Depletion of Smurf2 accelerates degradation of CNKSR2. (A) Depletion of Smurf2 by siRNA (siSmurf2) leads to decreased CNKSR2 protein levels in MDA-MB-231, MCF-7, SW480, and SCC131 cells. (B) Smurf2 does not affect CNKSR2 transcript level. RNA samples from MDA-MB-231 cells transfected with siSmurf2 were analyzed by qRT-PCR and further by (C) RT-PCR for the indicated transcripts. (D) MDA-MB-231 cells transfected with control siRNA and Smurf2 siRNA were treated with cycloheximide at 100 μg/ml, 48 hours post-transfection. Cells were harvested 0, 1, 2, or 3 h after cycloheximide addition and the cell lysate was subjected to SDS-PAGE followed by immunoblotting with specific antibodies. (E) Plot of CNKSR2 degradation rate was shown in the panel. (F) CNKSR2 depletion does not affect Smurf2 levels.