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. 1998 Dec 15;336(Pt 3):705–710. doi: 10.1042/bj3360705

Cell surface expression and biosynthesis of epithelial Na+ channels.

L S Prince 1, M J Welsh 1
PMCID: PMC1219923  PMID: 9841884

Abstract

The epithelial Na+ channel (ENaC) complex is composed of three homologous subunits: alpha, beta and gamma. Mutations in ENaC subunits can increase the number of channels on the cell surface, causing a hereditary form of hypertension called Liddle's syndrome, or can decrease channel activity, causing pseudohypoaldosteronism type I, a salt-wasting disease of infancy. To investigate surface expression, we studied ENaC subunits expressed in COS-7 and HEK293 cells. Using surface biotinylation and protease sensitivity, we found that when individual ENaC subunits are expressed alone, they traffic to the cell surface. The subunits are glycosylated with high-mannose oligosaccharides, but seem to have the carbohydrate removed before they reach the cell surface. Moreover, subunits form a complex that cannot be disrupted by several non-ionic detergents. The pattern of glycosylation and detergent solubility/insolubility persists when the N-teminal and C-terminal cytoplasmic regions of ENaC are removed. With co-expression of all three ENaC subunits, the insoluble complex is the predominant species. These results show that ENaC and its family members are unique in their trafficking, biochemical characteristics and post-translational modifications.

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Selected References

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  1. Adams C. M., Snyder P. M., Welsh M. J. Interactions between subunits of the human epithelial sodium channel. J Biol Chem. 1997 Oct 24;272(43):27295–27300. doi: 10.1074/jbc.272.43.27295. [DOI] [PubMed] [Google Scholar]
  2. Benos D. J., Awayda M. S., Ismailov I. I., Johnson J. P. Structure and function of amiloride-sensitive Na+ channels. J Membr Biol. 1995 Jan;143(1):1–18. doi: 10.1007/BF00232519. [DOI] [PubMed] [Google Scholar]
  3. Canessa C. M., Horisberger J. D., Rossier B. C. Epithelial sodium channel related to proteins involved in neurodegeneration. Nature. 1993 Feb 4;361(6411):467–470. doi: 10.1038/361467a0. [DOI] [PubMed] [Google Scholar]
  4. Canessa C. M., Schild L., Buell G., Thorens B., Gautschi I., Horisberger J. D., Rossier B. C. Amiloride-sensitive epithelial Na+ channel is made of three homologous subunits. Nature. 1994 Feb 3;367(6462):463–467. doi: 10.1038/367463a0. [DOI] [PubMed] [Google Scholar]
  5. Caplan S., Zeliger S., Wang L., Baniyash M. Cell-surface-expressed T-cell antigen-receptor zeta chain is associated with the cytoskeleton. Proc Natl Acad Sci U S A. 1995 May 23;92(11):4768–4772. doi: 10.1073/pnas.92.11.4768. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chang S. S., Grunder S., Hanukoglu A., Rösler A., Mathew P. M., Hanukoglu I., Schild L., Lu Y., Shimkets R. A., Nelson-Williams C. Mutations in subunits of the epithelial sodium channel cause salt wasting with hyperkalaemic acidosis, pseudohypoaldosteronism type 1. Nat Genet. 1996 Mar;12(3):248–253. doi: 10.1038/ng0396-248. [DOI] [PubMed] [Google Scholar]
  7. Firsov D., Schild L., Gautschi I., Mérillat A. M., Schneeberger E., Rossier B. C. Cell surface expression of the epithelial Na channel and a mutant causing Liddle syndrome: a quantitative approach. Proc Natl Acad Sci U S A. 1996 Dec 24;93(26):15370–15375. doi: 10.1073/pnas.93.26.15370. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. García-Añoveros J., Derfler B., Neville-Golden J., Hyman B. T., Corey D. P. BNaC1 and BNaC2 constitute a new family of human neuronal sodium channels related to degenerins and epithelial sodium channels. Proc Natl Acad Sci U S A. 1997 Feb 18;94(4):1459–1464. doi: 10.1073/pnas.94.4.1459. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Garty H., Palmer L. G. Epithelial sodium channels: function, structure, and regulation. Physiol Rev. 1997 Apr;77(2):359–396. doi: 10.1152/physrev.1997.77.2.359. [DOI] [PubMed] [Google Scholar]
  10. Hansson J. H., Nelson-Williams C., Suzuki H., Schild L., Shimkets R., Lu Y., Canessa C., Iwasaki T., Rossier B., Lifton R. P. Hypertension caused by a truncated epithelial sodium channel gamma subunit: genetic heterogeneity of Liddle syndrome. Nat Genet. 1995 Sep;11(1):76–82. doi: 10.1038/ng0995-76. [DOI] [PubMed] [Google Scholar]
  11. Helms J. B., Rothman J. E. Inhibition by brefeldin A of a Golgi membrane enzyme that catalyses exchange of guanine nucleotide bound to ARF. Nature. 1992 Nov 26;360(6402):352–354. doi: 10.1038/360352a0. [DOI] [PubMed] [Google Scholar]
  12. Hsu V. W., Yuan L. C., Nuchtern J. G., Lippincott-Schwartz J., Hammerling G. J., Klausner R. D. A recycling pathway between the endoplasmic reticulum and the Golgi apparatus for retention of unassembled MHC class I molecules. Nature. 1991 Aug 1;352(6334):441–444. doi: 10.1038/352441a0. [DOI] [PubMed] [Google Scholar]
  13. Ismailov I. I., Awayda M. S., Berdiev B. K., Bubien J. K., Lucas J. E., Fuller C. M., Benos D. J. Triple-barrel organization of ENaC, a cloned epithelial Na+ channel. J Biol Chem. 1996 Jan 12;271(2):807–816. doi: 10.1074/jbc.271.2.807. [DOI] [PubMed] [Google Scholar]
  14. Lingueglia E., Champigny G., Lazdunski M., Barbry P. Cloning of the amiloride-sensitive FMRFamide peptide-gated sodium channel. Nature. 1995 Dec 14;378(6558):730–733. doi: 10.1038/378730a0. [DOI] [PubMed] [Google Scholar]
  15. Lingueglia E., Voilley N., Waldmann R., Lazdunski M., Barbry P. Expression cloning of an epithelial amiloride-sensitive Na+ channel. A new channel type with homologies to Caenorhabditis elegans degenerins. FEBS Lett. 1993 Feb 22;318(1):95–99. doi: 10.1016/0014-5793(93)81336-x. [DOI] [PubMed] [Google Scholar]
  16. Lisanti M. P., Le Bivic A., Sargiacomo M., Rodriguez-Boulan E. Steady-state distribution and biogenesis of endogenous Madin-Darby canine kidney glycoproteins: evidence for intracellular sorting and polarized cell surface delivery. J Cell Biol. 1989 Nov;109(5):2117–2127. doi: 10.1083/jcb.109.5.2117. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Mallabiabarrena A., Fresno M., Alarcón B. An endoplasmic reticulum retention signal in the CD3 epsilon chain of the T-cell receptor. Nature. 1992 Jun 18;357(6379):593–596. doi: 10.1038/357593a0. [DOI] [PubMed] [Google Scholar]
  18. McDonald F. J., Price M. P., Snyder P. M., Welsh M. J. Cloning and expression of the beta- and gamma-subunits of the human epithelial sodium channel. Am J Physiol. 1995 May;268(5 Pt 1):C1157–C1163. doi: 10.1152/ajpcell.1995.268.5.C1157. [DOI] [PubMed] [Google Scholar]
  19. McDonald F. J., Snyder P. M., McCray P. B., Jr, Welsh M. J. Cloning, expression, and tissue distribution of a human amiloride-sensitive Na+ channel. Am J Physiol. 1994 Jun;266(6 Pt 1):L728–L734. doi: 10.1152/ajplung.1994.266.6.L728. [DOI] [PubMed] [Google Scholar]
  20. McDonald F. J., Welsh M. J. Binding of the proline-rich region of the epithelial Na+ channel to SH3 domains and its association with specific cellular proteins. Biochem J. 1995 Dec 1;312(Pt 2):491–497. doi: 10.1042/bj3120491. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. North R. A. Families of ion channels with two hydrophobic segments. Curr Opin Cell Biol. 1996 Aug;8(4):474–483. doi: 10.1016/s0955-0674(96)80023-8. [DOI] [PubMed] [Google Scholar]
  22. Price M. P., Snyder P. M., Welsh M. J. Cloning and expression of a novel human brain Na+ channel. J Biol Chem. 1996 Apr 5;271(14):7879–7882. doi: 10.1074/jbc.271.14.7879. [DOI] [PubMed] [Google Scholar]
  23. Rotin D., Bar-Sagi D., O'Brodovich H., Merilainen J., Lehto V. P., Canessa C. M., Rossier B. C., Downey G. P. An SH3 binding region in the epithelial Na+ channel (alpha rENaC) mediates its localization at the apical membrane. EMBO J. 1994 Oct 3;13(19):4440–4450. doi: 10.1002/j.1460-2075.1994.tb06766.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Sargiacomo M., Sudol M., Tang Z., Lisanti M. P. Signal transducing molecules and glycosyl-phosphatidylinositol-linked proteins form a caveolin-rich insoluble complex in MDCK cells. J Cell Biol. 1993 Aug;122(4):789–807. doi: 10.1083/jcb.122.4.789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Schweizer A., Rohrer J., Hauri H. P., Kornfeld S. Retention of p63 in an ER-Golgi intermediate compartment depends on the presence of all three of its domains and on its ability to form oligomers. J Cell Biol. 1994 Jul;126(1):25–39. doi: 10.1083/jcb.126.1.25. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Sheng Z., Vanderpuye O. A., Hull S. R., Carraway C. A., Carraway K. L. Topography and microfilament core association of a cell surface glycoprotein of ascites tumor cell microvilli. J Cell Biochem. 1989 Aug;40(4):453–466. doi: 10.1002/jcb.240400406. [DOI] [PubMed] [Google Scholar]
  27. Shimkets R. A., Warnock D. G., Bositis C. M., Nelson-Williams C., Hansson J. H., Schambelan M., Gill J. R., Jr, Ulick S., Milora R. V., Findling J. W. Liddle's syndrome: heritable human hypertension caused by mutations in the beta subunit of the epithelial sodium channel. Cell. 1994 Nov 4;79(3):407–414. doi: 10.1016/0092-8674(94)90250-x. [DOI] [PubMed] [Google Scholar]
  28. Snyder P. M., McDonald F. J., Stokes J. B., Welsh M. J. Membrane topology of the amiloride-sensitive epithelial sodium channel. J Biol Chem. 1994 Sep 30;269(39):24379–24383. [PubMed] [Google Scholar]
  29. Snyder P. M., Price M. P., McDonald F. J., Adams C. M., Volk K. A., Zeiher B. G., Stokes J. B., Welsh M. J. Mechanism by which Liddle's syndrome mutations increase activity of a human epithelial Na+ channel. Cell. 1995 Dec 15;83(6):969–978. doi: 10.1016/0092-8674(95)90212-0. [DOI] [PubMed] [Google Scholar]
  30. Tavernarakis N., Driscoll M. Molecular modeling of mechanotransduction in the nematode Caenorhabditis elegans. Annu Rev Physiol. 1997;59:659–689. doi: 10.1146/annurev.physiol.59.1.659. [DOI] [PubMed] [Google Scholar]
  31. Waldmann R., Champigny G., Voilley N., Lauritzen I., Lazdunski M. The mammalian degenerin MDEG, an amiloride-sensitive cation channel activated by mutations causing neurodegeneration in Caenorhabditis elegans. J Biol Chem. 1996 May 3;271(18):10433–10436. doi: 10.1074/jbc.271.18.10433. [DOI] [PubMed] [Google Scholar]
  32. Weng S., Spiro R. G. Demonstration of a peptide:N-glycosidase in the endoplasmic reticulum of rat liver. Biochem J. 1997 Mar 1;322(Pt 2):655–661. doi: 10.1042/bj3220655. [DOI] [PMC free article] [PubMed] [Google Scholar]

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