The effect of PI3-kinase and protein synthesis inhibition on the induction of KS-WNK1 by aldosterone. M1-MR+ clone 1 cells were grown in steroid-free medium for 2 days and then preincubated for 30 min with vehicle CHX (5 μg/ml) or anisomycin (aniso; 10 μM). Aldosterone (1 nM) was given (or not) at 30 min, and cells were incubated for an additional 4 h before RNA was prepared. KS-WNK1 and β-actin mRNA levels were determined by RT-PCR. *, P < 0.05, **, P < 0.01, ***, P < 0.001 vs. control, two-tailed unpaired t test. Comparisons were also made between values for each treatment obtained with and without aldosterone; P < 0.05 for vehicle vs. aldosterone; P < 0.05 for LY vs. LY + aldosterone; P = 0.07 for CHX vs. CHX + aldosterone; P < 0.05 for anisomycin vs. anisomycin + aldosterone (using one-tailed unpaired t tests). The values of KS-WNK1 mRNA levels in the presence aldosterone alone were not significantly different from those in cells incubated with aldosterone and LY or CHX or anisomycin (using two-tailed unpaired t tests). LY, LY294002.