Salt-sensitive hypertension and cardiac hypertrophy in mice deficient in the ubiquitin ligase Nedd4-2

Abstract

. Salt-sensitive hypertension and cardiac hypertrophy in mice deficient in the ubiquitin ligase Nedd4-2. Am J Physiol Renal Physiol 295: F462-F470, 2008. First published June 4, 2008 doi:10.1152 doi:10. /ajprenal.90300.2008 has been proposed to play a critical role in regulating epithelial Na ϩ channel (ENaC) activity. Biochemical and overexpression experiments suggest that Nedd4-2 binds to the PY motifs of ENaC subunits via its WW domains, ubiquitinates them, and decreases their expression on the apical membrane. Phosphorylation of Nedd4-2 (for example by Sgk1) may regulate its binding to ENaC, and thus ENaC ubiquitination. These results suggest that the interaction between Nedd4-2 and ENaC may play a crucial role in Na ϩ homeostasis and blood pressure (BP) regulation. To test these predictions in vivo, we generated Nedd4-2 null mice. The knockout mice had higher BP on a normal diet and a further increase in BP when on a high-salt diet. The hypertension was probably mediated by ENaC overactivity because 1) Nedd4-2 null mice had higher expression levels of all three ENaC subunits in kidney, but not of other Na ϩ transporters; 2) the downregulation of ENaC function in colon was impaired; and 3) NaClsensitive hypertension was substantially reduced in the presence of amiloride, a specific inhibitor of ENaC. Nedd4-2 null mice on a chronic high-salt diet showed cardiac hypertrophy and markedly depressed cardiac function. Overall, our results demonstrate that in vivo Nedd4-2 is a critical regulator of ENaC activity and BP. The absence of this gene is sufficient to produce salt-sensitive hypertension. This model provides an opportunity to further investigate mechanisms and consequences of this common disorder. ion channels; kidney; sodium channels HYPERTENSION AFFECTS MORE than 25% of the adult population in most Westernized countries and is a major cause of coronary artery disease and cerebrovascular disease (9, 10). Despite its prevalence, the etiology of more than 90% of hypertension cases is unknown. Numerous studies revealed that interactions between genetic and environmental factors, especially the generous intake of dietary salt, play a critical role in its pathogenesis and a large body of evidence implicates the inappropriate retention of Na(Cl) by the kidney in the pathophysiology of hypertension (25). One of the most important molecular determinants of Na ϩ excretion is the activity of the epithelial Na ϩ channel (ENaC), a highly regulated, threesubunit ion channel complex that is active in the distal nephron (15,29). The activity of ENaC is regulated by many first and second messengers, the most widely studied of which is aldosterone (13, 22). The importance of ENaC in Na ϩ homeostasis and hypertension is underscored by the discovery that patients with Liddle syndrome, an autosomal dominant form of hypertension, have gain-of-function mutations in ENaC (28).The mutations in the ␤-and ␥-ENaC subunits leading to Liddle syndrome have focused attention on the specific regions of the COOH termini whose modificati...

Deletion of the ubiquitin ligase Nedd4L in lung epithelia causes cystic fibrosis-like disease

“…Because their genetic background was similar to ours (although not identical), it is possible that those mice contain a shorter splice isoform of Nedd4L [suggested from a Nedd4L immunoblot (28)] that may help explain the milder phenotype. Another possibility is that because these mice lacked Nedd4L throughout embryonic development, other Nedd4 family E3 ligase(s) might have compensated for the loss of Nedd4L in the lung.…”

“…Knockout of Nedd4L in Lung Epithelia Leads to Increased ENaC Expression and Function. Because Nedd4L is known to inhibit ENaC when expressed in tissue culture cells or in Xenopus oocytes (11,14,15,28), we tested using several approaches, whether loss of Nedd4L in type II and Clara cells leads to increased ENaC expression and function in vivo in the lung. First, we assessed levels of ENaC protein expression in lungs using immunostaining of lung sections and isolated type II cells under nonpermeablilizing condition (biochemical analysis using immunoblotting was not feasible due to insufficient number of type II cells that can be isolated from the mice).…”

Renal tubular NEDD4-2 deficiency causes NCC-mediated salt-dependent hypertension

“…Indeed, these mice displayed impaired Na + excretion and hypertension, apparently mediated by ENaC overactivity (25). No effect on NCC was observed.…”

“…Generation of constitutive Nedd4L KO mice, by removing exons 6 to 8 of the Nedd4L gene (Nedd4L-Δ6-8 KO mice), demonstrated the importance of NEDD4-2 in the control of ENaC activity and blood pressure (25). Indeed, these mice displayed impaired Na + excretion and hypertension, apparently mediated by ENaC overactivity (25).…”

Epithelial Sodium Channel

“…A recent transgenic mouse model harboring a Nedd4-2 gene deletion was shown to develop a severe form of salt-sensitive hypertension, suggesting that Nedd4-2 is a limiting factor in the aldosterone-dependent signaling cascade. 52 Fava et al 53 reported that 24-hour ambulatory blood pressure monitoring was linked to chromosome 18q21-22, and genetic variation of Nedd4-2 associates with crosssectional and longitudinal blood pressure in Swedes. In a linkage study of 16 markers (including 2 SNP markers located within the Nedd4-2 gene) on chromosome 18 between 70 and 104 cM and ambulatory blood pressure, in 118 families, the strongest evidence of linkage was found for 24-hour and daytime systolic ambulatory blood pressure monitoring at the Nedd4-2 locus (82.25 cM).…”