FIGURE 4.

Depletion of Cdh1 attenuates TGF-β-induced KLF4 degradation. A, APC activity is enhanced in response to TGF-β signaling. Mv1Lu cells were treated with TGF-β and collected at the indicated time points. Endogenous APC was then purified by immunoprecipitation using antibody against Cdc27. Iodine 125-radiolabeled cyclin B recombinant protein was used as putative substrate for APC in vitro ubiquitylation assay. Upon stimulation with TGF-β, APC activity was profoundly enhanced as reflected by increased formation of cyclin B polyubiquitin conjugates. B, Cdh1 knockdown by RNAi in Mv1Lu cells. Mv1Lu cells were infected with Cdh1 shRNA expressing retrovirus and the stable cell line was generated. The knockdown effect of Cdh1 shRNA was evaluated by immunoblotting. The Cdh1 expression was normalized to the loading control β-actin. C, quantification of B. D, TGF-β-induced KLF4 degradation is attenuated in Cdh1-depleted Mv1Lu cells. Both Cdh1-depleted and control Mv1Lu cells were treated by TGF-β and harvested at different time points as indicated. KLF4 expression was measured by immunoblotting using antibody against KLF4. E, summary of D.