Fig. 4.

Disruption of tubular E3 ubiquitin-protein ligase. Neuronal precursor cell developmentally downregulated 2 (NEDD4-2) prevents dietary Mg²⁺ restriction-mediated downregulation of Na⁺-Cl⁻ cotransporter (NCC). A: control mice had lower abundance of total NCC (tNCC) and phosphorylated (p)NCC on Mg²⁺-deficient diet (D) compared with normal diet (N), whereas in inducible nephron-specific NEDD4-2 knockout (KO) mice, dietary Mg²⁺ restriction did not lead to lower abundance of NCC (two-way ANOVA with Tukey’s multiple-comparison test). B: abundance of serum and glucocorticoid-induced kinase 1 (SGK1) was comparable between normal diet and Mg²⁺-deficient diet after 3 days of dietary Mg²⁺ manipulation (two-tailed unpaired t-test). C: NCC mRNA did not differ among normal diet or short-term (3 days) and long-term (14 days) dietary Mg²⁺ restriction (one-way ANOVA with Tukey’s multiple-comparison test). For blot quantification, densitometric values were normalized using Coomassie-stained gels. Values are means ± SE; values in parentheses indicate n values. *P < 0.05; **P < 0.01; ***P < 0.0001.