Western blot analysis
Proteins were extracted using radio-immunoprecipitation assay buffer in MEF cells. Proteins were separated by 12% sodium dodecyl sulfate-polyacryamide gel electrophoresis and were transferred to a polyvinylidene fluoride membrane (Millipore, Bellerica, MA, USA). Membranes were blocked with 5% non-fat dry milk and were incubated with various antibodies. Antibodies to PC2 (anti-human and anti-mouse) and AQP1 were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Antibodies to β-actin were obtained from Sigma-Aldrich (St. Louis, MO, USA). Anti-Wnt4 was obtained from R&D Systems (Minneapolis, MN, USA). Each membrane was washed with phosphate-buffered saline-Tween, and immunocomplexes were detected by enhanced chemiluminescence (Amersham).