Fig. 5. ZEB2 is a direct transcriptional regulatory target of PAX6.
a Heatmap illustrating the expression of nine representative EMT-related genes in four groups of A549 cells. b RT-qPCR analysis of the mRNA expression of nine potentially differentiated genes based on the Human Tumor Metastasis RT2 Profiler PCR Array in A549 cells. c The expression levels of three representative genes were compared by western blotting. d A ChIP assay was performed using an antibody against PAX6 or control IgG in A549 cells. Immunoprecipitated DNA was analyzed by PCR followed by agarose gel electrophoresis. e Representative results of IHC staining (scale bar, 100 μm) and the relationship between PAX6 and ZEB2 protein levels in human lung cancer samples. f After ZEB2 knockdown (si-ZEB2), mRNA levels of ZEB2 and PAX6 were evaluated by RT-qPCR. g After ZEB2 knockdown, ZEB2, PAX6 and EMT markers were detected by western blot. h Representative images and i quantitative analysis of cell migration based on wound-healing assays (scale bar, 500μm). j, k Representative images (left panels) and quantitative analysis (right panels) of j cell migration and k invasion by transwell assay (scale bar, 100μm). l Analysis of EMT markers by IF staining (scale bar, 50μm). **P < 0.01, ***P < 0.001; two-tailed Student’s t-test