Nedd4-2 Modulates Renal Na+-Cl− Cotransporter via the Aldosterone-SGK1-Nedd4-2 Pathway

Abstract

Regulation of renal Naϩ transport is essential for controlling blood pressure, as well as Na ϩ and K ϩ homeostasis. Aldosterone stimulates Na ϩ reabsorption by the Na ϩ -Cl Ϫ cotransporter (NCC) in the distal convoluted tubule (DCT) and by the epithelial Na ϩ channel (ENaC) in the late DCT, connecting tubule, and collecting duct. Aldosterone increases ENaC expression by inhibiting the channel's ubiquitylation and degradation; aldosterone promotes serum-glucocorticoid-regulated kinase SGK1-mediated phosphorylation of the ubiquitin-protein ligase Nedd4-2 on serine 328, which prevents the Nedd4-2/ENaC interaction. It is important to note that aldosterone increases NCC protein expression by an unknown post-translational mechanism. Here, we present evidence that Nedd4-2 coimmunoprecipitated with NCC and stimulated NCC ubiquitylation at the surface of transfected HEK293 cells. In Xenopus laevis oocytes, coexpression of NCC with wild-type Nedd4-2, but not its catalytically inactive mutant, strongly decreased NCC activity and surface expression. SGK1 prevented this inhibition in a kinase-dependent manner. Furthermore, deficiency of Nedd4-2 in the renal tubules of mice and in cultured mDCT 15 cells upregulated NCC. In contrast to ENaC, Nedd4-2-mediated inhibition of NCC did not require the PY-like motif of NCC. Moreover, the mutation of Nedd4-2 at either serine 328 or 222 did not affect SGK1 action, and mutation at both sites enhanced Nedd4-2 activity and abolished SGK1-dependent inhibition. Taken together, these results suggest that aldosterone modulates NCC protein expression via a pathway involving SGK1 and Nedd4-2 and provides an explanation for the well-known aldosterone-induced increase in NCC protein expression.

The Thiazide-sensitive NaCl Cotransporter Is Targeted for Chaperone-dependent Endoplasmic Reticulum-associated Degradation

“…S6, right panel). These findings are compatible with previous reports by other groups that have employed HEK293 cells as an expression system to study NCC expression and regulation (50,51). These results indicate that when expressed in mammalian cells, NCC predominantly exists in its core glycosylated state, suggesting that the cotransporter undergoes extensive biosynthetic processing in the ER and is strongly sensitive to chaperone-dependent ERAD.…”

Hypertrophy in the Distal Convoluted Tubule of an 11β-Hydroxysteroid Dehydrogenase Type 2 Knockout Model

“…Dysfunction of the NaCl cotransporter (NCC) in the DCT perturbs BP not only in the extreme phenotypes of patients with Gitelman's 1 and Gordon's 2 syndromes but also in healthy individuals, 3 and the inhibition of NCC by thiazide diuretics has a potent antihypertensive effect. 4 In the short term, NCC activity is regulated at the molecular level; neurohormonal inputs converge on intracellular signaling networks (WNK-SPAK/OSR1 5 and SGK1/Nedd4-2 pathways 6 ) that shuttle NCC to and from the plasma membrane 7 and induce posttranslational protein modifications that modify transport function (phosphorylation 8 and ubiquitylation 6 ). Sustained physiologic perturbations promote structural remodeling of DCTepithelium.…”

“…42,43 It is pertinent that NCC is regulated by the SGK1/Nedd4-2 pathway in DCT cells. 6 Although this is a classic target of MR signaling, this pathway may also be activated by glucocorticoids.…”

Protein Phosphatase 1 Inhibitor-1 Deficiency Reduces Phosphorylation of Renal NaCl Cotransporter and Causes Arterial Hypotension

“…WNK1 and WNK4 are serine/ threonine kinases that interact in a complex cascade with the STE20-related proline-alanine-rich kinase (SPAK) to regulate NCC phosphorylation (e.g., Thr44, Thr53, Thr58, and Ser71 in mouse NCC) and finally, activity. 8,[10][11][12] In addition to the WNK-SPAK pathway, several other proteins were identified to control NCC, including parvalbumin (PV), 13 serum and glucocorticoid-inducible kinase Sgk1, ubiquitin ligase Nedd4-2, [14][15][16] kelch-like 3 (KLHL3), 17,18 cullin 3, 17,18 and protein phosphatase (PP) 4. 19 Remarkably, several of the above listed regulatory proteins are highly abundant in the DCT (e.g., WNK4, KS-WNK1, SPAK, PV, and KLHL3).…”