FIGURE 4:

YAP activity is required for GFP-CD–induced increase in proliferation. (A) MDCK cells expressing GFP or GFP-CD were infected with YAP shRNA lentivirus. Lysates were analyzed by Western blot for expression of YAP. YAP protein levels are decreased >90% in both the control and GFP-CD–expressing cells. (B) Changes in proliferation were determined by PHH3 labeling. Proliferation of GFP-CD cells is significantly decreased with YAP knockdown. Error bars, SEM; ***p < 0.005. YAP knockdown in control cells also resulted in decreased proliferation (0.74-fold of that of parent lines). (C) MDCK cells expressing GFP-CD were transfected with p130AMOT. Overexpression of p130-AMOT complements the GFP-CD increased-proliferation phenotype. Error bars, SEM; *p < 0.05. (D) Endogenous EDTB was knocked down in MDCK cells using two independent shRNAs, individually or together. Lysates were analyzed by Western blot to verify the reduction in EDTB expression. (E) Proliferation is decreased when EDTB expression is reduced. Error bars, SEM; *p < 0.05. (F) Pearson r of AMOTL2:YAP colocalization in EDTB knockdown cells. Knockdown of endotubin results in a significant increase in AMOTL2:YAP colocalization. Four to six images were quantified for each condition. Error bars, SEM; ***p < 0.005.