Fig. 1.

nSREBP2 overexpression induces lytic cell death. A: time curve of 0.3 μg/ml doxycycline (dox) treatment in HeLa-pTet-nSREBP2 cells. Cells were treated with dox for the indicated time and analyzed by crystal violet staining. B and C: quantification of cell viability of tet-on-nSREBP2 cells. Cells were treated with 0.3 μg/ml dox for the indicated time and analyzed by cell counting kit-8 (B) or LDH test (C). Data are presented as means ± SD. Statistical analyses, unpaired two-tailed Student’s t test. D and E, HeLa-pTet-nSREBP2 cells were treated with 0/0.3 μg/ml dox plus control, 25 mM D-mannitol or 10 mM glycine for 30 h, LDH (D) and cell viability (E) were tested. Data are presented as means ± SD. Statistical analyses, unpaired two-tailed Student’s t test. F, Time-lapse confocal images of HeLa-pTet-nSREBP2-5×MYC cells were taken at the indicated time points after 0.3 μg/ml dox induction for 36 h. Real time videos are included in supplemental Video S1. Scale bar, 25 μm G, Annexin V/PI flow cytometric analysis of dox-induced cell death phenotype. H, Two clones of HeLa-pTet-nSREBP1a/1c cells and HeLa-pTet-nSREBP2 cells were treated with or without 0.3 μg/ml dox for 36 h. Cell viability was analyzed by crystal violet staining. I, Time-lapse confocal images of HeLa-pTet-nSREBP1a/1c cells. HeLa-pTet-on-nSREBP1a/1c cells were induced by 0.3 μg/ml dox for 24 h, preloaded with 10 μg/ml PI for 15 min, then imaged at 40× magnification at 5 min intervals for 12 h. Real time videos are included in supplemental Videos S2 and S3. Scale bar, 25 μm. ∗∗ P < 0.01, ∗∗ P < 0.01, ∗∗∗∗ P < 0.0001, ns, not significant.