Correspondence to: Peter M. Snyder, 371EMRB, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, IA 52242. E-mail:psnyder{at}blue.weeg.uiowa.edu.
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The epithelial Na+ channel (ENaC) is comprised of three homologous subunits (, ß, and
). The channel forms the pathway for Na+ absorption in the kidney, and mutations cause disorders of Na+ homeostasis. However, little is known about the mechanisms that control the gating of ENaC. We investigated the gating mechanism by introducing bulky side chains at a position adjacent to the extracellular end of the second membrane spanning segment (549, 520, and 529 in
, ß, and
ENaC, respectively). Equivalent "DEG" mutations in related DEG/ENaC channels in Caenorhabditis elegans cause swelling neurodegeneration, presumably by increasing channel activity. We found that the Na+ current was increased by mutagenesis or chemical modification of this residue and adjacent residues in
, ß, and
ENaC. This resulted from a change in the gating of ENaC; modification of a cysteine at position 520 in ßENaC increased the open state probability from 0.12 to 0.96. Accessibility to this side chain from the extracellular side was state-dependent; modification occurred only when the channel was in the open conformation. Single-channel conductance decreased when the side chain contained a positive, but not a negative charge. However, alterations in the side chain did not alter the selectivity of ENaC. This is consistent with a location for the DEG residue in the outer vestibule. The results suggest that channel gating involves a conformational change in the outer vestibule of ENaC. Disruption of this mechanism could be important clinically since one of the mutations that increased Na+ current (
N530K) was identified in a patient with renal disease.
Key Words: hypertension, amiloride, sodium channel, epithelia, degenerin
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The epithelial Na+ channel (ENaC)1 is expressed at the apical membrane of epithelia, where it functions in Na+ absorption (, ß, and
ENaC subunits (
Little is known about the mechanisms that control the gating of ENaC. In contrast to voltage- and ligand-gated ion channels, ENaC conducts current in the absence of an identifiable stimulus ( and ß subunits (without
) had a very high Po, whereas channels derived from coexpression of
and
existed in either a high or low Po state (
To investigate the gating mechanisms of ENaC, we examined the effect of mutations in amino acids near the extracellular end of the second membrane-spanning segment. Several findings suggest that this domain might be involved in channel gating. First, in related Caenorhabditis elegans (C. elegans) channels (MEC-4, MEC-10, and DEG-1), dominant gain-of-function mutations in this domain cause neuronal swelling and degeneration, presumably by increasing channel activity (ENaC increased Na+ current (
|
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
DNA Constructs
cDNA constructs for human , ß, and
ENaC in pMT3, pcDNA3 (
Expression and Whole-Cell Electrophysiology
For measurement of whole-cell current, cDNAs encoding , ß, and
ENaC (0.2 ng each; in pMT3 or pcDNA3) were injected into the nucleus of Xenopus oocytes. After incubation in modified Barth's solution at 18°C for 1624 h, we measured whole-cell Na+ currents by two-electrode voltage clamp with the cells bathed in 116 mM NaCl, 2 mM KCl, 0.4 mM CaCl2, 1 mM MgCl2, and 5 mM Hepes, pH 7.4, with NaOH. Amiloride-sensitive current was determined at -60 mV by adding a maximal dose (100 µM) to the bathing solution. Currentvoltage relationships were obtained by stepping from -60 mV to potentials between -120 and +40 mV (20-mV steps) for 300 ms. Permeability ratios were calculated from changes in reversal potential with Na+, Li+, and K+ as the predominant cation in the extracellular bathing solution (
The response to methanethiosulfonate (MTS) compounds was determined by addition to the bathing solution of 1 mM MTSET ([2-(trimethylammonium)ethyl]methanethiosulfonate bromide), 10 mM MTSES (sodium (2-sulfonatoethyl)MTS), or 1 mM MTSEA-biotincap (N-biotinylcaproylaminoethyl MTS; Toronto Research Chemicals). These compounds have no significant effect on wild-type ENaC currents (
Single-Channel Currents
, ß (wild-type or ßS520C), and
ENaC were expressed in Xenopus oocytes by cytoplasmic injection of cRNA (2 ng each). 13 d after injection, single-channel currents were recorded from devitellinized oocytes by patch-clamp (cell attached configuration). The pipet solution contained 150 mM LiCl, 1 mM CaCl2, 1 mM MgCl2, 5 mM Hepes, pH 7.4, with LiOH. The bath solution contained 150 mM LiCl, 5 mM EDTA, and 5 mM Hepes, pH 7.4, with LiOH. Currents were amplified using an Axopatch 200B amplifier (Axon Instruments) and acquired at 2 kHz using Pulse software (version 8.09; HEKA). Currents were digitally filtered at 100 Hz and analyzed using TAC 3.0 (Bruxton Corporation). Slope conductance was determined between -100 and -40 mV (the conductance was identical in recordings filtered at 100 or 1,000 Hz). Open state probability (Po) was determined at -100 mV in patches containing one to three channels. The majority of patches contained a single channel. Mean open and closed times were determined from patches containing single channels.
To selectively modify channels in the patch, 1 mM MTSET or 10 mM MTSES were included in the patch pipet. In some experiments, we used a lower concentration of MTSET (10 µM), and filled the tip of the pipet with solution lacking MTSET, to allow us to record currents before and after modification of the channel.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Bulky DEG Residue Side Chains in ßENaC Increase Na+ Current
To test the hypothesis that mutations of the DEG residue in ENaC increase Na+ current, we mutated Ser520 in the ß subunit (Fig 1) to amino acids with larger and/or charged side chains. Expression of ßS520K (with wild-type and
ENaC) in Xenopus oocytes generated 3.2-fold more amiloride-sensitive Na+ current than wild-type ENaC (Fig 2A and Fig B). This mutation increased the size of the side chain at the DEG position, and added a positive charge. Mutation of Ser520 to an amino acid with a large neutral side chain (ßS520V) or a negatively charged side chain (ßS520E) also increased Na+ current (Fig 2 B). In contrast, a more conservative mutation that did not change the size or charge of the side chain (ßS520C) did not increase Na+ current (Fig 2 B).
|
Because the ßS520C mutation did not alter Na+ current, this allowed us to acutely change the composition of the DEG residue side chain by covalent modification of the cysteine. MTSET attaches an (S)-ethyl trimethyl ammonium group to the cysteine, increasing the size and adding a positive charge to the side chain. When we expressed ßS520C (with wild-type and
ENaC), extracellular MTSET rapidly increased Na+ current 4.3-fold (Fig 3A and Fig B). Amiloride completely blocked the stimulated current, indicating that it resulted from the stimulation of ENaC. Also consistent with this interpretation, MTSET had no significant effect on uninjected oocytes (not shown) or oocytes expressing wild-type ENaC (
|
To determine whether the size or the charge of the side chain (or both) was required for MTSET-induced stimulation of ENaC, we tested the effect of different MTS compounds. MTSES is similar in size to MTSET, but it carries a negative charge. Modification of ßS520C with MTSES increased Na+ current to the same extent as MTSET (Fig 3 B). A large neutral compound (MTSEA-biotincap) also increased Na+ current (Fig 3 B). These results, together with our finding that large neutral, positive, and negative amino acids increased Na+ current (Fig 2 B), suggest that the size of the DEG residue side chain, rather than its charge, was responsible for the stimulation of ENaC.
DEG Mutations in and
ENaC
The three ENaC subunits share significant sequence similarity, including the serine at the DEG position in all three subunits (Fig 1). Therefore, we tested the hypothesis that an increase in the size of the side chain at the DEG position in and
ENaC would increase Na+ current as it did in ßENaC. When we placed a cysteine at the DEG position in
(
S549C) or
ENaC (
S529C) (coexpressed with the other two wild-type subunits), Na+ currents were identical to wild-type ENaC (Fig 4 C). Modification of the cysteine introduced in
S549C with MTSET increased Na+ current (Fig 4 A), although to a lesser extent than modification of ßS520C (Fig 4 B). In contrast, modification of
S529C had minimal effect on Na+ current (Fig 4A and Fig B;
ENaC was not accessible to modification, or modification did not alter the current. In addition, mutation of
S529 to either valine or lysine also failed to increase the current (Fig 4 C), suggesting that, in
ENaC, large side chains at the DEG position do not alter channel function. Conversely, Na+ current decreased when we mutated the DEG residue (Ser549) to valine or lysine in
ENaC (Fig 4 C). This contrasts with the increase in current when
S549C was modified by MTSET (Fig 4 A).
|
We asked whether bulky side chains at positions surrounding the DEG residue would also stimulate the channel. When a cysteine was introduced at position 521 in ßENaC, MTSET increased Na+ current (Fig 4 B). Stimulation was identical when cysteines were modified at the equivalent position in or
ENaC (positions 550 and 530, respectively; Fig 4 B). In contrast, modification of a cysteine introduced at the other neighboring position in
and ßENaC (positions 548 and 519, respectively) did not increase Na+ current. However, in
ENaC, modification of this cysteine (position 528) produced a large increase in Na+ current (Fig 4 B). Thus, stimulation was not specific to the DEG position, but resulted from introduction of a bulky side chain at neighboring positions as well. The data suggest that the DEG region has a similar function in all three ENaC subunits, although there are differences in the function of the specific residues.
A sequence variation in this DEG domain was identified in a patient with diabetic nephropathy, changing Asn530 in ENaC to lysine (
N530K;
N530K (with wild-type
and ßENaC) increased Na+ current twofold compared with wild-type ENaC (Fig 4 D). Thus, a mutation in the DEG domain might alter the function of ENaC in humans.
Effect of Cysteine Modification on Selectivity
ENaC is highly selective for Na+ over K+, and is slightly more permeable to Li+ than Na+ (S540C and
S542C; Fig 1, shaded box) changed the selectivity of ENaC (Fig 5B and Fig C), similar to previous reports (
|
MTSET Locks ßS520C in a High Po State
The presence of a bulky side chain at the DEG position could stimulate ENaC by increasing either the single-channel conductance or the Po. To examine these mechanisms, we measured single-channel currents in cell-attached patches of cells expressing ßS520C with wild-type and
ENaC. To modify the DEG cysteine, MTSET was included in the patch pipet in an independent group of patches. Fig 6 A shows representative currents from patches containing single channels. Without MTSET, ßS520C displayed kinetics similar to previous reports for wild-type ENaC (Fig 6 A, top; (
|
In the absence of MTSET, ßS520C had a single-channel conductance (8.4 pS for Li+) similar to wild-type human ENaC (Fig 6 C;
Modification of ßS520C Is State Dependent
To investigate the conformational changes associated with the gating of ENaC, we tested the hypothesis that ßS520C was modified selectively in either the open or closed conformation. Modification of the channel by MTSET changes the gating and single-channel conductance, converting ENaC from a low Po, large single-channel conductance (OL) state to a high Po, small single-channel conductance (OS) state. If ßS520C was modified in the closed conformation, we predict that the next channel opening would be to the OS/high Po state. This concept is illustrated in Fig 7 A (top). In contrast, if modification occurred when the channel was open, we predict that the channel would first open into the OL state, followed by a decrease in current to the OS/high Po state at the time of modification (Fig 7 A, bottom). In the protocol used in Fig 6, the rate of modification was too fast to allow us to record channel activity at the time of modification. To delay modification, we used a lower concentration of MTSET (10 µM) in the patch pipet, and filled the tip of the pipet with solution lacking MTSET. Using this approach, we were able to observe the transition from the OL/low Po to the OS/high Po state in 10 experiments. An example is shown in Fig 7 B. The patch contained a single channel that opened only to the OL state during the first 4.6 min of recording (the last 8.5 s are shown, Before Modification). The Po during this time was very low (0.01). The channel then converted (Fig 7 B, Modification) to the OS/high Po state for the remainder of recording. After modification, the channel had brief infrequent closures (Fig 7 B, inset c) with a Po close to 1.0, and there was a significant decrease in the single-channel current amplitude (Fig 7 D), which is consistent with conversion from the OL to the OS state. In Fig 7 B, inset b shows an expanded time scale to focus on this conversion between states. The channel first opened into the OL state, followed by a decrease in current to the OS state (indicated by the arrowhead). This sequence is consistent with modification of the channel in the open conformation; it was observed in 10/10 experiments (Fig 7 A). A second example is shown in Fig 7 C. The three sweeps were taken from the same channel before, during, and after modification with MTSET. This record contains longer closures from the OS state. We did not observe channels open directly into the OS state (Fig 7 A). Thus, the data suggest that modification of the DEG residue is state-dependent, occurring selectively in the open conformation.
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Our data indicate that the DEG residue, and adjacent residues, are involved in the gating of ENaC. Large side chains at this position disrupt a conformational change in the outer vestibule, locking the channel in a high Po state.
Several findings suggest that the DEG residue influences the conduction pathway of ENaC. First, a cysteine introduced at this position was accessible to modification with water-soluble thiol-reactive compounds added to the extracellular bathing solution. Second, modification of ßS520C with MTSET decreased the single-channel conductance. Third, a positive charge was required; the negatively charged MTSES did not decrease single-channel conductance. This electrostatic effect suggests that the DEG residue is located within the conduction pathway, and is reminiscent of the effect of charged residues in the outer vestibule on the conductance of the nicotinic acetylcholine receptor (ENaC, residues in the DEG domain lie external to the site in the pore of amiloride block (
The presence of a bulky side chain at the DEG position produced a dramatic change in the gating of ENaC, converting the channel from a low Po state to a state in which the channel was almost always open. This resulted from a destabilization of the closed state and stabilization of the open state, as reflected by the large decrease in closed time and increase in open time, respectively. The requirement for a bulky side chain suggests that this may be a steric effect; perhaps the bulky side chain interferes with the conformational change required for the channel to close. Such a mechanism was previously proposed to explain the swelling neurodegeneration produced by equivalent mutations in C. elegans ( and
ENaC increased Na+ current by increasing Po, similar to the modification of ßS520C. However, we cannot exclude an increase in the single-channel conductance of these mutant channels.
We used two strategies to alter the side chain at the DEG position: mutagenesis to amino acids with large and/or charged side chains, and modification of an introduced cysteine. Both strategies produced equivalent results, with one exception; modification of S549C increased Na+ current, whereas mutation of this residue to valine or lysine decreased current. The reason for this difference is unclear. It is possible that large side chains at the DEG position in
ENaC disrupted the processing of ENaC to the cell surface, resulting in decreased Na+ current. This underscores a significant advantage of the cysteine modification strategy, which allowed us to determine the functional effect of an acute change in the size and/or charge of the side chain. A second possible mechanism involves the number of altered
S549 side chains in the channel complex. ENaC contains either two or three
subunits (
subunits were modified by MTSET; modification of only one cysteine might be sufficient to increase current. In contrast, when we mutated
S549 to valine or lysine, each
subunit contained a large side chain. Perhaps one large
side chain increases current, but current is decreased when all of the
subunits contain large side chains. Finally, the modified cysteine is not identical to lysine or valine; differences in the structure of the side chain could also explain the data. Future work will be required to distinguish between these potential mechanisms. The increase in whole-cell Na+ current with modification of ßS520C by MTSET (3.34.2 fold; Fig 3 B and 4 B) was less than predicted (5.4-fold increase) from the increase in Po (8-fold) and decrease in single-channel current (0.67-fold). It is possible that some channels or DEG cysteines may not have been modified by MTSET.
We found that a cysteine introduced at the DEG position was modified only when the channel was in the open conformation. This state-dependent modification suggests that channel gating results from a conformational change that alters the accessibility of the DEG residue to the extracellular bathing solution. Two potential models could explain these results. First, channel gating might result from the opening and closing of a gate in the extracellular domain of ENaC (Fig 8, top). If the gate was external to the DEG residue, channel closure would block access to this residue. Steric hindrance by the bulky side chain at this position might make it unfavorable for the gate to close. In this model, channel gating does not change the position of the DEG residue in relation to the pore of ENaC. This contrasts with a second potential model in which the DEG residue changes position during the gating conformational change (Fig 8, bottom). When the channel is open, the DEG residue side chain lines the vestibule where it is accessible to modification. Channel closure moves the DEG residue into a buried inaccessible position. A bulky side chain at the DEG position prevents this conformational change, disrupting channel closing. In this model, the gate lies internal to the DEG residue, either at the selectivity filter or within the intracellular vestibule, similar to K+ channels. The two models shown in Fig 8 are not mutually exclusive: elements of both models could be true. However, common to both models is the requirement for a conformational change in the outer vestibule in the gating of ENaC.
|
There are fundamental differences between the gating of ENaC and other members of the DEG/ENaC family of Na+ channels. Whereas nearly all of these channels require a ligand for the channel to open, ENaC is active in the absence of any known stimulus. However, there are also similarities. A bulky side chain at the DEG position increases the activity of several members of the DEG/ENaC family. In addition, in BNC1, the accessibility of a cysteine at the DEG position changed in response to acidic pH; this cysteine was relatively inaccessible at neutral pH, but became accessible at pH 5 (
DEG mutations produce pathology. In C. elegans, DEG mutations in MEC-4, MEC-10, and DEG-1 cause neuronal swelling, lysis, and touch insensitivity. In ENaC, a DEG mutation may also have clinical relevance. Melander and co-workers identified a patient with diabetic nephropathy who had a mutation in the subunit, changing Asn530 to lysine (
![]() |
Footnotes |
---|
1 Abbreviations used in this paper: ENaC, epithelial Na+ channel; MTS, methanethiosulfonate; MTSEA-biotincap, N-biotinylcaproylaminoethyl methanethiosulfonate; MTSES, sodium (2-sulfonatoethyl) methanethiosulfonate; MTSET, [2-(trimethylammonium)ethyl]methanethiosulfonate bromide; Po, open state probability.
![]() |
Acknowledgements |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
We thank Tien Vinh, Sarah Hestekin, and Fotene Gennatos for technical support and Michael Welsh, Christopher Adams, Christopher Benson, and our other laboratory colleagues for helpful discussions and critical review of this manuscript. We thank Ken Volk and John Stokes for providing ENaC subunits in pGEM-HE.
P.M. Snyder was supported by the Roy J. Carver Charitable Trust and by the National Heart, Lung and Blood Institute (grants No. HL-58812 and HL-03575) and National Institute of Diabetes and Digestive Kidney Diseases (grant No. DK-52617) of the National Institutes of Health.
Submitted: 11 May 2000
Revised: 13 October 2000
Accepted: 16 October 2000
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Benos, D.J., Awayda, M.S., Ismailov, I.I., and Johnson, J.P. 1995. Structure and function of amiloride-sensitive Na+ channels. J. Membr. Biol. 143:1-18[Medline].
Boucher, R.C., Stutts, M.J., Knowles, M.R., Cantley, L., and Gatzy, J.T. 1986. Na+ transport in cystic fibrosis respiratory epithelia. Abnormal basal rate and response to adenylate cyclase activation. J. Clin. Investig. 78:1245-1252[Medline].
Canessa, C.M., Schild, L., Buell, G., Thorens, B., Gautschi, I., Horisberger, J.D., and Rossier, B.C. 1994. Amiloride-sensitive epithelial Na+ channel is made of three homologous subunits. Nature 367:463-467[Medline].
Driscoll, M., and Chalfie, M. 1991. The mec-4 gene is a member of a family of Caenorhabditis elegans genes that can mutate to induce neuronal degeneration. Nature 349:588-593[Medline].
Eskandari, S., Snyder, P.M., Kreman, M., Zampighi, G.A., Welsh, M.J., and Wright, E.M. 1999. Number of subunits comprising the epithelial sodium channel. J. Biol. Chem. 274:27281-27286
Firsov, D., Gautschi, I., Merillat, A.M., Rossier, B.C., and Schild, L. 1998. The heterotetrameric architecture of the epithelial sodium channel (ENaC). EMBO (Eur. Mol. Biol. Organ.) J. 17:344-352
Fyfe, G.K., and Canessa, C.M. 1998. Subunit composition determines the single channel kinetics of the epithelial sodium channel. J. Gen. Physiol. 112:423-432
Garty, H., and Palmer, L.G. 1997. Epithelial sodium channels: function, structure, and regulation. Physiol. Rev. 77:359-396
Grunder, S., Firsov, D., Chang, S.S., Jaeger, N.F., Gautschi, I., Schild, L., Lifton, R.P., and Rossier, B.C. 1997. A mutation causing pseudohypoaldosteronism type 1 identifies a conserved glycine that is involved in the gating of the epithelial sodium channel. EMBO (Eur. Mol. Biol. Organ.) J. 16:899-907
Grunder, S., Jaeger, N.F., Gautschi, I., Schild, L., and Rossier, B.C. 1999. Identification of a highly conserved sequence at the N-terminus of the epithelial Na+ channel alpha subunit involved in gating. Pflügers Arch. 438:709-715.
Hille, B. 1992. Ionic Channels of Excitable Membranes. 2nd ed Sunderland, MA, Sinauer Associates Inc, pp. 692.
Hummler, E., Barker, P., Gatzy, J., Beermann, F., Verdumo, C., Schmidt, A., Boucher, R., and Rossier, B.C. 1996. Early death due to defective neonatal lung liquid clearance in alpha-ENaC-deficient mice. Nat. Genet. 12:325-328[Medline].
Imoto, K., Busch, C., Sakmann, B., Mishina, M., Konno, T., Nakai, J., Bujo, H., Mori, Y., Fukuda, K., and Numa, S. 1988. Rings of negatively charged amino acids determine the acetylcholine receptor channel conductance. Nature 335:645-648[Medline].
Ishikawa, T., Marunaka, Y., and Rotin, D. 1998. Electrophysiological characterization of the rat epithelial Na+ channel (rENaC) expressed in MDCK cells. Effects of Na+ and Ca2+. J. Gen. Physiol. 111:825-846
Kellenberger, S., Gautschi, I., and Schild, L. 1999a. A single point mutation in the pore region of the epithelial Na+ channel changes ion selectivity by modifying molecular sieving. Proc. Natl. Acad. Sci. USA. 96:4170-4175
Kellenberger, S., Hoffmann-Pochon, N., Gautschi, I., Schneeberger, E., and Schild, L. 1999b. On the molecular basis of ion permeation in the epithelial Na+ channel. J. Gen. Physiol. 114:13-30
Kosari, F., Sheng, S., Li, J., Mak, D.O., Foskett, J.K., and Kleyman, T.R. 1998. Subunit stoichiometry of the epithelial sodium channel. J. Biol. Chem. 273:13469-13474
Lifton, R.P. 1996. Molecular genetics of human blood pressure variation. Science 272:676-680[Abstract].
McDonald, F.J., Snyder, P.M., McCray, P.B., Jr., and Welsh, M.J. 1994. Cloning, expression, and tissue distribution of a human amiloride-sensitive Na+ channel. Am. J. Physiol. 266:L728-L734
McDonald, F.J., Price, M.P., Snyder, P.M., and Welsh, M.J. 1995. Cloning and expression of the beta- and gamma-subunits of the human epithelial sodium channel. Am. J. Physiol. 268:C1157-C1163
Melander, O., Orho, M., Fagerudd, J., Bengtsson, K., Groop, P.H., Mattiasson, I., Groop, L., and Hulthen, U.L. 1998. Mutations and variants of the epithelial sodium channel gene in Liddle's syndrome and primary hypertension. Hypertension 31:1118-11124
Palmer, L.G., and Frindt, G. 1986. Amiloride-sensitive Na channels from the apical membrane of the rat cortical collecting tubule. Proc. Natl. Acad. Sci. USA 83:2767-2770[Abstract].
Palmer, L.G., and Frindt, G. 1996. Gating of Na channels in the rat cortical collecting tubule: effects of voltage and membrane stretch. J. Gen. Physiol. 107:35-45[Abstract].
Price, M.P., Snyder, P.M., and Welsh, M.J. 1996. Cloning and expression of a novel human brain Na+ channel. J. Biol. Chem. 271:7879-7882
Schild, L., Canessa, C.M., Shimkets, R.A., Gautschi, I., Lifton, R.P., and Rossier, B.C. 1995. A mutation in the epithelial sodium channel causing Liddle disease increases channel activity in the Xenopus laevis oocyte expression system. Proc. Natl. Acad. Sci. USA. 92:5699-5703[Abstract].
Snyder, P.M. 2000. Liddle's syndrome mutations disrupt cAMP-mediated translocation of the epithelial Na+ channel to the cell surface. J. Clin. Investig. 105:45-53
Snyder, P.M., Price, M.P., McDonald, F.J., Adams, C.M., Volk, K.A., Zeiher, B.G., Stokes, J.B., and Welsh, M.J. 1995. Mechanism by which Liddle's syndrome mutations increase activity of a human epithelial Na+ channel. Cell 83:969-978[Medline].
Snyder, P.M., Cheng, C., Prince, L.S., Rogers, J.C., and Welsh, M.J. 1998. Electrophysiological and biochemical evidence that DEG/ENaC cation channels are composed of nine subunits. J. Biol. Chem. 273:681-684
Snyder, P.M., Olson, D.R., and Bucher, D.B. 1999. A pore segment in DEG/ENaC Na+ channels. J. Biol. Chem. 274:28484-28490
Tavernarakis, N., and Driscoll, M. 1997. Molecular modeling of mechanotransduction in the nematode Caenorhabditis elegans. Annu. Rev. Physiol. 59:659-689[Medline].
Volk, K.A., Husted, R.F., Snyder, P.M., and Stokes, J.B. 2000. Kinase regulation of hENaC mediated through a region in the COOH-terminal portion of the alpha-subunit. Am. J. Physiol. 47:C1047-C1054.
Volk, K.A., Sigmund, R.D., Snyder, P.M., McDonald, F.J., Welsh, M.J., and Stokes, J.B. 1995. rENaC is the predominant Na+ channel in the apical membrane of the rat renal inner medullary collecting duct. J. Clin. Investig. 96:2748-2757[Medline].
Waldmann, R., Champigny, G., Voilley, N., Lauritzen, I., and Lazdunski, M. 1996. The mammalian degenerin MDEG, an amiloride-sensitive cation channel activated by mutations causing neurodegeneration in Caenorhabditis elegans. J. Biol. Chem. 271:10433-10436
Zabner, J., Smith, J.J., Karp, P.H., Widdicombe, J.H., and Welsh, M.J. 1998. Loss of CFTR chloride channels alters salt absorption by cystic fibrosis airway epithelia in vitro. Mol. Cell 2:397-403[Medline].