Fig. 4

CXCR4 enhances β-catenin nuclear translocation and activates WNT5A/β-catenin pathway in palbociclib-resistant breast cancer. a RT-qPCR showing the mRNA level of WNT5A in parental and palbociclib-resistance MCF-7 and T47D cells. b, c Western blotting showing the protein level of WNT5A and β-catenin in parental and palbociclib-resistance MCF-7 (b) and T47D(c) cells. d–g Palbociclib-resistance MCF-7 and T47D cells were transfected with NC or one of the two CXCR4 siRNAs to knock down the expression of CXCR4. RT-qPCR (d, e) showing the mRNA level of CXCR4 and WNT5A and Western blotting (f, g) showing the protein level of CXCR4, WNT5A and β-catenin. h, i Western blotting showing the protein level of CXCR4, WNT5A and β-catenin in control and CXCR4-overexpressed MCF7-Pa (h) and T47D-Pa (i) cells. j, k The nuclear and cytoplasmic protein was extracted and then was subjected to western blotting analysis, the results showing the levels of nuclear and cytoplasmic β-catenin in parental and palbociclib-resistance MCF-7 (j) and T47D (k) cells. l Palbociclib-resistance MCF-7 cells were transfected with NC or one of the two CXCR4 siRNAs to knock down the expression of CXCR4, western blotting showing the levels of nuclear and cytoplasmic β-catenin in control and CXCR4 knocked down palbociclib-resistance MCF-7 cells. m Immunofluorescence assay showing the expression and the nuclear and cytoplasmic distribution of β-catenin and CXCR4 in parental and palbociclib-resistance MCF-7 cells and CXCR4 knocked down MCF7-palR cells. n Western blotting showing the levels of nuclear and cytoplasmic β-catenin in control and CXCR4-ovexpressed MCF7-Pa cells. For a, d, e, n = 3 biologically independent experiments, the p values were calculated by Student’s t-test for two group comparison and one-way ANOVAs for multiple groups, ***p < 0.001, **p < 0.01. For b, c, f, g, h, i, GAPDH as a loading control. For j–l, n, GAPDH and Lamin B as cytoplasmic or nuclear loading control, respectively. For b, c, f–n, representative images of three biologically independent experiments were shown