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Synergistic activation of ENaC by three membrane-bound channel-activating serine proteases (mCAP1, mCAP2, and mCAP3) and serum- and glucocorticoid-regulated kinase (Sgk1) in Xenopus Oocytes.

ABSTRACT: Sodium balance is maintained by the precise regulation of the activity of the epithelial sodium channel (ENaC) in the kidney. We have recently reported an extracellular activation of ENaC-mediated sodium transport (I(Na)) by a GPI-anchored serine protease (mouse channel-activating protein, mCAP1) that was isolated from a cortical collecting duct cell line derived from mouse kidney. In the present study, we have identified two additional membrane-bound serine proteases (mCAP2 and mCAP3) that are expressed in the same cell line. We show that each of these proteases is able to increase I(Na) 6-10-fold in the Xenopus oocyte expression system. I(Na) and the number (N) of channels expressed at the cell surface (measured by binding of a FLAG monoclonal I(125)-radioiodinated antibody) were measured in the same oocyte. Using this assay, we show that mCAP1 increases I(Na) 10-fold (P < 0.001) but N remained unchanged (P = 0.9), indicating that mCAP1 regulates ENaC activity by increasing its average open probability of the whole cell (wcP(o)). The serum- and glucocorticoid-regulated kinase (Sgk1) involved in the aldosterone-dependent signaling cascade enhances I(Na) by 2.5-fold (P < 0.001) and N by 1.6-fold (P < 0.001), indicating a dual effect on N and wcP(o). Compared with Sgk1 alone, coexpression of Sgk1 with mCAP1 leads to a ninefold increase in I(Na) (P < 0.001) and 1.3-fold in N (P < 0.02). Similar results were observed for mCAP2 and mCAP3. The synergism between CAPs and Sgk1 on I(Na) was always more than additive, indicating a true potentiation. The synergistic effect of the two activation pathways allows a large dynamic range for ENaC-mediated sodium regulation crucial for a tight control of sodium homeostasis.

SUBMITTER: Vuagniaux G

PROVIDER: S-EPMC2234457 | biostudies-literature | 2002 Aug

REPOSITORIES: biostudies-literature

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Synergistic activation of ENaC by three membrane-bound channel-activating serine proteases (mCAP1, mCAP2, and mCAP3) and serum- and glucocorticoid-regulated kinase (Sgk1) in Xenopus Oocytes.

Vuagniaux Grégoire G Vallet Véronique V Jaeger Nicole Fowler NF Hummler Edith E Rossier Bernard C BC

The Journal of general physiology 20020801 2

Sodium balance is maintained by the precise regulation of the activity of the epithelial sodium channel (ENaC) in the kidney. We have recently reported an extracellular activation of ENaC-mediated sodium transport (I(Na)) by a GPI-anchored serine protease (mouse channel-activating protein, mCAP1) that was isolated from a cortical collecting duct cell line derived from mouse kidney. In the present study, we have identified two additional membrane-bound serine proteases (mCAP2 and mCAP3) that are ...[more]

PMID: 12149280

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Project description:BackgroundPulmonary edema is one of the pathological characteristics of acute respiratory distress syndrome (ARDS). The epithelial sodium channel (ENaC) is thought to be the rate-limiting factor for alveolar fluid clearance (AFC) during pulmonary edema. The peroxisome proliferator-activated receptor γ (PPARγ) agonist rosiglitazone was shown to stimulate ENaC-mediated salt absorption in the kidney. However, its role in the lung remains unclear. Here, we investigated the role of the PPARγ agonist in the lung to find out whether it can regulate AFC during acute lung injury (ALI). We also attempted to elucidate the mechanism for this.MethodsOur ALI model was established through intratracheal instillation of lipopolysaccharide (LPS) in C57BL/6 J mice. The mice were randomly divided into 4 groups of 10. The control group underwent a sham operation and received an equal quantity of saline. The three experimental groups underwent intratracheal instillation of 5 mg/kg LPS, followed by intraperitoneal injection of 4 mg/kg rosiglitazone, 4 mg/kg rosiglitazone plus 1 mg/kg GW9662, or only equal quantity of saline. The histological morphology of the lung, the levels of TNF-α and IL-1β in the bronchoalveolar lavage fluid (BALF), the level of AFC, and the expressions of αENaC and serum and glucocorticoid-induced kinase-1 (SGK1) were determined. Type 2 alveolar (AT II) cells were incubated with rosiglitazone (15 μM) with or without GW9662 (10 μM). The expressions of αENaC and SGK1 were determined 24 h later.ResultsA mouse model of ALI was successfully established. Rosiglitazone significantly ameliorated the lung injury, decreasing the TNF-α and IL-1β levels in the BALF, enhancing AFC, and promoting the expressions of αENaC and SGK1 in ALI mice, which were abolished by the specific PPARγ blocker GW9662. In vitro, rosiglitazone increased the expressions of αENaC and SGK1. This increase was prevented by GW9662.ConclusionsRosiglitazone ameliorated the lung injury and promoted ENaC-mediated AFC via a PPARγ/SGK1-dependent signaling pathway, alleviating pulmonary edema in a mouse model of ALI.

Dataset Information

Synergistic activation of ENaC by three membrane-bound channel-activating serine proteases (mCAP1, mCAP2, and mCAP3) and serum- and glucocorticoid-regulated kinase (Sgk1) in Xenopus Oocytes.

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Synergistic activation of ENaC by three membrane-bound channel-activating serine proteases (mCAP1, mCAP2, and mCAP3) and serum- and glucocorticoid-regulated kinase (Sgk1) in Xenopus Oocytes.

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