Figure 4.

The SREBPs support proliferation, growth, and survival in breast cancer cell lines. (a) Effect of SREBP1 and SREBP2 depletion on proliferation in breast cancer cells. MDA-MB-468, MDA-MB-453 and Hs578T cells were transfected with siRNAs targeting SREBP1, SREBP2, or both and were switched to lipid-reduced serum 24 h after the knockdown (t = 0 h). For all proliferation graphs, data are shown as mean ± s.e.m., n=3. * P-value < 0.05 compared to control cells at the final time point. (b) Rescue of human SREBP2 knockdown with mouse SREBP2 expression. MDA-MB-468 cells stably expressing mouse SREBP2 were transfected with siRNA targeting human SREBP2. Cells were lysed for immunoblotting 72 h post-transfection, following 16 h serum starvation. To measure proliferation, cells were cultured in lipid-reduced serum and counted every 24 h. (c) Effects of SREBP2 shRNA on SREBP target expression and proliferation. MDA-MB-468 cells stably expressing four different shRNA sequences targeting SREBP2 were either serum starved for 16 h for immunoblot analysis or cultured in lipid-reduced serum to measure proliferation. (d) Effects of SREBP knockdown on cell size. Cell diameter was measured at 48 h in cells from a, in solution. Color-coded P-values, compared to cells with control siRNAs, correspond to the color-coding in the legend (>1000 cells measured for each). (e) Effect of SREBP knockdown on breast cancer cell viability. Percent cell death was determined by counting Annexin-V and/or propidium iodide positive cells treated as in a by flow cytometry 72 h after siRNA transfection. Data are shown as mean ± s.e.m. relative to cells transfected with non-targeting siRNA, n=2. * P-value < 0.05.