Fig. 6

EGF and EGFR regulated the AJAP1-mediated β-catenin nuclear translocation. a-b The changes of AJAP1 and β-catenin were observed through immunofluorescence assay of both cells with EGF stimulation (100 ng/ml). (magnification, × 400) c Statistical analysis of the membrane and nuclear expressions of β-catenin with EGF treatment by Image J. d protein changes of AJAP1 and β-catenin expression in T47D and MDA-MB-231 cells under EGF stimulation e Immunoprecipitation assays were conducted to evaluate the interaction between AJAP1 and β-catenin under EGF treatment. f Relative luciferase activities of TOP/FOP dual-luciferase reporter performed to observe the effects of EGF (100 ng/ml) on both cell lines (left) qRT-PCR was performed to assess the effects of EGF stimulation on the β-catenin downstream genes C-myc and CyclinD1 in the experimental groups. g Western blot and qRT-PCR were conducted to examine the AJAP1 expression in EGFR-depleted without EGF stimulating cells (first and second panel). The results showed that EGFR-depletion in AJAP1 knocked down cells promote the β-catenin transfer to the nucleus and its transcription activity (third and fourth panels) and increased the downstream gene levels (third and last panels). h the results of tumor growth, H&E and immunohistochemistry staining of C-myc,CyclinD1, and β-catenin nuclear expression in vivo assays (magnification, × 400). Data were expressed as mean ± SD. **p < 0.01, ***p < 0.001 i A mechanism model showed that the β-Catenin nuclear localization positively fed back on the EGF/EGFR attenuated AJAP1 expression in breast cancer to promote breast cancer progression and metastasis