Nedd4-2 isoforms differentially associate with ENaC and regulate its activity

Omar A Itani; John B Stokes; Christie P Thomas

doi:10.1152/ajprenal.00394.2004

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Nedd4-2 isoforms differentially associate with ENaC and regulate its activity

Journal article

Open access

Peer reviewed

Nedd4-2 isoforms differentially associate with ENaC and regulate its activity

Omar A Itani, John B Stokes and Christie P Thomas

American journal of physiology. Renal physiology, Vol.289(2), pp.F334-346

08/2005

DOI: 10.1152/ajprenal.00394.2004

PMID: 15814530

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Abstract

Mutations that disrupt a PY motif in epithelial Na(+) channel (ENaC) subunits increase surface expression of Na(+) channels in the collecting duct, resulting in greater Na(+) reabsorption. Nedd4 and Nedd4-2 have been identified as ubiquitin ligases that can interact with ENaC via its PY motifs to regulate channel activity. We recently reported that human Nedd4-2 (hNedd4-2) is expressed as many isoforms because of alternative promoter usage and/or variable splicing. To understand the relevance of hNedd4-2 isoforms for collecting duct Na(+) transport, we studied the interaction with ENaC and the intracellular localization and function of the following three naturally occurring hNedd4-2 isoforms: full-length Nedd4-2 (Nedd4-2), Nedd4-2 lacking the NH(2)-terminal C2 domain (Nedd4-2DeltaC2), and Nedd4-2 lacking the C2 domain and WW domains 2 and 3 (Nedd4-2DeltaWW2,3). Nedd4-2 and Nedd4-2DeltaC2 associate with ENaC and robustly reduce Na(+) transport in Xenopus oocytes, whereas the interaction with and functional effect of Nedd4-2DeltaWW2,3 on ENaC is weak. Nedd4-2 is expressed in the mouse collecting duct, and overexpression of Nedd4-2 reduces endogenous ENaC activity in a collecting duct cell line. This reduction in ENaC activity can be reversed early with exposure to dexamethasone, an effect that is associated with an increase in sgk1 abundance. The C2 domain is required to target Nedd4-2 to the plasma membrane in response to elevation of intracellular Ca(2+) concentration ([Ca(2+)](i)) in MDCK cells, although it does not appear to mediate the inhibitory effect of [Ca(2+)](i) on Na(+) transport. Our data illustrate that naturally occurring hNedd4-2 isoforms differentially associate with ENaC to regulate its activity.

Xenopus

Immunoprecipitation

Cercopithecus aethiops

Molecular Sequence Data

Cytoplasm - metabolism

Male

Sodium Channels - physiology

Isomerism

RNA - genetics

Epithelial Sodium Channels

Transfection

Sodium Channels - metabolism

Adenoviridae - genetics

Female

Nedd4 Ubiquitin Protein Ligases

Endosomal Sorting Complexes Required for Transport

Oocytes - metabolism

Mice, Inbred C57BL

Ubiquitin-Protein Ligases - metabolism

Xenopus Proteins

Reverse Transcriptase Polymerase Chain Reaction

Blotting, Western

Kidney Tubules, Collecting - metabolism

RNA - biosynthesis

Animals

Nuclease Protection Assays

Homeostasis - physiology

Fluorescent Antibody Technique

Glucocorticoids - pharmacology

Mice

In Vitro Techniques

COS Cells

Details

Title: Subtitle: Nedd4-2 isoforms differentially associate with ENaC and regulate its activity
Creators: Omar A Itani - Department of Internal Medicine, University of Iowa, Iowa City, 52242, USA
John B Stokes
Christie P Thomas
Resource Type: Journal article
Publication Details: American journal of physiology. Renal physiology, Vol.289(2), pp.F334-346
DOI: 10.1152/ajprenal.00394.2004
PMID: 15814530
NLM abbreviation: Am J Physiol Renal Physiol
ISSN: 1931-857X
eISSN: 1522-1466
Publisher: United States
Grant note: DK-54348 / NIDDK NIH HHS HL-71664 / NHLBI NIH HHS R01 DK054348 / NIDDK NIH HHS DK-52617 / NIDDK NIH HHS R01 HL071664 / NHLBI NIH HHS
Language: English
Date published: 08/2005
Academic Unit: Stead Family Department of Pediatrics; Obstetrics and Gynecology; Internal Medicine
Record Identifier: 9983985903602771

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