The Journal of General Physiology
Scientifica: Experts in Electrophysiology
  Home | Help | Feedback | Subscriptions | Archive | Search | Table of Contents
Published online
doi:10.1085/jgp.200709800
The Journal of General Physiology, Vol. 130, No. 4, 399-413
The Rockefeller University Press, 0022-1295 $30.00
© Pochynyuk et al.
This Article
Right arrow Full Text
Right arrow Full Text (PDF, 4350K)
Right arrow PPT slides of all figures
Right arrow Alert me when this article is cited
Right arrow Citation Map
Services
Right arrow Email this article
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new content in the JGP
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via CrossRef
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Pochynyuk, O.
Right arrow Articles by Stockand, J. D.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Pochynyuk, O.
Right arrow Articles by Stockand, J. D.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Facebook   Add to Reddit   Add to Technorati   Add to Twitter  
What's this?

ARTICLE

 Molecular Determinants of PI(4,5)P2 and PI(3,4,5)P3 Regulation of the Epithelial Na+ Channel



Oleh Pochynyuk1, Qiusheng Tong1, Jorge Medina1, Alain Vandewalle2,3, Alexander Staruschenko4, Vladislav Bugaj1, and James D. Stockand1

1 University of Texas Health Science Center, Department of Physiology, San Antonio, TX 78229
2 INSERM U773, Centre de Recherche Biomedicale Bichat-Beaujon, CRB3 Paris F-75018, France
3 Universite Paris 7, Denis Diderot, site Bichat, Paris F-75018, France
4 Medical College of Wisconsin, Department of Physiology and Kidney Disease Center, Milwaukee, WI 53226

Correspondence to James D. Stockand: stockand{at}uthscsa.edu

Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P3) are physiologically important second messengers. These molecules bind effector proteins to modulate activity. Several types of ion channels, including the epithelial Na+ channel (ENaC), are phosphoinositide effectors capable of directly interacting with these signaling molecules. Little, however, is known of the regions within ENaC and other ion channels important to phosphoinositide binding and modulation. Moreover, the molecular mechanism of this regulation, in many instances, remains obscure. Here, we investigate modulation of ENaC by PI(3,4,5)P3 and PI(4,5)P2 to begin identifying the molecular determinants of this regulation. We identify intracellular regions near the inner membrane interface just following the second transmembrane domains in ß- and {gamma}- but not {alpha}-ENaC as necessary for PI(3,4,5)P2 but not PI(4,5)P2 modulation. Charge neutralization of conserved basic amino acids within these regions demonstrated that these polar residues are critical to phosphoinositide regulation. Single channel analysis, moreover, reveals that the regions just following the second transmembrane domains in ß- and {gamma}-ENaC are critical to PI(3,4,5)P3 augmentation of ENaC open probability, thus, defining mechanism. Unexpectedly, intracellular domains within the extreme N terminus of ß- and {gamma}-ENaC were identified as being critical to down-regulation of ENaC activity and Po in response to depletion of membrane PI(4,5)P2. These regions of the channel played no identifiable role in a PI(3,4,5)P3 response. Again, conserved positive-charged residues within these domains were particularly important, being necessary for exogenous PI(4,5)P2 to increase open probability. We conclude that ß and {gamma} subunits bestow phosphoinositide sensitivity to ENaC with distinct regions of the channel being critical to regulation by PI(3,4,5)P3 and PI(4,5)P2. This argues that these phosphoinositides occupy distinct ligand-binding sites within ENaC to modulate open probability.

Abbreviations used in this paper: ENaC, epithelial Na+ channel; PH, pleckstrin homology; PI3-K, phosphatidylinositide 3-OH kinase; PI(4,5)P2, phosphatidylinositol 4,5-bisphosphate; PI(3,4,5)P3, phosphatidylinositol 3,4,5-trisphosphate; TIRF, total internal reflection fluorescence.


Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Facebook Facebook   Add to Reddit Reddit   Add to Technorati Technorati   Add to Twitter Twitter    What's this?


This article has been cited by other articles:


Home page
 J. Am. Soc. Nephrol.Home page
V. Vallon, E. Hummler, T. Rieg, O. Pochynyuk, V. Bugaj, J. Schroth, G. Dechenes, B. Rossier, R. Cunard, and J. Stockand
Thiazolidinedione-Induced Fluid Retention Is Independent of Collecting Duct {alpha}ENaC Activity
J. Am. Soc. Nephrol., April 1, 2009; 20(4): 721 - 729.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
O. Pochynyuk, V. Bugaj, T. Rieg, P. A. Insel, E. Mironova, V. Vallon, and J. D. Stockand
Paracrine Regulation of the Epithelial Na+ Channel in the Mammalian Collecting Duct by Purinergic P2Y2 Receptor Tone
J. Biol. Chem., December 26, 2008; 283(52): 36599 - 36607.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Renal Physiol.Home page
V. Bugaj, O. Pochynyuk, E. Mironova, A. Vandewalle, J. L. Medina, and J. D. Stockand
Regulation of the epithelial Na+ channel by endothelin-1 in rat collecting duct
Am J Physiol Renal Physiol, October 1, 2008; 295(4): F1063 - F1070.
[Abstract] [Full Text] [PDF]

Home page
 J. Physiol.Home page
O. J. Mace, A. M. Woollhead, and D. L. Baines
AICAR activates AMPK and alters PIP2 association with the epithelial sodium channel ENaC to inhibit Na+ transport in H441 lung epithelial cells
J. Physiol., September 15, 2008; 586(18): 4541 - 4557.
[Abstract] [Full Text] [PDF]


  Home | Help | Feedback | Subscriptions | Archive | Search | Table of Contents