Figure 6

Transducin (β)-like 1 X-linked receptor 1 (TBLR1) activates β-catenin signaling. (A) Western blotting shows the protein levels of cyclin D1, pRb and total Rb in breast cancer cell lines; α-tubulin was used as a loading control. (B) Real-time PCR analysis shows the relative expression of cyclin D1 mRNA in breast cancer cell lines. Expression levels were normalized against glyceraldehyde-3-phosphate dehydrogenase. (C) Indicated cells transfected with TOPflash or FOPflash and Renilla pRL-TK plasmids were subjected to dual-luciferase assays 48 hours after transfection. Reporter activity detected was normalized by Renilla luciferase activity. (D) Real-time PCR analysis indicates an apparent overlap between β-catenin-dependent genes and TBLR1-regulated genes. The pseudocolor represents the intensity scale of TBLR1 versus vector control, or TBLR1 siRNA versus negative control, generated by a log2 transformation. (E) Chromatin immunoprecipitation assays showing the levels of endogenous β-catenin protein bound to cyclin D1 and MYC promoters in TBLR1 breast cancer cells and vector control cells; Error bars represents the mean ± SD of three independent experiments; *P <0.05.