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J. Biol. Chem. 283 (41): 27477-27482

© 2008 by The American Society for Biochemistry and Molecular Biology, Inc.

Intracellular Sodium Regulates Proteolytic Activation of the Epithelial Sodium Channel*

Kristin K. Knight, Danielle M. Wentzlaff, , and Peter M. Snyder1

Departments of Internal Medicine and Molecular Physiology and Biophysics, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa 52242

Abstract: Na+ transport across epithelia is mediated in part by the epithelial Na+ channel ENaC. Previous work indicates that Na+ is an important regulator of ENaC, providing a negative feedback mechanism to maintain Na+ homeostasis. ENaC is synthesized as an inactive precursor, which is activated by proteolytic cleavage of the extracellular domains of the {alpha} and {gamma} subunits. Here we found that Na+ regulates ENaC in part by altering proteolytic activation of the channel. When the Na+ concentration was low, we found that the majority of ENaC at the cell surface was in the cleaved/active state. As Na+ increased, there was a dose-dependent decrease in ENaC cleavage and, hence, ENaC activity. This Na+ effect was dependent on Na+ permeation; cleavage was increased by the ENaC blocker amiloride and by a mutation that decreases ENaC activity ({alpha}H69A) and was reduced by a mutation that activates ENaC (βS520K). Moreover, the Na+ ionophore monensin reversed the effect of the inactivating mutation ({alpha}H69A) on ENaC cleavage, suggesting that intracellular Na+ regulates cleavage. Na+ did not alter activity of Nedd4-2, an E3 ubiquitin ligase that modulates ENaC cleavage, but Na+ reduced ENaC cleavage by exogenous trypsin. Our findings support a model in which intracellular Na+ regulates cleavage by altering accessibility of ENaC cleavage sites to proteases and provide a molecular explanation for the earlier observation that intracellular Na+ inhibits Na+ transport via ENaC (Na+ feedback inhibition).

Received for publication May 30, 2008.

* This work was supported, in whole or in part, by the National Institutes of Health. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 Supported by the National Institutes of Health. To whom correspondence should be addressed: 371 EMRB, University of Iowa, Iowa City, IA 52242. E-mail: peter-snyder{at}

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