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This Article Full Text Full Text (PDF) All Versions of this Article: 294/1/F47    most recent 00399.2007v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Web of Science (9) Citing Articles via Google Scholar Google Scholar Articles by Carattino, M. D. Articles by Kleyman, T. R. Search for Related Content PubMed PubMed Citation Articles by Carattino, M. D. Articles by Kleyman, T. R.

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Aldosterone and Epithelial Na⁺ Channels

Defining an inhibitory domain in the -subunit of the epithelial sodium channel

¹Renal-Electrolyte Division and ³Division of Pulmonary, Allergy, and Critical Care Medicine, Department of Medicine, and ⁴Department of Cell Biology and Physiology, University of Pittsburgh, Pittsburgh, Pennsylvania; and ²NMR Facility, Department of Chemistry, University of Tennessee, Knoxville, Tennessee

Submitted 29 August 2007 ; accepted in final form 19 November 2007

Epithelial sodium channels (ENaC) are processed by proteases as they transit the biosynthetic pathway. We recently observed that furin-dependent processing of the -subunit of ENaC at two sites within its extracellular domain is required for channel activation due to release of a 26-residue inhibitory domain. While channels with -subunits lacking the furin sites are not cleaved and have very low activity, channels lacking the furin consensus sites as well as the tract between these sites (D206–R231) are active. We analyzed channels with a series of deletions in the tract D206–R231 and lacking the -subunit furin consensus sites in Xenopus laevis oocytes. We found an eight-residue tract that, when deleted, restored channel activity to the level found in oocytes expressing wild-type ENaC. A synthetic peptide, LPHPLQRL, representing the tract L211–L218, inhibited wild-type ENaC expressed in oocytes with an IC₅₀ of 0.9 µM, and inhibited channels expressed in collecting duct cells and human primary airway epithelial cells with an IC₅₀s of between 50 and 100 µM. Analyses of peptides with deletions within this inhibitory tract indicate that eight residues is the minimal backbone length that is required for ENaC inhibition. Analyses of 8-mer peptides with conserved and nonconserved substitutions suggest that L¹, P², H³, P⁴, and L⁸ are required for inhibitory activity. Our findings suggest that this eight-residue tract is a key conserved inhibitory domain that provides epithelial cells with a reserve of inactive channels that can be activated as required by proteases.

peptide inhibitors; proteases; furin

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