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Originally published In Press as doi:10.1074/jbc.C100737200 on January 23, 2002

J. Biol. Chem., Vol. 277, Issue 10, 7641-7644, March 8, 2002
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ACCELERATED PUBLICATION
Anionic Phospholipids Regulate Native and Expressed Epithelial Sodium Channel (ENaC)*

He-Ping MaDagger , Sunil Saxena, and David G. Warnock

From the Department of Medicine, Division of Nephrology, The University of Alabama at Birmingham, Birmingham, Alabama 35294

Using patch clamp techniques, we found that the epithelial sodium channel (ENaC) activity in the apical membrane of A6 distal nephron cells showed a sudden rundown beginning at 4 min after forming the inside-out configuration. This sudden rundown was prevented by addition of anionic phospholipids such as phosphatidylinositol 4,5-bisphosphate (PIP2), phosphatidylinositol 3,4,5-trisphosphate (PIP3), and phosphatidylserine (PS) to the "cytoplasmic" bath. Conversely, chelation of endogenous PIP2 with anti-PIP2 antibody, hydrolysis of PIP2 with either exogenous phospholipase C (PLC) or activation of endogenous PLC by extracellular ATP, or application of the positively charged molecule, poly-L-lysine, accelerated channel rundown. However, neutral phosphatidylcholine had no effect on ENaC activity. By two-electrode voltage clamp recordings, we demonstrated that PIP2 and PIP3 significantly increased amiloride-sensitive current in Xenopus oocytes injected with cRNAs of rat alpha -, beta -, and gamma -ENaC. However, PIP2 and PIP3 did not affect surface expression of ENaC, indicating that PIP2 and PIP3 regulate ENaC at the level of the inner plasma membrane through a mechanism that is independent of ENaC trafficking. These data suggest that anionic phospholipids may mediate the regulation of ENaC by PLC- or phosphoinositide 3-kinase-coupled receptors.

* This work was supported by a National Kidney Foundation Young Investigator Award (to H.-P. M.).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.

Dagger To whom all correspondence should be addressed: The University of Alabama at Birmingham, Dept. of Medicine, Division of Nephrology, 1530 Third Ave. South, Sparks Bldg. 865, Birmingham, AL 35294-0017. Tel.: 205-934-3907; Fax: 205-934-1147; E-mail: hepingma@uab.edu.

Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.


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