Fig. 1.

Forskolin (FSK) dose and time dependently induces UT-A1 ubiquitination. UT-A1 Madin-Darby canine kidney (MDCK) cells were treated with different doses of FSK for 1 h (A) or with 10 μM FSK for different time periods (B). Some cells as indicated in A were pretreated with 20 μM H89 for 1 h. After treatment, cells were lysed with radioimmunoprecipitation assay (RIPA) buffer. Equal amounts of lysates were immunoprecipitated with UT-A1 antibody and Western blotted with anti-ubiquitin (Ub) P4D1. Immunoprecipitation without primary (UT-A1) Ab was set as a negative control. The bands were quantified with NIH ImageJ from 3 different experiments. The ubiquitinated UT-A1 was normalized to the immunoprecipitated UT-A1. The relative density of control (or time point 0) was set as 1. *P < 0.05. **P < 0.01.