Figure 2.

c-Abl counteracts YAP-induced phenotype in MCF10A. (a) c-Abl downregulates TEAD target genes. MCF10A cells were infected with either empty vector (pBabe zeo) or YAP, and were then subjected to a second infection with either empty vector (pBabe puro) or active c-Abl. Expression of genes was determined by quantitative RT-PCR and compared with vector control cells. (b) c-Abl inhibits TEAD activation by YAP in a kinase-dependent manner. The designated plasmids were co-transfected with a TEAD-responsive element luciferase reporter plasmid (8 × GTIIC-Luc) into HEK293 cells. Luciferase activity was measured and normalized to co-transfected TK-Renilla. Km, kinase mutant. (c) c-Abl interferes with YAP activity over time. The designated plasmids were co-transfected with a TEAD-responsive element luciferase reporter plasmid (8 × GTIIC-Luc) into HEK293 cells and luciferase activity was recorded over 5 days using real-time bioluminescence detector. (d) c-Abl inhibits YAP-induced EMT. The 2D morphology – representative images of cells grown in monolayer cultures; 3D morphology – cells were cultured on Matrigel. Images were taken on day 4. (e) c-Abl inhibits YAP-induced enhanced migration potential as measured in a wound healing assay. Confluent cells were scratched using 1 ml pipette and grown for 24 h in medium containing 2% serum and no EGF. (f) c-Abl inhibits YAP-induced anchorage-independence. Images were taken and colonies counted 21 days after cells were seeded in soft agar. Colonies >50 μm in diameter were counted as positive for growth. Upper panel: images of colonies; lower panel; quantification of colonies from three independent experiments