, several new mechanisms including monoubiquitination, serum and glucocorticoid-inducible kinase (SGK), and with-no-lysine-kinase 4 (WNK4) have been discovered in the regulation of ROMK expression in plasma membrane.
Monoubiquitination
Ubiquitination plays an important role in the regulation of protein degradation and recycling by attaching ubiquitin molecules to lysine residues of substrate protein (8, 17, 47). Ubiquitination can further be classified into monoubiquitination by adding only one or two ubiquitin molecules to the substrate protein or polyubiquitination by attaching more than four ubiquitin molecules. The polyubiquitinated protein is subjected to degradation, whereas monoubiquitinated proteins are targeted to internalization and possibly recycling to the cell membrane (11, 18, 50). A large body of evidence indicates that polyubiquitination plays an important role in the regulation of cell signaling and ENaC (52, 53). However, the role of monoubiquitination in the regulation of membrane transporters and ion channels is not clear. Recently, it has been shown that ROMK channel activity could be regulated by monoubiquitination (31). First, immunoprecipitation of ROMK from renal tissue or from cell culture revealed a 50-kDa protein band that was also recognized by an ubiquitin antibody. Second, mutation of lysine residue 22 to arginine (R1K22R) in the NH2 terminus of ROMK1 increased ROMK channel activity. Third, no ubiquitinated ROMK can be detected in oocytes injected with a ROMK1 mutant, R1K22R. Moreover, the biophysical properties of R1K22R such as channel conductance and open probability are the same as that of wt ROMK. Thus an increase in ROMK channel activity must be the result of augmentation of membrane expression of ROMK1 channels. This view is also supported by the observation that a higher biotin-labeled ROMK1, an indication of membrane expressed ROMK1, was observed in cells transfected with ROMK1 mutant than those of ROMK1.
The physiological importance of monoubiquitination in the regulation of ROMK channel is still not clear. Because ROMK channel activity in the CCD decreases in response to stimulation of PTK, it would be interesting to determine whether monoubiquitination is required for the PTK-induced internalization of ROMK1 in the CCD. Figure 3 is a model of a cell illustrating a possible role of ubiquitination in the regulation of ROMK trafficking. Stimulation of PTK has been shown to activate ubiquitin ligase activity (23). It is possible that PTK could enhance the ubiquitination of ROMK and accordingly internalization.
SGK1
Aldosterone has been shown to stimulate glucocorticoid-inducible kinase 1 (SGK1) levels (41), which are possibly involved in mediating the effect of aldosterone on ENaC activity (1, 7). SGK1 was first thought to have no effect on ROMK channel activity (7). As a matter of fact, ROMK was used as a negative control to demonstrate that the stimulatory effect of SGK1 on ENaC was specific. However, three separate studies have later demonstrated that SGK1 can also increase the activity of ROMK in the presence of Na/H exchange-regulating factor (NHERF) (70–72). It is possible that NHERF is served as a scaffold protein required for the stimulatory effect of SGK1 on ROMK. However, the role of NHERF, specifically NHERF2 in mediating the SGK effect, is controversial. Yoo et al. (71) observed that SGK could stimulate ROMK channel without NHERH. Moreover, they observed a modest stimulation of ROMK surface expression by NHERF alone. They suggested that NHERF is required for the effect of SGK on ROMK only when abundance of one of them is low. The mechanism of the SGK1 effect may be through stimulation of the serine phosphorylation of ROMK1. Confocal microscopy and cell surface antibody-binding assay have further shown that stimulation of SGK-induced phosphorylation increases the expression of ROMK1 channels in plasma membrane. The phosphorylation site of ROMK1 induced by SGK is located on Ser44 in the NH2 terminus, which is also a putative PKA phosphorylation site (69) because mutation of Ser44 to alanine abolished the effect of SGK (71). Also, Yoo et al. (71) suggested that effect of SGK on ROMK activity in oocytes depends on the basal level of ROMK phosphorylation such that the stimulatory effect of SGK could be blunted when the basal phosphorylation level of ROMK is high. The role of SGK on K secretion is also demonstrated in SGK null mice in which renal K secretion is impaired (21). However, it is still not known whether the stimulatory effect of SGK on renal K secretion is the result of increased ENaC activity that, in turn, augments the electrochemical gradient for K secretion or due to a direct stimulation of ROMK channel insertion. Confocal microscopy has shown that ROMK expression in the apical membrane of the TAL and CCD is actually increased rather than decreased in SGK null mice. This suggests that the role of SGK in stimulating ROMK insertion may not be essential or can be replaced by other kinases such as cAMP-dependent protein kinase A. A possible role of SGK is to regulate renal K secretion in response to a daily dietary K intake. It has been shown that a high dietary K intake for 12 h stimulates SGK expression in the kidney (71). Interestingly, it has been shown that ROMK channel activity significantly increased in the CCD from rats on a high-K diet for only 6 h (46). Thus SGK may regulate ROMK channel activity in the CT and CCD in response to a daily variation in dietary K intake.
WNK4
WNK4 is a serine-threonine kinase and has been identified in both distal convoluted tubule (DCT) and the CCD. Wild-type WNK4 has been shown to suppress the expression of the Na-Cl cotransporter in the plasma membrane of the DCT and, accordingly, to inhibit Na absorption (65). The gene product encoding inactivated WNK4 causes pseudohypoaldosteronism type II, a disease with characteristics of hypertension and low aldosterone levels. WNK4 is also expressed in the CCD, and expression of WNK4 in oocytes decreased the expression of ROMK1 (22). However, the inhibitory effect of WNK4 on ROMK1 does not require the kinase activity because coexpression of inactivated WNK4 also resulted in an inhibition of the expression of ROMK1 in Xenopus laevis oocytes. The physiological role of WNK4 may be that it serves as a switch between Na transport and K secretion. The WNK4-induced inhibition of Na transport in the DCT is expected to increase Na delivery to the CCD and enhance Na absorption, which leads to augmentation of the driving force for K secretion. However, because WNK4 inhibits ROMK channel activity, increases in electrochemical gradient for K secretion would have a diminished effect on renal K secretion. Mutation of WNK4 would release the inhibition of the Na-Cl cotransporter and lead to stimulation of Na transport in the DCT. Moreover, inactivated WNK4 further decreases the surface expression of ROMK, reduces K secretion in the CCD, and causes hyperkalemia. However, it is possible that decreases in Na delivery to the CCD may also be responsible for the diminished K secretion in patients with pseudohypoaldosteronism type II.
SUMMARY
Figure 3 is a model of the principal cell, illustrating the possible mechanisms by which ROMK channel expression in plasma membrane is regulated by WNK4, SGK, and ubiquitin. Stimulation of PTK decreases, whereas activation of SGK1 increases, the expression of ROMK. Also, increased WNK4 expression should diminish ROMK channel activity.
GRANTS
This work is supported by National Institutes of Health Grants DK-54983, DK-47402, and HL-34300.
ACKNOWLEDGMENTS
The author thanks Melody Steinberg for editorial assistance.
FOOTNOTES
REFERENCES
Alvarez de la Rosa D, Zhang P, Naaray-Fejes-Toth A., Fejes-Toth G, and Canessa CM. The serum and glucocorticoid kinase sgk increases the abundance of epithelial sodium channels in the plasma membrane of Xenopus oocytes. J Biol Chem 274: 37834–37839, 1999.
Babilonia E, Wei Y, Sterling H, Kaminski P, and Wolin MS. Superoxide anions are involved in mediating the effect of low K intake on c-Src expression and renal K secretion in the cortical collecting duct. J Biol Chem 280: 10790–10796, 2005.
Bleich M, Schlatter E, and Greger R. The luminal K+ channel of the thick ascending limb of Henle's loop. Pflügers Arch 415: 449–460, 1990.
Boim MA, Ho K, Schuck ME, Bienkowski MJ, Block JH, Slightom JL, Yang Y, Brenner BM, and Hebert SC. The ROMK inwardly rectifying ATP-sensitive K channel. II. Cloning and intra-renal distribution of alternatively spliced forms. Am J Physiol Renal Fluid Electrolyte Physiol 268: F1132–F1140, 1995.
Brandt D, Gimona M, Hillmann M, Haller H, and Mischak H. Protein kinase C induces actin reorganization via a Src- and Rho-dependent pathway. J Biol Chem 277: 20903–20910, 2002.
Brumell JH, Burkhardt AL, Bolen JB, and Grinstein S. Endogenous reactive oxygen intermediates activate tyrosine kinase in human neutrophils. J Biol Chem 271: 1455–1461, 1996.
Chen SY, Bhargava A, Mastroberardino L, Meijer OC, Wang J, Buse P, Firestone GL, Verrey F, and Pearce D. Epithelial sodium channel regulation by aldosterone-induced protein SGK. Proc Natl Acad Sci USA 96: 2514–2519, 1999.
Ciechanover A. The ubiquitin-proteasome proteolytic pathway. Cell 79: 13–21, 1994.
Dahlmann A, Li M, Gao ZH, McGarrigle D, Sackin H, and Palmer LG. Regulation of Kir channels by intracellular pH and extracellular K: mechanism of coupling. J Gen Physiol 123: 441–454, 2004.
Denu JM and Tanner KG. Specific and reversible inactivation of protein tyrosine phosphatases by hydrogen peroxide: evidence for a sulfenic acid intermediate and implications for redox regulation. Biochemistry 37: 5633–5642, 1998.
DiFiore PPD, Polo S, and Hofmann K. When ubiquitin meets ubiquitin receptors: a signalling connection. Nat Rev Mol Cell Biol 4: 491–497, 2003.
Droge W. Free radicals in the physiological control of cell function. Physiol Rev 82: 47–95, 2002.
Ecelbarger CA, Kim CH, Knepper MA, Liu J, Tate M, Welling PA, and Wade JB. Regulation of potassium channel Kir 1.1 (ROMK) abundance in the thick ascending limb of Henle's loop. J Am Soc Nephrol 12: 10–18, 2001.
Fakler B, Schultz JH, Yang J, Schulte U, Braendle U, Zenner HP, Jan LY, and Ruppersberg J. Identification of a titratable lysine residue that determines sensitivity of kidney potassium channels (ROMK) to intracellular pH. EMBO J 15: 4093–4099, 1996.
Geiszt M and Leto T. The nox family of NAD(p)H oxidase: host defense and beyond. J Bio Chem 279: 51715–51718, 2004.
Hebert SC, Desir GV, Giebisch G, and Wang WH. Molecular diversity and regulation of renal potassium channels. Physiol Rev 85: 319–371, 2005.
Hershko A and Ciechanover A. The ubiquitin system for protein degradation. Annu Rev Biochem 61: 761–807, 1992.
Hicke L and Dunn R. Regulation of membrane protein transport by ubiquitin and ubiquitin-binding proteins. Annu Rev Cell Dev Biol 19: 172, 2003.
Ho K, Nichols CG, Lederer WJ, Lytton J, Vassilev PM, Kanazirska MV, and Hebert SC. Cloning and expression of an inwardly rectifying ATP-regulated potassium channel. Nature 362: 31–38, 1993.
Huang CL, Feng S, and Hilgemann DW. Direct activation of inward rectifier potassium channels by PIP2 and its stabilization by G. Nature 391: 803–806, 1998.
Huang DY, Wulff P, Volkl H, Loffing J, Richter K, Kuhl D, Lang F, and Vallon V. Impaired regulation of renal K elimination in the sgk1-knockout mouse. J Am Soc Nephrol 15: 885–891, 2004.
Kahle KT, Wilson FH, Leng Q, Lalioti MD, Connell AD, Dong K, Rapson AK, MacGregor GG, Giebisch G, Hebert SC, and Lifton RP. WNK4 regulates the balance between renal NaCl reabsorption and K secretion. Nat Genet 35: 372–376, 2003.
Kassenbrock CK and Anderson SM. Regulation of ubiquitin protein ligase activity in c-Cbl by phosphorylation-induced conformational change and constitutive activation by tyrosine to glutamate point mutations. J Biol Chem 279: 28017–28027, 2004.
Konstas AA, Dabrowski M, Korbmacher C, and Tucker SJ. Intrinsic sensitivity of Kir1.1 (ROMK) to glibenclamide in the absence of sur2b. J Biol Chem 277: 21346–21351, 2002.
Konstas AA, Koch JP, Tucker SJ, and Korbmacher C. Cystic fibrosis transmembrane conductance regulator-dependent up-regulation of Kir 1.1 (ROMK) renal K channels by the epithelial sodium channel. J Biol Chem 277: 25377–25384, 2002.
Krapivinsky G, Gordon EA, Wickman K, Velimirovic B, Krapivinsky L, and Clapham DE. The G-protein-gated atrial K+ channel I KACh is a heteromultimer of two inwardly rectifying K+-channel proteins. Nature 374: 135–141, 1995.
Krishna MC, Grahame DA, Samuni A, Mitchell JB, and Russo A. Oxoammonium cation intermediate in the nitroxide-catalyzed dismutation of superoxide. Proc Natl Acad Sci USA 89: 5537–5541, 1992.
Kubokawa M, Wang WH, McNicholas CM, and Giebisch G. Role of Ca2+/CaMK II in Ca2+-induced K+ channel inhibition in rat CCD principal cell. Am J Physiol Renal Fluid Electrolyte Physiol 268: F211–F219, 1995.
Lin DH, Sterling H, Lerea KM, Giebisch G, and Wang WH. Protein kinase C (PKC)-induced phosphorylation of ROMK1 is essential for the surface expression of ROMK1 channels. J Biol Chem 277: 44332–44338, 2002.
Lin DH, Sterling H, Lerea KM, Welling P, Jin L, Giebisch G, and Wang WH. K depletion increases the protein tyrosine-mediated phosphorylation of ROMK. Am J Physiol Renal Physiol 283: F671–F677, 2002.
Lin DH, Sterling H, Wang Z, Babilonia E, Yang B, Dong K, Hebert SC, Giebisch G, and Wang WH. ROMK1 channel activity is regulated by monoubiquitination. Proc Natl Acad Sci USA 102: 4306–4311, 2005.
Lin DH, Sterling H, Yang B, Hebert SC, Giebisch G, and Wang WH. Protein tyrosine kinase is expressed and regulates ROMK1 location in the cortical collecting duct. Am J Physiol Renal Physiol 286: F881–F892, 2004.
Liou HH, Zhou SS, and Huang CL. Regulation of ROMK1 channel by protein kinase A via a phosphatidylinositol 4,5-bisphosphate-dependent mechanism. Proc Natl Acad Sci USA 96: 5820–5825, 1999.
Lu M, Wang T, Yan Q, Yang X, Dong K, Knepper MA, Wang WH, Giebisch G, Shull GE, and Hebert SC. Absence of small conductance K channel (SK) activity in apical membrane of thick ascending limb and cortical collecting duct in ROMK (Bartter's) knockout mice. J Biol Chem 277: 37881–37887, 2002.
Lu M, Wang T, Yan QS, Wang WH, Giebisch G, and Hebert SC. ROMK is required for expression of the 70 pS K channel in the thick ascending limb. Am J Physiol Renal Physiol 286: F490–F495, 2004.
Lu M, Zhu Y, Balazy M, Falck JR, and Wang WH. Effect of angiotensin II on the apical K channels in the thick ascending limb of the rat kidney. J Gen Physiol 108: 537–547, 1996.
Mahadev K, Zilbering A, Zhu L, and Goldstein BJ. Insulin-stimulated hydrogen peroxide reversibly inhibits protein tyrosine phosphatase 1B in vivo and enhances the early insulin action cascade. J Biol Chem 276: 21938–21942, 2001.
McNicholas CM, Guggino WB, Schwiebert EM, Hebert SC, Giebisch G, and Egan ME. Sensitivity of a renal K+ channel (ROMK2) to the inhibitory sulfonylurea compound glibenclamide is enhanced by coexpression with the ATP-binding cassette transporter cystic fibrosis transmembrane regulator. Proc Natl Acad Sci USA 93: 8083–8088, 1996.
Mohazzab HM and Wolin MS. Sites of superoxide anion production detected by lucigenin in calf pulmonary artery smooth muscle. Am J Physiol Lung Cell Mol Physiol 267: L815–L822, 1994.
Mohazzab KM, Fayngersh RP, Kaminski PM, and Wolin MS. Potential role of NADH oxidoreductase-derived reactive O2 species in calf pulmonary arterial PO2-elicited responses. Am J Physiol Lung Cell Mol Physiol 269: L637–L644, 1995.
Naaray-Fejes-Toth A and Fejes-Toth G. The sgk, an aldosterone-induced gene in mineralocorticoids target cells, regulates the epithelial sodium channel. Kidney Int 57: 1290–1294, 2000.
Nakamura H, Hori T, Sato N, Sugie K, Kawakami T, and Yodoi J. Redox regulation of a Src family protein tyrosine kinase p56Lck in T cells. Oncogene 8: 3133–3139, 1993.
Palmer LG. Potassium secretion and the regulation of distal nephron K channels. Am J Physiol Renal Physiol 277: F821–F825, 1999.
Palmer LG, Antonian L, and Frindt G. Regulation of apical K and Na channels and Na/K pumps in rat cortical collecting tubule by dietary K. J Gen Physiol 104: 693–710, 1994.
Palmer LG, Choe H, and Frindt G. Is the secretory K channel in the rat CCT ROMK Am J Physiol Renal Physiol 273: F404–F410, 1997.
Palmer LG and Frindt G. Regulation of apical K channels in the rat cortical collecting tubule during changes in dietary K intake. Am J Physiol Renal Physiol 277: F805–F812, 1999.
Reed SI. Ratchets and clocks: the cell cycle, ubiquitylation and protein turnover. Nat Rev Mol Cell Biol 4: 855–864, 2003.
Ruknudin A, Schulze DH, Sullivan SK, Lederer WJ, and Welling P. Novel subunit composition of a renal epithelial KATP channel. J Biol Chem 273: 14165–14171, 1998.
Sackin H, Palmer LG, and Krambis M. Potassium-dependent slow inactivation of Kir1.1(ROMK) channels. Biophys J 86: 2145–2155, 2004.
Schnell JD and Hicke L. Non-traditional functions of ubiquitin and ubiquitin-binding proteins. J Biol Chem 278: 35857–35860, 2003.
Simon DB, Karet FE, Rodriguez J, Hamdan JH, DiPietro A, Trachtman H, Sanjad SA, and Lifton RP. Genetic heterogeneity of Bartter's syndrome revealed by mutations in the K channel, ROMK. Nat Genet 14: 152–156, 1996.
Snyder PM, Price MP, McDonald FJ, Adams CM, Volk KA, Zeiher BG, Stokes JB, and Welsh MJ. Mechanism by which Liddle's Syndrome mutations increase activity of a human epithelial Na+ channel. Cell 83: 969–978, 1995.
Staub O, Gautschi I, Ishikawa T, Breitschopf K, Ciechanover A, Schild L, and Rotin D. Regulation of stability and function of the epithelial Na+ channel (ENaC) by ubiquitination. EMBO J 16: 6325–6336, 1997.
Sterling H, Lin DH, Chen YJ, Wei Y, Wang ZJ, Lai J, and Wang WH. PKC expression is regulated by dietary K intake and mediates internalization of SK channels in the CCD. Am J Physiol Renal Physiol 286: F1072–F1078, 2004.
Sterling H, Lin DH, Gu RM, Dong K, Hebert SC, and Wang WH. Inhibition of protein-tyrosine phosphatase stimulates the dynamin-dependent endocytosis of ROMK1. J Biol Chem 277: 4317–4323, 2002.
Sterling H, Lin DH, Wei Y, and Wang WH. Tetanus toxin abolishes exocytosis of ROMK1 induced by inhibition of protein tyrosine kinase. Am J Physiol Renal Physiol 284: F510–F517, 2003.
Stutts MJ, Canessa CM, Olsen JC, Hamrick M, Cohn JA, Rossier BC, and Boucher RC. CFTR as a cAMP-dependent regulator of sodium channels. Science 269: 847–850, 1995.
Sun TJ, Zeng WZ, and Huang CL. Inhibition of ROMK potassium channel by syntaxin 1A. Am J Physiol Renal Physiol 288: F284–F289, 2005.
Wang WH. Two types of K+ channel in thick ascending limb of rat kidney. Am J Physiol Renal Fluid Electrolyte Physiol 267: F599–F605, 1994.
Wang WH and Giebisch G. Dual modulation of renal ATP-sensitive K+ channel by protein kinases A and C. Proc Natl Acad Sci USA 88: 9722–9725, 1991.
Wang WH, Hebert SC, and Giebisch G. Renal K channels: structure and function. Ann Rev Physiol 59: 413–436, 1997.
Wang WH, White S, Geibel J, and Giebisch G. A potassium channel in the apical membrane of the rabbit thick ascending limb of Henle's loop. Am J Physiol Renal Fluid Electrolyte Physiol 258: F244–F253, 1990.
Wei Y, Bloom P, Gu RM, and Wang WH. Protein-tyrosine phosphatase reduces the number of apical small conductance K channels in the rat cortical collecting duct. J Biol Chem 275: 20502–20507, 2000.
Wei Y, Bloom P, Lin DH, Gu RM, and Wang WH. Effect of dietary K intake on the apical small-conductance K channel in the CCD: role of protein tyrosine kinase. Am J Physiol Renal Physiol 281: F206–F212, 2001.
Wilson FH, Kahle KT, Sabath E, Lalioti MD, Rapson AK, Hoover RC, Hebert SC, Gamba G, and Lifton RP. Molecular pathogenesis of inherited hypertension with hyperkalemia: the Na-Cl cotransporter is inhibited by wild type but not mutant WNK4. Proc Natl Acad Sci USA 100: 680–684, 2003.
Woda CB, Bragin A, Kleyman T, and Satlin LM. Flow-dependent K secretion in the cortical collecting duct (CCD) is mediated by a TEA-sensitive channel. Am J Physiol Renal Physiol 280: F786–F793, 2001.
Woda CB, Leite M Jr, Rohatgi R, and Satlin LM. Effects of luminal flow and nucleotides on Ca2+ in rabbit cortical collecting duct. Am J Physiol Renal Physiol 283: F437–F446, 2002.
Wolin MS. Reactive oxygen species and vascular signal transduction mechanisms. Microcirculation 3: 1–17, 1996.
Xu ZC, Yang Y, and Hebert SC. Phosphorylation of the ATP-sensitive, inwardly-rectifying K channel, ROMK, by cyclic AMP-dependent protein kinase. J Biol Chem 271: 9313–9319, 1996.
Yoo D, Flagg TP, Olsen O, Raghuram V, Foskett JK, and Welling PA. Assembly and trafficking of a multiprotein ROMK (Kir 1.1) channel complex by PDZ interactions. J Biol Chem 279: 6863–6873, 2004.
Yoo D, Kim BY, Campo C, Nance L, King A, Maouya D, and Welling PA. Cell surface expression of the ROMK (Kir1.1) channel is regulated by aldosterone-induced kinase, SGK, and protein kinase A. J Biol Chem 278: 23066–23075, 2003.
Yun CC, Palmada M, Embark HM, Fedorenko O, Feng Y, Henke G, Setiawan I, Boehmer C, Weinman EJ, Sandrasagra S, Korbmacher C, Cohen P, Pearce D, and Lang F. The serum and glucorticoid-inducible kinase SGK1 and Na/H exchange regulating factor NHERF2 synergize to stimulate the renal outer medullary K channel ROMK1. J Am Soc Nephrol 13: 2823–2830, 2002.
Zeng WZ, Babich V, Ortega B, Quigley R, White SJ, Welling PA, and Huang CL. Evidence for endocytosis of ROMK potassium channel via clathrin-coated vesicles. Am J Physiol Renal Physiol 283: F630–F639, 2002.
Zeng WZ, Li XJ, Hilgemann DW, and Huang CL. Protein kinase C inhibits ROMK1 channel activity via a phosphatidylinositol 4,5-biphosphate-dependent mechanism. J Biol Chem 278: 16852–16856, 2003.(Wen-Hui Wang)
ABSTRACT
This brief review attempts to provide an overview regarding recent developments in the regulation of ROMK channels. Studies performed in ROMK null mice suggest that ROMK cannot only form hometetramers such as the small-conductance (30-pS) K channels but also construct heterotetramers such as the 70-pS K channel in the thick ascending limb (TAL). The expression of ROMK channels in the plasma membrane is regulated by protein tyrosine kinase (PTK), serum and glucorticoid-induced kinase (SGK), and with-no-lysine-kinase 4. PTK is involved in mediating the effect of low K intake on ROMK channel activity. Increases in superoxide anions induced by low dietary K intake are responsible for the stimulation of PTK expression and tyrosine phosphorylation of ROMK channels. Finally, a recent study indicated that ROMK channels can be monoubiquitinated and monoubiquitination regulates the surface expression of ROMK channels.
THE ROMK CHANNEL PLAYS AN important role in K recycling in the thick ascending limb (TAL) and K secretion in the connecting tubule (CT) and cortical collecting duct (CCD) (16, 19, 34, 43, 45, 61). Although maxi-K channels are also possibly involved in the regulation of K secretion in the CCD (66, 67), it is generally agreed that ROMK channels are mainly responsible for K secretion under normal conditions. The regulation of ROMK channels has been extensively studied, and a recent review has provided extensive coverage regarding the regulatory mechanism of ROMK in the past (16). Thus this review is meant to emphasize the newest developments in the field, which may not have been included in previous reviews. Several studies have identified new amino acids that are involved in the regulation of pH sensitivity of ROMK1 channels in addition to lysine residue 80 (14). Also, it has been shown that changes in extracellular K concentrations can affect the sensitivity of ROMK channels to cell pH (9, 49). Although these topics are important to an understanding of the function of ROMK channels, they are beyond the focus of the present review.
ROMK IS A KEY COMPONENT OF THE APICAL K CHANNELS IN THE TAL
Apical K channels play an important role in K recycling, which is essential for maintaining the normal function of the Na-K-Cl cotransporter in the TAL (16, 61). Patch-clamp experiments demonstrated that two types of K channels, a 30- to 40- and a 70- to 80-pS, are expressed in the apical membrane of the TAL (3, 59, 62). It is well established that the 30-pS K channel is related to ROMK because it has similar biophysical properties and regulatory mechanisms to that in native tubules (61). However, it is not clear whether ROMK is also involved in forming the apical 70-pS K channel. The observation that the loss-of-function mutations of ROMK result in severe salt wasting and metabolic alkalosis (Bartter's disease) (34, 51) indicates strongly that ROMK is an important component of both the 30- and 70-pS K channels in the TAL. Because 70-pS K channels could contribute as much as 60–70% of apical K conductance in the TAL of the rat kidney (36), it is hard to imagine that loss-of-function mutations of ROMK could impair the NaCl transport in the TAL if ROMK is only responsible for the construction of the 30-pS K channels. This puzzle is partially resolved by the patch-clamp experiments performed in the TAL from ROMK null mice in which neither 70- nor 30-pS K channels are detected. In addition, ROMK null mice have demonstrated a typical phenotype of Bartter's syndrome: severe salt wasting, metabolic alkalosis, and hypokalemia (34). This suggests strongly that the 70-pS K channel is a heterotetramer including ROMK and an unidentified subunit. Alternatively, deletion of the ROMK gene could indirectly suppress the expression of the 70-pS K channel. However, the first possibility is further suggested by the observation that high K intake doubled the number of the 70-pS K channels while reducing the number of the 30-pS K channels in the TAL from both wild-type and ROMK (+/–) mice (35). It is possible that high K intake facilitates the formation of the 70-pS K channels by decreasing the formation of the 30-pS K channels. This is achieved by increasing reconstruction between ROMK and other unidentified subunits (Fig. 1) to form the heterotetramer (16). The conductance of the heterotetramer K channel could be different from that of the homotetramer. Relevant to this is the acetylcholine-activated K channel, which is composed of GIRK and cardiac inwardly rectifying K (CIR) channels. Expression of CIR alone in oocytes produces a K channel with single-channel conductance of 15 pS, whereas the coexpression of CIR and GIRK forms an acetylcholine-activated K channel with a single-channel conductance of 37 pS (26). It is possible that the 70-pS K channel may be formed by ROMK and another inwardly rectifying K channel. However, the nature of the unidentified subunit of the 70-pS K channel remains to be determined.
The abundance of ROMK channels in the TAL is regulated by hormones and Na diet. It has been shown that infusion of vasopressin into Brattleboro rats for 7 days increased the apical labeling of ROMK in the TAL (13). The role of vasopressin in the regulation of ROMK channel expression is further indicated by the finding that restriction of water intake can also significantly increase ROMK expression in the TAL. Also, ROMK expression is upregulated in the TAL in response to high Na intake (13). This suggests that ROMK channels and Na-K-Cl cotransporters are possibly coordinately regulated in the TAL by vasopressin and Na intake.
NEW ASPECTS OF THE EFFECTS OF PKC AND CFTR ON ROMK
Regulation by PKC
ROMK1 channels have three putative PKC phosphorylation sites: serine residues (Ser) 4, 181, and 201 (19). In vitro phosphorylation has shown that ROMK1 can be phosphorylated by PKC on Ser4 and Ser201 (29). Moreover, stimulation of PKC has been shown to inhibit ROMK channel activity in native tubules (28, 60). However, the K current in oocytes injected with ROMK1, S4D or ROMK1, S4D/S201D in which the serine residue was mutated to aspartate was either unchanged or increased, suggesting that the PKC-induced inhibition of ROMK1 may not be the result of direct phosphorylation. This possibility was further suggested by the finding that stimulation of PKC with phorbol ester reduced phosphatidylinositol 4,5-bisphosphate (PIP2) content in oocyte membrane and inhibition of PKC abolished the phorbol ester-induced decrease in PIP2 (74). Because PIP2 has been shown to maintain ROMK channels in an open state (20, 33), it is possible that decreases in PIP2 concentration induced by stimulating PKC may contribute to the inhibition of ROMK channel activity. In addition, PKC has been shown to regulate the surface expression of ROMK and endocytosis of ROMK induced by stimulation of PTK (29, 54). We demonstrated that mutation of Ser4 and Ser201 to alanine significantly suppressed the surface expression of ROMK1 in oocytes (29). This suggests that direct phosphorylation of ROMK1 by PKC is important for the surface expression of K channels in oocytes. On the other hand, we also observed that inhibition of PKC blocked the internalization of ROMK1 induced by stimulation of PTK (54). This seeming discrepancy may be the result of different PKC targets. It is possible that the role of PKC in mediating the internalization of ROMK channels may be achieved by targeting factors other than ROMK. Relevant to this hypothesis is the observation that PKC is involved in actin reorganization (5). Thus the complexity of PKC's effects on ROMK may be induced because PKC regulates ROMK channel activity by direct and indirect mechanisms.
Regulation by CFTR
The role of CFTR in the regulation of ROMK channel activity has been demonstrated previously. CFTR has been shown to play an important role in mediating sulfonylurea agent-induced inhibition and ATP sensitivity (38, 48), although it has also been reported that ROMK channels have an intrinsic sensitivity to sulfonylurea agents (24). A recent study suggests that CFTR may play a role in mediating the interaction between ROMK and the epithelial Na channel (ENaC). Previous studies have shown the interaction between ENaC and CFTR such that stimulation of CFTR reduced ENaC activity (57), whereas coexpression of ENaC increased CFTR expression (25). Recently, it also has been reported that CFTR may link the activity of ENaC to ROMK. In the presence of CFTR, expression of ROMK channels was augmented in oocytes injected with ENaC (25). Also, the PDZ domain has been shown to facilitate the interaction between ROMK and CFTR (70). In contrast, coexpression of ENaC had no effect on ROMK expression in the absence of CFTR. This suggests that CFTR synchronizes the activity of ENaC and ROMK. However, whether such interaction takes place in the native tubule is not clear and needs to be examined.
SUPEROXIDE IS INVOLVED IN THE REGULATION OF ROMK CHANNEL ACTIVITY
ROMK1 is exclusively located in the CT and CCD (4) and plays an important role in K secretion. A large body of evidence indicates that the number of ROMK channels in the CCD is varied such that high K intake increases (44, 46), whereas low K intake decreases, ROMK channel numbers in the CCD (2). The response of rats to low K is very sensitive because decreases in K content from 1.1 to 0.7% significantly reduced ROMK channel activity in the CCD to an extent not significantly different from that in rats on a K-deficient diet. This could be the reason the previous study failed to detect a significant difference in ROMK channel activity in the CCD between rats on a K-deficient diet and "normal K diet (0.7%)" (64). Several lines of evidence strongly suggest that the PTK-dependent signal transduction pathway plays a key role in mediating the effect of low K intake on ROMK channel activity. First, low K intake significantly increases the expression Src-family PTK such as c-Src and c-Yes in the kidney (64). Second, the Src family PTK is expressed in principal cells of the CCD and CT (32). Third, low K intake enhances whereas high K intake diminishes the tyrosine phosphorylation of ROMK (30). Finally, inhibition of PTK increases whereas blockade of protein tyrosine phosphatase (PTP) decreases the ROMK channel activity in the CCD (63, 64). Studies with confocal microscopy have further indicated that stimulation of PTK-induced phosphorylation of ROMK1 facilitates the internalization (55, 56). These data suggest that low K intake stimulates the activity of Src family PTK which, in turn, enhances the tyrosine phosphorylation of ROMK channels in the CCD. As a consequence, ROMK channels are internalized and the density of ROMK in the apical membrane decreases.
Although the role of PTK in mediating the effect of low K intake on ROMK channel activity has been established, the upstream signaling which regulates PTK activity is not completely understood. A recent study suggests the possible role of superoxide in the regulation of expression of Src family PTK. Using a chemiluminescence technique (68), it was found that low K intake significantly increased the concentration of superoxide anions in the renal cortex and outer medulla (2) and that the superoxide production induced by low K intake was blunted by tempol, a superoxide dismutase mimetic (27). The role of superoxide in mediating the effect of low K intake on PTK expression was further suggested by observations that treatment of M1 cell, a mouse principal cell line, with hydrogen peroxide significantly increased the expression of c-Src. The effect of hydrogen peroxide on c-Src expression was abolished by either cycloheximide or actinomycin A. This indicates that the stimulatory effect of hydrogen peroxide on c-Src expression is the result of increased transcription and translation. Indeed, Western blot analysis revealed that low K intake increased the phosphorylation of c-Jun on Ser73, an indication of activation of c-Jun. The view that increases in superoxide anions may stimulate transcription of PTK was also suggested by experiments in which treatment of M1 cells with hydrogen peroxide significantly increased the serine phosphorylation of c-Jun. Moreover, tempol treatment completely abolished the effect of low K intake on c-Jun activation and c-Src expression in the kidney of rats fed a K-deficient diet.
The possibility that superoxide is involved in the regulation of ROMK channel activity and renal K excretion is supported by several lines of evidence (2). First, lowering superoxide production in the kidney by tempol decreased the tyrosine phosphorylation of ROMK in the renal cortex and outer medulla and increased the ROMK channel activity in the CCD. Second, metabolic cage studies showed that the renal K excretion in the tempol-treated rats was 10 times higher than those in rats on a K-deficient diet alone. Third, animals receiving tempol developed a more severe hypokalemia than those without tempol treatment. Figure 2 is a model of principal cell illustrating the mechanism by which low K intake inhibits ROMK1 channel activity. Low K intake increases the production of superoxide, which, in turn, stimulates PTK expression. As a consequence, tyrosine phosphorylation of ROMK channels in the CCD increases and the ROMK channel is endocytosed. In addition to stimulation of PTK expression, superoxide and the related product have also been shown to stimulate PTK (6, 42) or inhibit the activity of PTP (10, 37). Thus it is possible that superoxide may be able to acutely regulate ROMK channel activity through modulation of PTK and PTP interaction. Indeed, we have observed that acute application of hydrogen peroxide inhibited ROMK channel activity in the rat CCD and that the effect was blocked by herbimycin A (unpublished observations), indicating that the effect of hydrogen peroxide on ROMK channel activity is due to stimulation of PTK activity. Although the role of superoxide in mediating the effect of low K intake on renal K secretion has been suggested, the mechanism by which low K intake stimulates superoxide production is still not clear. Because the major source of superoxide is from activation of NAD(p)H oxidase (12, 15, 39, 40), it would be interesting to determine the role of NAD(p)H oxidase in the regulation of renal K secretion.
REGULATION OF ROMK CHANNEL TRAFFICKING
A large body of evidence indicates that ROMK channel activity is regulated by endocytosis and exocytosis. ROMK channels have been shown to interact with clathrin via AP-2 protein (73). Recently, it has been demonstrated that syntaxin 1A interacts with the COOH terminus of ROMK and that coexpression of syntaxin 1A inhibits ROMK channel activity in oocytes (58). Also, several new mechanisms including monoubiquitination, serum and glucocorticoid-inducible kinase (SGK), and with-no-lysine-kinase 4 (WNK4) have been discovered in the regulation of ROMK expression in plasma membrane.
Monoubiquitination
Ubiquitination plays an important role in the regulation of protein degradation and recycling by attaching ubiquitin molecules to lysine residues of substrate protein (8, 17, 47). Ubiquitination can further be classified into monoubiquitination by adding only one or two ubiquitin molecules to the substrate protein or polyubiquitination by attaching more than four ubiquitin molecules. The polyubiquitinated protein is subjected to degradation, whereas monoubiquitinated proteins are targeted to internalization and possibly recycling to the cell membrane (11, 18, 50). A large body of evidence indicates that polyubiquitination plays an important role in the regulation of cell signaling and ENaC (52, 53). However, the role of monoubiquitination in the regulation of membrane transporters and ion channels is not clear. Recently, it has been shown that ROMK channel activity could be regulated by monoubiquitination (31). First, immunoprecipitation of ROMK from renal tissue or from cell culture revealed a 50-kDa protein band that was also recognized by an ubiquitin antibody. Second, mutation of lysine residue 22 to arginine (R1K22R) in the NH2 terminus of ROMK1 increased ROMK channel activity. Third, no ubiquitinated ROMK can be detected in oocytes injected with a ROMK1 mutant, R1K22R. Moreover, the biophysical properties of R1K22R such as channel conductance and open probability are the same as that of wt ROMK. Thus an increase in ROMK channel activity must be the result of augmentation of membrane expression of ROMK1 channels. This view is also supported by the observation that a higher biotin-labeled ROMK1, an indication of membrane expressed ROMK1, was observed in cells transfected with ROMK1 mutant than those of ROMK1.
The physiological importance of monoubiquitination in the regulation of ROMK channel is still not clear. Because ROMK channel activity in the CCD decreases in response to stimulation of PTK, it would be interesting to determine whether monoubiquitination is required for the PTK-induced internalization of ROMK1 in the CCD. Figure 3 is a model of a cell illustrating a possible role of ubiquitination in the regulation of ROMK trafficking. Stimulation of PTK has been shown to activate ubiquitin ligase activity (23). It is possible that PTK could enhance the ubiquitination of ROMK and accordingly internalization.
SGK1
Aldosterone has been shown to stimulate glucocorticoid-inducible kinase 1 (SGK1) levels (41), which are possibly involved in mediating the effect of aldosterone on ENaC activity (1, 7). SGK1 was first thought to have no effect on ROMK channel activity (7). As a matter of fact, ROMK was used as a negative control to demonstrate that the stimulatory effect of SGK1 on ENaC was specific. However, three separate studies have later demonstrated that SGK1 can also increase the activity of ROMK in the presence of Na/H exchange-regulating factor (NHERF) (70–72). It is possible that NHERF is served as a scaffold protein required for the stimulatory effect of SGK1 on ROMK. However, the role of NHERF, specifically NHERF2 in mediating the SGK effect, is controversial. Yoo et al. (71) observed that SGK could stimulate ROMK channel without NHERH. Moreover, they observed a modest stimulation of ROMK surface expression by NHERF alone. They suggested that NHERF is required for the effect of SGK on ROMK only when abundance of one of them is low. The mechanism of the SGK1 effect may be through stimulation of the serine phosphorylation of ROMK1. Confocal microscopy and cell surface antibody-binding assay have further shown that stimulation of SGK-induced phosphorylation increases the expression of ROMK1 channels in plasma membrane. The phosphorylation site of ROMK1 induced by SGK is located on Ser44 in the NH2 terminus, which is also a putative PKA phosphorylation site (69) because mutation of Ser44 to alanine abolished the effect of SGK (71). Also, Yoo et al. (71) suggested that effect of SGK on ROMK activity in oocytes depends on the basal level of ROMK phosphorylation such that the stimulatory effect of SGK could be blunted when the basal phosphorylation level of ROMK is high. The role of SGK on K secretion is also demonstrated in SGK null mice in which renal K secretion is impaired (21). However, it is still not known whether the stimulatory effect of SGK on renal K secretion is the result of increased ENaC activity that, in turn, augments the electrochemical gradient for K secretion or due to a direct stimulation of ROMK channel insertion. Confocal microscopy has shown that ROMK expression in the apical membrane of the TAL and CCD is actually increased rather than decreased in SGK null mice. This suggests that the role of SGK in stimulating ROMK insertion may not be essential or can be replaced by other kinases such as cAMP-dependent protein kinase A. A possible role of SGK is to regulate renal K secretion in response to a daily dietary K intake. It has been shown that a high dietary K intake for 12 h stimulates SGK expression in the kidney (71). Interestingly, it has been shown that ROMK channel activity significantly increased in the CCD from rats on a high-K diet for only 6 h (46). Thus SGK may regulate ROMK channel activity in the CT and CCD in response to a daily variation in dietary K intake.
WNK4
WNK4 is a serine-threonine kinase and has been identified in both distal convoluted tubule (DCT) and the CCD. Wild-type WNK4 has been shown to suppress the expression of the Na-Cl cotransporter in the plasma membrane of the DCT and, accordingly, to inhibit Na absorption (65). The gene product encoding inactivated WNK4 causes pseudohypoaldosteronism type II, a disease with characteristics of hypertension and low aldosterone levels. WNK4 is also expressed in the CCD, and expression of WNK4 in oocytes decreased the expression of ROMK1 (22). However, the inhibitory effect of WNK4 on ROMK1 does not require the kinase activity because coexpression of inactivated WNK4 also resulted in an inhibition of the expression of ROMK1 in Xenopus laevis oocytes. The physiological role of WNK4 may be that it serves as a switch between Na transport and K secretion. The WNK4-induced inhibition of Na transport in the DCT is expected to increase Na delivery to the CCD and enhance Na absorption, which leads to augmentation of the driving force for K secretion. However, because WNK4 inhibits ROMK channel activity, increases in electrochemical gradient for K secretion would have a diminished effect on renal K secretion. Mutation of WNK4 would release the inhibition of the Na-Cl cotransporter and lead to stimulation of Na transport in the DCT. Moreover, inactivated WNK4 further decreases the surface expression of ROMK, reduces K secretion in the CCD, and causes hyperkalemia. However, it is possible that decreases in Na delivery to the CCD may also be responsible for the diminished K secretion in patients with pseudohypoaldosteronism type II.
SUMMARY
Figure 3 is a model of the principal cell, illustrating the possible mechanisms by which ROMK channel expression in plasma membrane is regulated by WNK4, SGK, and ubiquitin. Stimulation of PTK decreases, whereas activation of SGK1 increases, the expression of ROMK. Also, increased WNK4 expression should diminish ROMK channel activity.
GRANTS
This work is supported by National Institutes of Health Grants DK-54983, DK-47402, and HL-34300.
ACKNOWLEDGMENTS
The author thanks Melody Steinberg for editorial assistance.
FOOTNOTES
REFERENCES
Alvarez de la Rosa D, Zhang P, Naaray-Fejes-Toth A., Fejes-Toth G, and Canessa CM. The serum and glucocorticoid kinase sgk increases the abundance of epithelial sodium channels in the plasma membrane of Xenopus oocytes. J Biol Chem 274: 37834–37839, 1999.
Babilonia E, Wei Y, Sterling H, Kaminski P, and Wolin MS. Superoxide anions are involved in mediating the effect of low K intake on c-Src expression and renal K secretion in the cortical collecting duct. J Biol Chem 280: 10790–10796, 2005.
Bleich M, Schlatter E, and Greger R. The luminal K+ channel of the thick ascending limb of Henle's loop. Pflügers Arch 415: 449–460, 1990.
Boim MA, Ho K, Schuck ME, Bienkowski MJ, Block JH, Slightom JL, Yang Y, Brenner BM, and Hebert SC. The ROMK inwardly rectifying ATP-sensitive K channel. II. Cloning and intra-renal distribution of alternatively spliced forms. Am J Physiol Renal Fluid Electrolyte Physiol 268: F1132–F1140, 1995.
Brandt D, Gimona M, Hillmann M, Haller H, and Mischak H. Protein kinase C induces actin reorganization via a Src- and Rho-dependent pathway. J Biol Chem 277: 20903–20910, 2002.
Brumell JH, Burkhardt AL, Bolen JB, and Grinstein S. Endogenous reactive oxygen intermediates activate tyrosine kinase in human neutrophils. J Biol Chem 271: 1455–1461, 1996.
Chen SY, Bhargava A, Mastroberardino L, Meijer OC, Wang J, Buse P, Firestone GL, Verrey F, and Pearce D. Epithelial sodium channel regulation by aldosterone-induced protein SGK. Proc Natl Acad Sci USA 96: 2514–2519, 1999.
Ciechanover A. The ubiquitin-proteasome proteolytic pathway. Cell 79: 13–21, 1994.
Dahlmann A, Li M, Gao ZH, McGarrigle D, Sackin H, and Palmer LG. Regulation of Kir channels by intracellular pH and extracellular K: mechanism of coupling. J Gen Physiol 123: 441–454, 2004.
Denu JM and Tanner KG. Specific and reversible inactivation of protein tyrosine phosphatases by hydrogen peroxide: evidence for a sulfenic acid intermediate and implications for redox regulation. Biochemistry 37: 5633–5642, 1998.
DiFiore PPD, Polo S, and Hofmann K. When ubiquitin meets ubiquitin receptors: a signalling connection. Nat Rev Mol Cell Biol 4: 491–497, 2003.
Droge W. Free radicals in the physiological control of cell function. Physiol Rev 82: 47–95, 2002.
Ecelbarger CA, Kim CH, Knepper MA, Liu J, Tate M, Welling PA, and Wade JB. Regulation of potassium channel Kir 1.1 (ROMK) abundance in the thick ascending limb of Henle's loop. J Am Soc Nephrol 12: 10–18, 2001.
Fakler B, Schultz JH, Yang J, Schulte U, Braendle U, Zenner HP, Jan LY, and Ruppersberg J. Identification of a titratable lysine residue that determines sensitivity of kidney potassium channels (ROMK) to intracellular pH. EMBO J 15: 4093–4099, 1996.
Geiszt M and Leto T. The nox family of NAD(p)H oxidase: host defense and beyond. J Bio Chem 279: 51715–51718, 2004.
Hebert SC, Desir GV, Giebisch G, and Wang WH. Molecular diversity and regulation of renal potassium channels. Physiol Rev 85: 319–371, 2005.
Hershko A and Ciechanover A. The ubiquitin system for protein degradation. Annu Rev Biochem 61: 761–807, 1992.
Hicke L and Dunn R. Regulation of membrane protein transport by ubiquitin and ubiquitin-binding proteins. Annu Rev Cell Dev Biol 19: 172, 2003.
Ho K, Nichols CG, Lederer WJ, Lytton J, Vassilev PM, Kanazirska MV, and Hebert SC. Cloning and expression of an inwardly rectifying ATP-regulated potassium channel. Nature 362: 31–38, 1993.
Huang CL, Feng S, and Hilgemann DW. Direct activation of inward rectifier potassium channels by PIP2 and its stabilization by G. Nature 391: 803–806, 1998.
Huang DY, Wulff P, Volkl H, Loffing J, Richter K, Kuhl D, Lang F, and Vallon V. Impaired regulation of renal K elimination in the sgk1-knockout mouse. J Am Soc Nephrol 15: 885–891, 2004.
Kahle KT, Wilson FH, Leng Q, Lalioti MD, Connell AD, Dong K, Rapson AK, MacGregor GG, Giebisch G, Hebert SC, and Lifton RP. WNK4 regulates the balance between renal NaCl reabsorption and K secretion. Nat Genet 35: 372–376, 2003.
Kassenbrock CK and Anderson SM. Regulation of ubiquitin protein ligase activity in c-Cbl by phosphorylation-induced conformational change and constitutive activation by tyrosine to glutamate point mutations. J Biol Chem 279: 28017–28027, 2004.
Konstas AA, Dabrowski M, Korbmacher C, and Tucker SJ. Intrinsic sensitivity of Kir1.1 (ROMK) to glibenclamide in the absence of sur2b. J Biol Chem 277: 21346–21351, 2002.
Konstas AA, Koch JP, Tucker SJ, and Korbmacher C. Cystic fibrosis transmembrane conductance regulator-dependent up-regulation of Kir 1.1 (ROMK) renal K channels by the epithelial sodium channel. J Biol Chem 277: 25377–25384, 2002.
Krapivinsky G, Gordon EA, Wickman K, Velimirovic B, Krapivinsky L, and Clapham DE. The G-protein-gated atrial K+ channel I KACh is a heteromultimer of two inwardly rectifying K+-channel proteins. Nature 374: 135–141, 1995.
Krishna MC, Grahame DA, Samuni A, Mitchell JB, and Russo A. Oxoammonium cation intermediate in the nitroxide-catalyzed dismutation of superoxide. Proc Natl Acad Sci USA 89: 5537–5541, 1992.
Kubokawa M, Wang WH, McNicholas CM, and Giebisch G. Role of Ca2+/CaMK II in Ca2+-induced K+ channel inhibition in rat CCD principal cell. Am J Physiol Renal Fluid Electrolyte Physiol 268: F211–F219, 1995.
Lin DH, Sterling H, Lerea KM, Giebisch G, and Wang WH. Protein kinase C (PKC)-induced phosphorylation of ROMK1 is essential for the surface expression of ROMK1 channels. J Biol Chem 277: 44332–44338, 2002.
Lin DH, Sterling H, Lerea KM, Welling P, Jin L, Giebisch G, and Wang WH. K depletion increases the protein tyrosine-mediated phosphorylation of ROMK. Am J Physiol Renal Physiol 283: F671–F677, 2002.
Lin DH, Sterling H, Wang Z, Babilonia E, Yang B, Dong K, Hebert SC, Giebisch G, and Wang WH. ROMK1 channel activity is regulated by monoubiquitination. Proc Natl Acad Sci USA 102: 4306–4311, 2005.
Lin DH, Sterling H, Yang B, Hebert SC, Giebisch G, and Wang WH. Protein tyrosine kinase is expressed and regulates ROMK1 location in the cortical collecting duct. Am J Physiol Renal Physiol 286: F881–F892, 2004.
Liou HH, Zhou SS, and Huang CL. Regulation of ROMK1 channel by protein kinase A via a phosphatidylinositol 4,5-bisphosphate-dependent mechanism. Proc Natl Acad Sci USA 96: 5820–5825, 1999.
Lu M, Wang T, Yan Q, Yang X, Dong K, Knepper MA, Wang WH, Giebisch G, Shull GE, and Hebert SC. Absence of small conductance K channel (SK) activity in apical membrane of thick ascending limb and cortical collecting duct in ROMK (Bartter's) knockout mice. J Biol Chem 277: 37881–37887, 2002.
Lu M, Wang T, Yan QS, Wang WH, Giebisch G, and Hebert SC. ROMK is required for expression of the 70 pS K channel in the thick ascending limb. Am J Physiol Renal Physiol 286: F490–F495, 2004.
Lu M, Zhu Y, Balazy M, Falck JR, and Wang WH. Effect of angiotensin II on the apical K channels in the thick ascending limb of the rat kidney. J Gen Physiol 108: 537–547, 1996.
Mahadev K, Zilbering A, Zhu L, and Goldstein BJ. Insulin-stimulated hydrogen peroxide reversibly inhibits protein tyrosine phosphatase 1B in vivo and enhances the early insulin action cascade. J Biol Chem 276: 21938–21942, 2001.
McNicholas CM, Guggino WB, Schwiebert EM, Hebert SC, Giebisch G, and Egan ME. Sensitivity of a renal K+ channel (ROMK2) to the inhibitory sulfonylurea compound glibenclamide is enhanced by coexpression with the ATP-binding cassette transporter cystic fibrosis transmembrane regulator. Proc Natl Acad Sci USA 93: 8083–8088, 1996.
Mohazzab HM and Wolin MS. Sites of superoxide anion production detected by lucigenin in calf pulmonary artery smooth muscle. Am J Physiol Lung Cell Mol Physiol 267: L815–L822, 1994.
Mohazzab KM, Fayngersh RP, Kaminski PM, and Wolin MS. Potential role of NADH oxidoreductase-derived reactive O2 species in calf pulmonary arterial PO2-elicited responses. Am J Physiol Lung Cell Mol Physiol 269: L637–L644, 1995.
Naaray-Fejes-Toth A and Fejes-Toth G. The sgk, an aldosterone-induced gene in mineralocorticoids target cells, regulates the epithelial sodium channel. Kidney Int 57: 1290–1294, 2000.
Nakamura H, Hori T, Sato N, Sugie K, Kawakami T, and Yodoi J. Redox regulation of a Src family protein tyrosine kinase p56Lck in T cells. Oncogene 8: 3133–3139, 1993.
Palmer LG. Potassium secretion and the regulation of distal nephron K channels. Am J Physiol Renal Physiol 277: F821–F825, 1999.
Palmer LG, Antonian L, and Frindt G. Regulation of apical K and Na channels and Na/K pumps in rat cortical collecting tubule by dietary K. J Gen Physiol 104: 693–710, 1994.
Palmer LG, Choe H, and Frindt G. Is the secretory K channel in the rat CCT ROMK Am J Physiol Renal Physiol 273: F404–F410, 1997.
Palmer LG and Frindt G. Regulation of apical K channels in the rat cortical collecting tubule during changes in dietary K intake. Am J Physiol Renal Physiol 277: F805–F812, 1999.
Reed SI. Ratchets and clocks: the cell cycle, ubiquitylation and protein turnover. Nat Rev Mol Cell Biol 4: 855–864, 2003.
Ruknudin A, Schulze DH, Sullivan SK, Lederer WJ, and Welling P. Novel subunit composition of a renal epithelial KATP channel. J Biol Chem 273: 14165–14171, 1998.
Sackin H, Palmer LG, and Krambis M. Potassium-dependent slow inactivation of Kir1.1(ROMK) channels. Biophys J 86: 2145–2155, 2004.
Schnell JD and Hicke L. Non-traditional functions of ubiquitin and ubiquitin-binding proteins. J Biol Chem 278: 35857–35860, 2003.
Simon DB, Karet FE, Rodriguez J, Hamdan JH, DiPietro A, Trachtman H, Sanjad SA, and Lifton RP. Genetic heterogeneity of Bartter's syndrome revealed by mutations in the K channel, ROMK. Nat Genet 14: 152–156, 1996.
Snyder PM, Price MP, McDonald FJ, Adams CM, Volk KA, Zeiher BG, Stokes JB, and Welsh MJ. Mechanism by which Liddle's Syndrome mutations increase activity of a human epithelial Na+ channel. Cell 83: 969–978, 1995.
Staub O, Gautschi I, Ishikawa T, Breitschopf K, Ciechanover A, Schild L, and Rotin D. Regulation of stability and function of the epithelial Na+ channel (ENaC) by ubiquitination. EMBO J 16: 6325–6336, 1997.
Sterling H, Lin DH, Chen YJ, Wei Y, Wang ZJ, Lai J, and Wang WH. PKC expression is regulated by dietary K intake and mediates internalization of SK channels in the CCD. Am J Physiol Renal Physiol 286: F1072–F1078, 2004.
Sterling H, Lin DH, Gu RM, Dong K, Hebert SC, and Wang WH. Inhibition of protein-tyrosine phosphatase stimulates the dynamin-dependent endocytosis of ROMK1. J Biol Chem 277: 4317–4323, 2002.
Sterling H, Lin DH, Wei Y, and Wang WH. Tetanus toxin abolishes exocytosis of ROMK1 induced by inhibition of protein tyrosine kinase. Am J Physiol Renal Physiol 284: F510–F517, 2003.
Stutts MJ, Canessa CM, Olsen JC, Hamrick M, Cohn JA, Rossier BC, and Boucher RC. CFTR as a cAMP-dependent regulator of sodium channels. Science 269: 847–850, 1995.
Sun TJ, Zeng WZ, and Huang CL. Inhibition of ROMK potassium channel by syntaxin 1A. Am J Physiol Renal Physiol 288: F284–F289, 2005.
Wang WH. Two types of K+ channel in thick ascending limb of rat kidney. Am J Physiol Renal Fluid Electrolyte Physiol 267: F599–F605, 1994.
Wang WH and Giebisch G. Dual modulation of renal ATP-sensitive K+ channel by protein kinases A and C. Proc Natl Acad Sci USA 88: 9722–9725, 1991.
Wang WH, Hebert SC, and Giebisch G. Renal K channels: structure and function. Ann Rev Physiol 59: 413–436, 1997.
Wang WH, White S, Geibel J, and Giebisch G. A potassium channel in the apical membrane of the rabbit thick ascending limb of Henle's loop. Am J Physiol Renal Fluid Electrolyte Physiol 258: F244–F253, 1990.
Wei Y, Bloom P, Gu RM, and Wang WH. Protein-tyrosine phosphatase reduces the number of apical small conductance K channels in the rat cortical collecting duct. J Biol Chem 275: 20502–20507, 2000.
Wei Y, Bloom P, Lin DH, Gu RM, and Wang WH. Effect of dietary K intake on the apical small-conductance K channel in the CCD: role of protein tyrosine kinase. Am J Physiol Renal Physiol 281: F206–F212, 2001.
Wilson FH, Kahle KT, Sabath E, Lalioti MD, Rapson AK, Hoover RC, Hebert SC, Gamba G, and Lifton RP. Molecular pathogenesis of inherited hypertension with hyperkalemia: the Na-Cl cotransporter is inhibited by wild type but not mutant WNK4. Proc Natl Acad Sci USA 100: 680–684, 2003.
Woda CB, Bragin A, Kleyman T, and Satlin LM. Flow-dependent K secretion in the cortical collecting duct (CCD) is mediated by a TEA-sensitive channel. Am J Physiol Renal Physiol 280: F786–F793, 2001.
Woda CB, Leite M Jr, Rohatgi R, and Satlin LM. Effects of luminal flow and nucleotides on Ca2+ in rabbit cortical collecting duct. Am J Physiol Renal Physiol 283: F437–F446, 2002.
Wolin MS. Reactive oxygen species and vascular signal transduction mechanisms. Microcirculation 3: 1–17, 1996.
Xu ZC, Yang Y, and Hebert SC. Phosphorylation of the ATP-sensitive, inwardly-rectifying K channel, ROMK, by cyclic AMP-dependent protein kinase. J Biol Chem 271: 9313–9319, 1996.
Yoo D, Flagg TP, Olsen O, Raghuram V, Foskett JK, and Welling PA. Assembly and trafficking of a multiprotein ROMK (Kir 1.1) channel complex by PDZ interactions. J Biol Chem 279: 6863–6873, 2004.
Yoo D, Kim BY, Campo C, Nance L, King A, Maouya D, and Welling PA. Cell surface expression of the ROMK (Kir1.1) channel is regulated by aldosterone-induced kinase, SGK, and protein kinase A. J Biol Chem 278: 23066–23075, 2003.
Yun CC, Palmada M, Embark HM, Fedorenko O, Feng Y, Henke G, Setiawan I, Boehmer C, Weinman EJ, Sandrasagra S, Korbmacher C, Cohen P, Pearce D, and Lang F. The serum and glucorticoid-inducible kinase SGK1 and Na/H exchange regulating factor NHERF2 synergize to stimulate the renal outer medullary K channel ROMK1. J Am Soc Nephrol 13: 2823–2830, 2002.
Zeng WZ, Babich V, Ortega B, Quigley R, White SJ, Welling PA, and Huang CL. Evidence for endocytosis of ROMK potassium channel via clathrin-coated vesicles. Am J Physiol Renal Physiol 283: F630–F639, 2002.
Zeng WZ, Li XJ, Hilgemann DW, and Huang CL. Protein kinase C inhibits ROMK1 channel activity via a phosphatidylinositol 4,5-biphosphate-dependent mechanism. J Biol Chem 278: 16852–16856, 2003.(Wen-Hui Wang)