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Respiratory distress and perinatal lethality in Nedd4-2-deficient mice.

Boase NA, Rychkov GY, Townley SL, Dinudom A, Candi E, Voss AK, Tsoutsman T, Semsarian C, Melino G, Koentgen F, Cook DI, Kumar S - Nat Commun (2011)

Bottom Line: This increased ENaC activity is the likely reason for premature fetal lung fluid clearance in Nedd4-2(-/-) animals, resulting in a failure to inflate lungs and perinatal lethality.A small percentage of Nedd4-2(-/-) animals survive up to 22 days, and these animals also show increased ENaC expression and develop lethal sterile inflammation of the lung.Thus, we provide critical in vivo evidence that Nedd4-2 is essential for correct regulation of ENaC expression, fetal and postnatal lung function and animal survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Haematology, Centre for Cancer Biology, SA Pathology, PO Box 14, Rundle Mall, Adelaide, South Australia 5000, Australia.

ABSTRACT
The epithelial sodium channel (ENaC) is essential for sodium homoeostasis in many epithelia. ENaC activity is required for lung fluid clearance in newborn animals and for maintenance of blood volume and blood pressure in adults. In vitro studies show that the ubiquitin ligase Nedd4-2 ubiquitinates ENaC to regulate its cell surface expression. Here we show that knockout of Nedd4-2 in mice leads to increased ENaC expression and activity in embryonic lung. This increased ENaC activity is the likely reason for premature fetal lung fluid clearance in Nedd4-2(-/-) animals, resulting in a failure to inflate lungs and perinatal lethality. A small percentage of Nedd4-2(-/-) animals survive up to 22 days, and these animals also show increased ENaC expression and develop lethal sterile inflammation of the lung. Thus, we provide critical in vivo evidence that Nedd4-2 is essential for correct regulation of ENaC expression, fetal and postnatal lung function and animal survival.

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Increased ENaC expression in Nedd4-2−/− mice.(a) α and βENaC expression in E18.5 lung and kidney. (b) Quantification of ENaC expression from (a) normalized to β-actin and relative to wild type. (c) βENaC expression (green) in E18.5 lung and kidney. Scale bars, 50 μm.
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f3: Increased ENaC expression in Nedd4-2−/− mice.(a) α and βENaC expression in E18.5 lung and kidney. (b) Quantification of ENaC expression from (a) normalized to β-actin and relative to wild type. (c) βENaC expression (green) in E18.5 lung and kidney. Scale bars, 50 μm.

Mentions: At the molecular level, absence of Nedd4-2 is expected to result in decreased ENaC ubiquitination, and therefore less internalization and degradation. To determine whether ENaC expression was altered in Nedd4-2−/− mice, immunoblot analysis was performed using antibodies generated against α or β subunits of ENaC (specificity of antibodies shown in Supplementary Fig. S3). Nedd4-2−/− E18.5 lung and kidney tissue lysates displayed an increase in α and βENaC expression compared with wild type, with heterozygous mice showing intermediate levels of ENaC expression (Fig. 3a,b). Immunostaining of E18.5 lung sections demonstrated dramatically higher βENaC expression on alveolar luminal surfaces in the Nedd4-2−/− lung compared with wild type (Fig. 3c). E18.5 kidneys from Nedd4-2−/− animals also showed a clear increase of βENaC in the membranes of collecting ducts compared with wild-type littermates (Fig. 3c). Together, these results are consistent with a role for Nedd4-2 in ENaC regulation.


Respiratory distress and perinatal lethality in Nedd4-2-deficient mice.

Boase NA, Rychkov GY, Townley SL, Dinudom A, Candi E, Voss AK, Tsoutsman T, Semsarian C, Melino G, Koentgen F, Cook DI, Kumar S - Nat Commun (2011)

Increased ENaC expression in Nedd4-2−/− mice.(a) α and βENaC expression in E18.5 lung and kidney. (b) Quantification of ENaC expression from (a) normalized to β-actin and relative to wild type. (c) βENaC expression (green) in E18.5 lung and kidney. Scale bars, 50 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3104547&req=5

f3: Increased ENaC expression in Nedd4-2−/− mice.(a) α and βENaC expression in E18.5 lung and kidney. (b) Quantification of ENaC expression from (a) normalized to β-actin and relative to wild type. (c) βENaC expression (green) in E18.5 lung and kidney. Scale bars, 50 μm.
Mentions: At the molecular level, absence of Nedd4-2 is expected to result in decreased ENaC ubiquitination, and therefore less internalization and degradation. To determine whether ENaC expression was altered in Nedd4-2−/− mice, immunoblot analysis was performed using antibodies generated against α or β subunits of ENaC (specificity of antibodies shown in Supplementary Fig. S3). Nedd4-2−/− E18.5 lung and kidney tissue lysates displayed an increase in α and βENaC expression compared with wild type, with heterozygous mice showing intermediate levels of ENaC expression (Fig. 3a,b). Immunostaining of E18.5 lung sections demonstrated dramatically higher βENaC expression on alveolar luminal surfaces in the Nedd4-2−/− lung compared with wild type (Fig. 3c). E18.5 kidneys from Nedd4-2−/− animals also showed a clear increase of βENaC in the membranes of collecting ducts compared with wild-type littermates (Fig. 3c). Together, these results are consistent with a role for Nedd4-2 in ENaC regulation.

Bottom Line: This increased ENaC activity is the likely reason for premature fetal lung fluid clearance in Nedd4-2(-/-) animals, resulting in a failure to inflate lungs and perinatal lethality.A small percentage of Nedd4-2(-/-) animals survive up to 22 days, and these animals also show increased ENaC expression and develop lethal sterile inflammation of the lung.Thus, we provide critical in vivo evidence that Nedd4-2 is essential for correct regulation of ENaC expression, fetal and postnatal lung function and animal survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Haematology, Centre for Cancer Biology, SA Pathology, PO Box 14, Rundle Mall, Adelaide, South Australia 5000, Australia.

ABSTRACT
The epithelial sodium channel (ENaC) is essential for sodium homoeostasis in many epithelia. ENaC activity is required for lung fluid clearance in newborn animals and for maintenance of blood volume and blood pressure in adults. In vitro studies show that the ubiquitin ligase Nedd4-2 ubiquitinates ENaC to regulate its cell surface expression. Here we show that knockout of Nedd4-2 in mice leads to increased ENaC expression and activity in embryonic lung. This increased ENaC activity is the likely reason for premature fetal lung fluid clearance in Nedd4-2(-/-) animals, resulting in a failure to inflate lungs and perinatal lethality. A small percentage of Nedd4-2(-/-) animals survive up to 22 days, and these animals also show increased ENaC expression and develop lethal sterile inflammation of the lung. Thus, we provide critical in vivo evidence that Nedd4-2 is essential for correct regulation of ENaC expression, fetal and postnatal lung function and animal survival.

Show MeSH
Related in: MedlinePlus