Fig. 4.

Effect of NHERF-1 expression on dopamine-mediated Na⁺-K⁺-ATPase α1-subunit regulation. Vector-transfected WT and NHERF-1-deficient OK (OKH) cells, or OKH cells transfected with full-length NHERF-1 (NF), PDZ-1 (Z1)-mutated, or PDZ-2 (Z2)-mutated NHERF-1 were treated for 15 min with 1 μM dopamine at 37°C. A: ouabain-sensitive K⁺-pNPPase activity was measured as described previously (18). Each bar represents activity as mean ± SE (μmole pNPP released·mg protein⁻¹·min⁻¹) from four independent experiments. *P < 0.05 by ANOVA followed by Bonferroni's analysis from the respective vehicle-treated cells. B: ouabain-sensitive ⁸⁶Rb uptake was measured as described previously (18). Each bar represents ⁸⁶Rb uptake as means ± SE (nmole ⁸⁶Rb uptake·mg protein⁻¹·10 min⁻¹). *P < 0.05 by ANOVA followed by Bonferroni's analysis from the respective vehicle-treated cells. C: cells were lysed after treatment with dopamine as above. Na⁺-K⁺-ATPase α1-subunit was immunoprecipitated from crude membranes and analyzed by Western blot analysis using phosphoserine antibodies. Bar diagram represents data as A.U. (ratio of phosphoserine band density to Na⁺-K⁺-ATPase α1-subunit) means ± SE from four independent experiments. *P < 0.05 by ANOVA followed by Bonferroni's analysis from the respective vehicle-treated cells.