|
|
|
|||||||
|
|||||||||
J. Biol. Chem., Vol. 277, Issue 12, 9772-9779, March 22, 2002
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
From the Hospital for Sick Children and the Department of
Biochemistry, University of Toronto,
Toronto, Ontario M5G 1X8, Canada
The apically located epithelial
Na+ channel (
Trafficking and Cell Surface Stability of the Epithelial
Na+ Channel Expressed in Epithelial Madin-Darby Canine
Kidney Cells*
,
-ENaC) plays a key role
in the regulation of salt and fluid transport in the kidney and other
epithelia, yet its mode of trafficking to the plasma membrane and its
cell surface stability in mammalian cells are poorly understood.
Because the expression of ENaC in native tissues/cells is very low, we
generated epithelial Madin-Darby canine kidney (MDCK) cells
stably expressing -ENaC, where each subunit is tagged
differentially at the intracellular C terminus and the -subunit is
also Myc-tagged at the ectodomain
(HAMyc,T7FLAG). ENaC expression in these cells was verified by immunoblotting with
antibodies to the tags, and patch clamp analysis has confirmed that the
tagged channel is functional. Moreover, using electron microscopy, we
demonstrated apical, but not basal, membrane localization of ENaC in
these cells. The glycosylation pattern of the intracellular pool of
ENaC revealed peptide N-glycosidase F and
endoglycosidase H sensitivity. Surprisingly, the cell surface pool of
ENaC, analyzed by surface biotinylation, was also core glycosylated and
lacked detectable endoglycosidase H-resistant channels. Extraction of the channel from cells in Triton X-100 demonstrated that both intracellular and cell surface pools of ENaC are largely soluble. Moreover, floatation assays to analyze the presence of ENaC in lipid
rafts showed that both intracellular and cell surface pools of this
channel are not associated with rafts. We have shown previously that
the total cellular pool of ENaC is turned over rapidly
(t1/2 ~ 1-2 h). Using cycloheximide
treatment and surface biotinylation we now demonstrate that the cell
surface pool of ENaC has a similarly short half-life
(t1/2 ~1 h), unlike the long half-life
reported recently for the Xenopus A6 cells. Collectively,
these results help elucidate key aspects of ENaC trafficking and
turnover rates in mammalian kidney epithelial cells.
*
This work was supported in part by the Canadian Cystic
Fibrosis Foundation, the Canadian Institute of Health Research, and by
the International Human Frontier Science Program.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Supported by a University of Toronto open studentship.
§
Present address: Dept. of Biomedical Sciences, Graduate School of
Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan.
¶
Supported by a Canadian Institute of Health Research
investigator award. To whom correspondence should be addressed: Program in Cell Biology, the Hospital for Sick Children, 555 University Ave.,
Toronto, Ontario M5G 1X8, Canada. Tel.: 416-813-5098; Fax: 416-813-5771; E mail: drotin{at}sickkids.ca.
Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  A. C.G. van Ginneken MinK: a double-hearted partner in cardiac repolarization Cardiovasc Res, June 1, 2009; 82(3): 390 - 391. [Full Text] [PDF]
|
|
|
|
 |
|
 H.-W. Su, S.-W. Wang, F. K. Ghishan, P. R. Kiela, and M.-J. Tang Cell confluency-induced Stat3 activation regulates NHE3 expression by recruiting Sp1 and Sp3 to the proximal NHE3 promoter region during epithelial dome formation Am J Physiol Cell Physiol, January 1, 2009; 296(1): C13 - C24. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. B. Butterworth, R. S. Edinger, R. A. Frizzell, and J. P. Johnson Regulation of the epithelial sodium channel by membrane trafficking Am J Physiol Renal Physiol, January 1, 2009; 296(1): F10 - F24. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Yu, M. N. Helms, Q. Yue, and D. C. Eaton Single-channel analysis of functional epithelial sodium channel (ENaC) stability at the apical membrane of A6 distal kidney cells Am J Physiol Renal Physiol, November 1, 2008; 295(5): F1519 - F1527. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Obeid ERP57 Membrane Translocation Dictates the Immunogenicity of Tumor Cell Death by Controlling the Membrane Translocation of Calreticulin J. Immunol., August 15, 2008; 181(4): 2533 - 2543. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Kabra, K. K. Knight, R. Zhou, and P. M. Snyder Nedd4-2 Induces Endocytosis and Degradation of Proteolytically Cleaved Epithelial Na+ Channels J. Biol. Chem., March 7, 2008; 283(10): 6033 - 6039. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. Annaba, Z. Sarwar, P. Kumar, S. Saksena, J. R. Turner, P. K. Dudeja, R. K. Gill, and W. A. Alrefai Modulation of ileal bile acid transporter (ASBT) activity by depletion of plasma membrane cholesterol: association with lipid rafts Am J Physiol Gastrointest Liver Physiol, February 1, 2008; 294(2): G489 - G497. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W. G. Hill, M. B. Butterworth, H. Wang, R. S. Edinger, J. Lebowitz, K. W. Peters, R. A. Frizzell, and J. P. Johnson The Epithelial Sodium Channel (ENaC) Traffics to Apical Membrane in Lipid Rafts in Mouse Cortical Collecting Duct Cells J. Biol. Chem., December 28, 2007; 282(52): 37402 - 37411. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. B. Butterworth, R. S. Edinger, H. Ovaa, D. Burg, J. P. Johnson, and R. A. Frizzell The Deubiquitinating Enzyme UCH-L3 Regulates the Apical Membrane Recycling of the Epithelial Sodium Channel J. Biol. Chem., December 28, 2007; 282(52): 37885 - 37893. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Adebamiro, Y. Cheng, U. S. Rao, H. Danahay, and R. J. Bridges A Segment of {gamma} ENaC Mediates Elastase Activation of Na+ Transport J. Gen. Physiol., November 26, 2007; 130(6): 611 - 629. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. Pruliere-Escabasse, C. Planes, E. Escudier, P. Fanen, A. Coste, and C. Clerici Modulation of Epithelial Sodium Channel Trafficking and Function by Sodium 4-Phenylbutyrate in Human Nasal Epithelial Cells J. Biol. Chem., November 23, 2007; 282(47): 34048 - 34057. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. M. Mueller, O. B. Kashlan, J. B. Bruns, A. B. Maarouf, M. Aridor, T. R. Kleyman, and R. P. Hughey Epithelial Sodium Channel Exit from the Endoplasmic Reticulum Is Regulated by a Signal within the Carboxyl Cytoplasmic Domain of the {alpha} Subunit J. Biol. Chem., November 16, 2007; 282(46): 33475 - 33483. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Tiwari, L. Nordquist, V. K. M. Halagappa, and C. A. Ecelbarger Trafficking of ENaC subunits in response to acute insulin in mouse kidney Am J Physiol Renal Physiol, July 1, 2007; 293(1): F178 - F185. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Grasberger, X. De Deken, F. Miot, J. Pohlenz, and S. Refetoff Missense Mutations of Dual Oxidase 2 (DUOX2) Implicated in Congenital Hypothyroidism Have Impaired Trafficking in Cells Reconstituted with DUOX2 Maturation Factor Mol. Endocrinol., June 1, 2007; 21(6): 1408 - 1421. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H.-W. Su, H.-H. Yeh, S.-W. Wang, M.-R. Shen, T.-L. Chen, P. R. Kiela, F. K. Ghishan, and M.-J. Tang Cell Confluence-induced Activation of Signal Transducer and Activator of Transcription-3 (Stat3) Triggers Epithelial Dome Formation via Augmentation of Sodium Hydrogen Exchanger-3 (NHE3) Expression J. Biol. Chem., March 30, 2007; 282(13): 9883 - 9894. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Hinzpeter, J. Fritsch, F. Borot, S. Trudel, D.-L. Vieu, F. Brouillard, M. Baudouin-Legros, J. Clain, A. Edelman, and M. Ollero Membrane Cholesterol Content Modulates ClC-2 Gating and Sensitivity to Oxidative Stress J. Biol. Chem., January 26, 2007; 282(4): 2423 - 2432. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. J. D. Espiritu, A. A. Bernardo, and J. A. L. Arruda Role of NH2 and COOH termini in targeting, stability, and activity of sodium bicarbonate cotransporter 1 Am J Physiol Renal Physiol, September 1, 2006; 291(3): F588 - F596. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 O. Staub and F. Verrey Impact of Nedd4 Proteins and Serum and Glucocorticoid-Induced Kinases on Epithelial Na+ Transport in the Distal Nephron J. Am. Soc. Nephrol., November 1, 2005; 16(11): 3167 - 3174. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Adebamiro, Y. Cheng, J. P. Johnson, and R. J. Bridges Endogenous Protease Activation of ENaC: Effect of Serine Protease Inhibition on ENaC Single Channel Properties J. Gen. Physiol., September 26, 2005; 126(4): 339 - 352. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. A. Itani, J. B. Stokes, and C. P. Thomas Nedd4-2 isoforms differentially associate with ENaC and regulate its activity Am J Physiol Renal Physiol, August 1, 2005; 289(2): F334 - F346. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Kuliawat, J. Ramos-Castaneda, Y. Liu, and P. Arvan Intracellular Trafficking of Thyroid Peroxidase to the Cell Surface J. Biol. Chem., July 29, 2005; 280(30): 27713 - 27718. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Malik, Q. Yue, G. Yue, X. J. Chen, S. R. Price, W. E. Mitch, and D. C. Eaton Role of Nedd4-2 and polyubiquitination in epithelial sodium channel degradation in untransfected renal A6 cells expressing endogenous ENaC subunits Am J Physiol Renal Physiol, July 1, 2005; 289(1): F107 - F116. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. A. Caldwell, R. C. Boucher, and M. J. Stutts Neutrophil elastase activates near-silent epithelial Na+ channels and increases airway epithelial Na+ transport Am J Physiol Lung Cell Mol Physiol, May 1, 2005; 288(5): L813 - L819. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Y. Sautin, M. Lu, A. Gaugler, L. Zhang, and S. L. Gluck Phosphatidylinositol 3-Kinase-Mediated Effects of Glucose on Vacuolar H+-ATPase Assembly, Translocation, and Acidification of Intracellular Compartments in Renal Epithelial Cells Mol. Cell. Biol., January 15, 2005; 25(2): 575 - 589. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. B. Butterworth, R. S. Edinger, J. P. Johnson, and R. A. Frizzell Acute ENaC Stimulation by cAMP in a Kidney Cell Line is Mediated by Exocytic Insertion from a Recycling Channel Pool J. Gen. Physiol., December 28, 2004; 125(1): 81 - 101. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. M. Sedensky, J. M. Siefker, J. Y. Koh, D. M. Miller III, and P. G. Morgan A stomatin and a degenerin interact in lipid rafts of the nervous system of Caenorhabditis elegans Am J Physiol Cell Physiol, August 1, 2004; 287(2): C468 - C474. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Mohan, J. R. Bruns, K. M. Weixel, R. S. Edinger, J. B. Bruns, T. R. Kleyman, J. P. Johnson, and O. A. Weisz Differential Current Decay Profiles of Epithelial Sodium Channel Subunit Combinations in Polarized Renal Epithelial Cells J. Biol. Chem., July 30, 2004; 279(31): 32071 - 32078. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Dagenais, R. Frechette, Y. Yamagata, T. Yamagata, J.-F. Carmel, M.-E. Clermont, E. Brochiero, C. Masse, and Y. Berthiaume Downregulation of ENaC activity and expression by TNF-{alpha} in alveolar epithelial cells Am J Physiol Lung Cell Mol Physiol, February 1, 2004; 286(2): L301 - L311. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. A. Weisz and J. P. Johnson Noncoordinate regulation of ENaC: paradigm lost? Am J Physiol Renal Physiol, November 1, 2003; 285(5): F833 - F842. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Ishikawa, C. Jiang, M. J. Stutts, Y. Marunaka, and D. Rotin Regulation of the Epithelial Na+ Channel by Cytosolic ATP J. Biol. Chem., October 3, 2003; 278(40): 38276 - 38286. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. B. Bruns, B. Hu, Y. J. Ahn, S. Sheng, R. P. Hughey, and T. R. Kleyman Multiple epithelial Na+ channel domains participate in subunit assembly Am J Physiol Renal Physiol, October 1, 2003; 285(4): F600 - F609. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. P. Hughey, G. M. Mueller, J. B. Bruns, C. L. Kinlough, P. A. Poland, K. L. Harkleroad, M. D. Carattino, and T. R. Kleyman Maturation of the Epithelial Na+ Channel Involves Proteolytic Processing of the {alpha}- and {gamma}-Subunits J. Biol. Chem., September 26, 2003; 278(39): 37073 - 37082. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Auberson, N. Hoffmann-Pochon, A. Vandewalle, S. Kellenberger, and L. Schild Epithelial Na+ channel mutants causing Liddle's syndrome retain ability to respond to aldosterone and vasopressin Am J Physiol Renal Physiol, September 1, 2003; 285(3): F459 - F471. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. E. Booth and J. D. Stockand Targeted degradation of ENaC in response to PKC activation of the ERK1/2 cascade Am J Physiol Renal Physiol, May 1, 2003; 284(5): F938 - F947. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. W. Leung, R. G. Contreras, C. Flores-Maldonado, M. Cereijido, and E. Rodriguez-Boulan Inhibitors of glycosphingolipid biosynthesis reduce transepithelial electrical resistance in MDCK I and FRT cells Am J Physiol Cell Physiol, April 1, 2003; 284(4): C1021 - C1030. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. G. Shlyonsky, F. Mies, and S. Sariban-Sohraby Epithelial sodium channel activity in detergent-resistant membrane microdomains Am J Physiol Renal Physiol, January 1, 2003; 284(1): F182 - F188. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Planes, M. Blot-Chabaud, M. A. Matthay, S. Couette, T. Uchida, and C. Clerici Hypoxia and beta 2-Agonists Regulate Cell Surface Expression of the Epithelial Sodium Channel in Native Alveolar Epithelial Cells J. Biol. Chem., November 27, 2002; 277(49): 47318 - 47324. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. Jin, T. M. Weiger, and I. B. Levitan Reciprocal Modulation between the alpha and beta 4 Subunits of hSlo Calcium-dependent Potassium Channels J. Biol. Chem., November 8, 2002; 277(46): 43724 - 43729. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Guipponi, G. Vuagniaux, M. Wattenhofer, K. Shibuya, M. Vazquez, L. Dougherty, N. Scamuffa, E. Guida, M. Okui, C. Rossier, et al. The transmembrane serine protease (TMPRSS3) mutated in deafness DFNB8/10 activates the epithelial sodium channel (ENaC) in vitro Hum. Mol. Genet., November 1, 2002; 11(23): 2829 - 2836. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W. G. Hill, B. An, and J. P. Johnson Endogenously Expressed Epithelial Sodium Channel Is Present in Lipid Rafts in A6 Cells J. Biol. Chem., September 6, 2002; 277(37): 33541 - 33544. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Kamynina and O. Staub Concerted action of ENaC, Nedd4-2, and Sgk1 in transepithelial Na+ transport Am J Physiol Renal Physiol, September 1, 2002; 283(3): F377 - F387. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. C. Rossier Hormonal Regulation of the Epithelial Sodium Channel ENaC: N or Po? J. Gen. Physiol., June 24, 2002; 120(1): 67 - 70. [Full Text] [PDF]
|
|
|
|
|
|
D. A. de la Rosa, H. Li, and C. M. Canessa Effects of Aldosterone on Biosynthesis, Traffic, and Functional Expression of Epithelial Sodium Channels in A6 Cells J. Gen. Physiol., April 15, 2002; 119(5): 427 - 442. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. A. de la Rosa, H. Li, and C. M. Canessa Effects of Aldosterone on Biosynthesis, Traffic, and Functional Expression of Epithelial Sodium Channels in A6 Cells J. Gen. Physiol., April 15, 2002; 119(5): 427 - 442. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| All ASBMB Journals | Molecular and Cellular Proteomics |
| Journal of Lipid Research | ASBMB Today |