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. 2015 Jun 8;4(6):e156. doi: 10.1038/oncsis.2015.16

Figure 6.

Figure 6

FTY720 induces ERα expression in ERα-negative human and murine breast cancer cells and sensitizes them to tamoxifen. 4T1 (a) and MDA-MB-231 (b) cells were treated with FTY720 (5 μM) or SAHA (1 μM) for 24 h. ERα, PR and ERβ mRNA levels were determined by quantitative real-time PCR (QPCR) and normalized to GAPDH. (c) Proteins in MDA-MB-231 nuclear extracts were analyzed by immunoblotting with the indicated antibodies. LaminA/C was used as a loading control. (d) MDA-MB-231 cells were subjected to chromatin immunoprecipitation (ChIP) analyses with antibodies to H3-ac, H3 or normal rabbit IgG, as indicated. The precipitated DNA was analyzed by real-time PCR with primers amplifying the core promoter sequence of the ERα gene. Relative binding to the promoter is expressed as fold enrichment compared with input. Data are mean±s.d. *P<0.003 compared with vehicle. (e) MDA-MB-231 cells were treated with FTY720 (1 nM) without or with 10 nM E2 for the indicated days and cell proliferation was determined by WST assay. (f, g) MDA-MB-231 cells (f) or 4T1 cells (g) were treated with the indicated concentrations of TAM or FTY720, or with 2.5 μM FTY720 with increasing concentrations of TAM for 48 h and cell proliferation determined. Data are expressed as % of untreated control.