Figure 7
SREBP1 is essential for cell viability and in vivo tumor growth. (A) U87-GFP-Tet-pLKO-shSREBP1 cells were treated with 1 μg/ml doxycycline or solvent (ethanol) for 48 hours before being placed in medium containing either 10% FCS or 1% LPDS for a further 24 hours. Expression of SREBP1 and SREBP2 was determined. Graphs show mean and range of two independent experiments. (B) Cells were treated as in A and acetate-dependent de novo lipid synthesis was determined. Graph shows mean ± SEM of three independent experiments. (C) Expression of stearoyl-CoA desaturase (SCD) in U87 cells depleted of SREBP1. Graph shows mean and range of two independent experiments. (D) Induction of apoptosis was determined in cells depleted of SREBP1. Cells were treated with 1 μg/ml doxycycline or solvent for 48 hours before being placed in medium containing either 10% FCS or 1% lipoprotein depleted serum (LPDS) for the final 64 hours. Graph shows mean ± SEM of three independent experiments. (E) Expression of CHOP in cells treated as in A. Graph shows mean and range of two independent experiments. (F) Cells were treated with 1 μg/ml doxycycline or solvent for 48 hours before being placed in medium containing either 10% FCS or 1% LPDS for the final 24 hours. Lysates were analyzed for cleaved PARP and PERK and eIF2α phosphorylation. (G) Cells were treated as in D but 10 mM NAC was added prior to placing into lipoprotein-deplete conditions. Graph shows mean ± SEM of three independent experiments. (H) Nude mice (nu/nu, 6 per group) were injected subcutaneously with 5x106 U87-GFP-Tet-pLKO-shSREBP1 cells. Silencing was induced in the treatment group by addition of doxycycline to the food (day 8). Tumor volumes were determined over 30 days. Graph shows mean ± SEM. (I) Weight of tumors at day 30. Graph shows mean ± SEM. (J) Expression of SREBP1 in tumors at day 30. Graph shows mean ± SEM. (K) Histological analysis of tumors (hematoxylin and eosin staining). *P < 0.05; **P < 0.01.