Figure 1. mTORC1 is not involved in fragment N-induced cell survival.

HeLa cells were transfected with a GFP-expressing plasmid together with empty pcDNA3 or with pcDNA3 encoding the D157A caspase-resistant form of fragment N. Twenty-four hours later, the cells were incubated 90 minutes with 20 ng/ml rapamycin or with vehicle alone (DMSO). A. The cells were lysed after an additional 24-hour period and the extent of S6K phosphorylation was assessed by Western blot. B-C. Following the rapamycin pre-incubation period, the cells were treated or not with 10 µM cisplatin (panel B) or with 5 ng/ml Fas ligand (panel C) for 24 hours and apoptosis was scored (mean ±95% CI of 5 and 3 independent experiments, respectively, performed in monoplicate). D. HeLa cells were transfected or not with siRNAs directed at Raptor and one day later transfected with empty pcDNA3 or with pcDNA3 encoding fragment N. After an additional 24-, 48-, or 72-hour period, the cells were lysed to assess the levels of Raptor expression. E. Alternatively, one day following the pcDNA3 plasmid transfection, the cells were treated with cisplatin (30 µM) for 24 more hours and apoptosis was then scored (mean ±95% CI of 3 independent experiments performed in monoplicate). Asterisks denote statistically significant differences [one-way ANOVAs followed by pair-wise Dunn (Bonferroni) post hoc t tests].