Journal
JOURNAL OF GENERAL PHYSIOLOGY
Volume 130, Issue 4, Pages 399-413Publisher
ROCKEFELLER UNIV PRESS
DOI: 10.1085/jgp.200709800
Keywords
-
Categories
Funding
- NIDDK NIH HHS [R01DK070571, R01DK59594, R01 DK070571, R01 DK059594] Funding Source: Medline
Ask authors/readers for more resources
Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P-2) and phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P-3) 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 beta- 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 beta- 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 beta- and gamma-ENaC were identified as being critical to downregulation of ENaC activity and P-o 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 beta 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 ligandbinding sites within ENaC to modulate open probability.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available