Fig. 2
From: α-catenin interaction with YAP/FoxM1/TEAD-induced CEP55 supports liver cancer cell migration
Functional relevance of α‐catenin in hepatocarcinogenesis. A Western blot analysis detecting α‐catenin in human liver cancer cell line lysates (n = 8) and the immortalized human hepatocyte cell line HHT4. B Immunofluorescence images of HLF, HepG2, and Hep3B cells stained for α‐catenin. Nuclei are visualized by DAPI. Scale bar: 10 µm. C Representative Western immunoblot and qPCR of α‐catenin in HLF cells after transfection of two independent α-catenin-specific siRNAs (#1, #2). Samples were isolated 72 h after transfection. D Cell viability assay of HLF cells after siRNA-mediated silencing of α‐catenin at indicated time points. E Colony formation assay using HLF cells after siRNA-mediated α‐catenin knockdown followed by the quantification of relative colony formation (software: ImageJ). Colony density was measured 2 weeks after seeding cells. F Proliferation of HLF cells was measured using a BrdU ELISA. Incorporation of the base thymidine analog was detected 96 h after transfection of α‐catenin-specific siRNAs. G HLF cell apoptosis was detected utilizing a CellTox™ Green reagent 48 h after inhibition of α‐catenin. H Lateral migration of HLF cells was detected at indicated time points using a ‘scratch’ assay after siRNA-mediated α‐catenin inhibition. Cells were pretreated with mitomycin-C to prevent cell mitosis. Relative lateral migration was quantified using ImageJ. I Hydrodynamic gene delivery of expression vectors in FVB/N mice. Exemplary pictures of livers for all injected gene combinations are shown (myr-AKT, α‐catenin, and myr-AKT/α‐catenin). The gene combinations myr-AKT/YAPS127A and YAPS127/β-catenin lead to the formation of multiple tumors within 12 and 8.5 weeks after injection (positive controls). Representative H&E stains with indicated non-tumor (N) and tumor (T) areas are shown. The bar graph illustrates the number of animals with tumor formation. Scale bar: 60 µm. For all RNAi experiments, scramble (scr.) siRNA-transfected cells served as controls. All results were normalized to respective controls. Statistical test: Mann–Whitney U test, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001