Fig. 2: RACK1 promotes proliferation by affecting β-catenin expression in breast cancer cells.

A Coomassie brilliant blue staining of anti-Flag antibody immunoprecipitation products from control and RACK1-flag expressing MDA-231 cells. B The Venn diagram of 110 proteins interacting with RACK1. C Bubble chart showing the KEGG enrichment analysis of RACK1-interacting proteins. D Bubble chart showing the KEGG enrichment analysis of RACK1 co-expression genes in the TCPA database. E Western blotting analysis showed that silencing of RACK1 inhibited β-catenin expression in MDA-231 and BT549 cells. F Western blotting analysis showed that restoration of RACK1 in RACK1-silenced cells rescued the expression of β-catenin in MDA-231 cells. G, H IHC staining of RACK1 and β-catenin in paraffin sections of mouse tumor specimens. Scale bar, 50 μm. I The positive expression rate of RACK1 and β-catenin in mouse tumor specimens. J The nuclear expression rate of β-catenin in mouse tumor specimens. K, L TOPflash assays showed that basal (K) and WNT3A-induced (L) β-catenin-dependent TCF/LEF transcriptional activity was significantly decreased in RACK1-silenced cells compared with control cells. M Western blotting analysis showed that silencing of β-catenin did not affect RACK1 expression in breast cancer cells. N–O Knockdown of β-catenin inhibited the proliferative capacity of breast cancer cells as measured by CCK-8 (N) and colony formation assay (O). P Cell cycle analysis of control and β-catenin-silenced breast cancer cells. Q Western blotting analysis of the expression of cyclin B1, cyclin D1, and cyclin E1 in control and β-catenin-silenced cells. The relative expression of the proteins was quantified by grayscale using Image J software. All data are expressed as the mean ± SD; *p < 0.05. **p < 0.01, ***p < 0.001, ****p < 0.0001 and ns p > 0.05 versus control, N = 3.