Fig. 5.

Myc is necessary and sufficient for estrogen-independent cell proliferation and antiestrogen resistance, but does not affect invasiveness. A Western blot showing Myc protein levels after treatment of T47D, TYS and TDG cells with 100 nM non-coding (NC) or Myc SMARTpool siRNA for 24h, followed by treatment with vehicle or 1 nM E₂ for 24h. B Proliferation of T47D, TYS and TDG cells treated with 100 nM NC or Myc SMARTpool siRNA, followed by treatment with vehicle or 1 nM E₂ for 96 h (mean ± s.e.m., n=6). C Western blot showing Myc levels following treatment of cells with vehicle or 1 nM E₂ with, or without, 1 µM JQ1 for 8h. D Proliferation of T47D, TYS and TDG cells treated with vehicle, E₂, or E₂ plus 0.5 or 1 µM JQ1 for 96 h (mean ± s.e.m., n=6). E Western blot showing ERα, Myc and Cyclin E levels in T47D, TYS and TDG cells treated with Myc-lentivirus or control luciferase-lentivirus, followed by the indicated concentrations of E₂, OHT and ICI for 24h. F Proliferation of T47D, TYS and TDG cells, after transduction with Myc or control lentivirus containing medium for 24h, followed by treatment with the indicated concentrations of E₂, OHT and ICI for 4 days. (mean ± s.e.m., n=6). B,D,F ‘•’ denotes cell number at day 0. C,E Myc protein levels were quantitated using a PhosphorImager and ImageQuant. B,D * indicates a significant difference among groups using one-way ANOVA followed by Tukey’s post hoc test. *P<0.05, **P<0.005, ***P<0.001. ns, not significant. F Different letters indicate a significant difference among groups (P<0.05) using one-way ANOVA followed by Tukey’s post hoc test. G,H IDR assay of T47D cells transduced with Myc-lentivirus or luciferase-lentivirus with collagen- G or matrigel-coated H membrane and chamber (mean ± s.e.m., n=5). Cells were transduced with virus and after 1 day total invaded cells and cells that invaded, then dissociated and rebound (IDR) were measured using their luciferase activity and a standard curve for each cell line of luciferase activity versus cell number.