Fig. 2: TeloC dilncRNAs are essential to maintain ALT cells viability.

a Cells were transfected with 10 nM ASO, and growth rate was calculated by Incucyte; data are presented as mean values +/− SEM, n = 3 biologically independent experiments; two-way ANOVA, for HeLa control vs antiteloC 24 h *p = 0.0334, 28 h **p = 0.0063, 32 h *p = 0.0372; for HeLa antiteloG vs antiteloC 24 h *p = 0.0133, 28 h ***p = 0.0009, 32 h **p = 0.0092; for U2OS control vs antiteloC from 24 h to 72 h ****p < 0.0001; for U2OS antiteloG vs antiteloC from 24 h to 32 h ****p < 0.000, 36 h ***p = 0.003, 40 h ***p = 0.0002, 44 h ***p = 0.0007, 48 h ***p = 0.0004, from 52 h to 64 h ***p = 0.0002, 68 h ***p = 0.0003, 72 h ***p = 0.0005. b Cells were transfected with 20 nM ASO and growth rate was calculated three days later by resazurin; data are presented as mean values +/− SEM, n = 3 biologically independent experiments for HeLa (df = 8, F = 2.865) and G292 (df = 8, F = 8.454), n = 4 biologically independent experiments for BJhTERT (df = 11, F = 0.5318), U2OS (df = 11, F = 14.50) and SAOS2 (df = 11, F = 7.493); one-way ANOVA, ns = non-significant, for U2OS control vs antiteloC **p = 0.0024 antiteloG vs antiteloC **p = 0.0041; for G292 control vs antiteloC *p = 0.0224 antiteloG vs antiteloC *p = 0.0356; for SAOS2 control vs antiteloC **p = 0.0206 antiteloG vs antiteloC *p = 0.0212;. c Cells were treated with a range of concentrations of ASO antiteloC without transfection reagent and GR₅₀ was calculated three days later with RealTime-Glo. When not determined, GR₅₀ was underestimated to 100 μM (the highest concentration used in this experiment); data are presented as mean values +/− SEM, n = 3 biologically independent experiments for SI-14 and SI-24 (df = 4, t = 14.37), for JFCF-6/T1C and 1D (df = 4, t = 6.267), and for JFCF-6/T.1J6B and 1.3 C (df = 4, t = 3.349) and n = 4 biologically independent experiments for 6C3 and 8G12 (df = 6, t = 2.382); two-tailed unpaired t-test, for SI-14 vs SI-24 ***p = 0.0001; for JFCF-6/T1C and 1D **p = 0.0033; for JFCF-6/T.1J6B and 1.3 C *p = 0.0286 d Cells treated as in (a), and apoptotic cells counted by Incucyte three days after transfection; values reported as mean values +/- SEM, n = 5 biologically independent experiments; one-way ANOVA, df = 14, F = 34.71, HeLa vs U2OS ***p = 0.0004, BJhTERT vs U2OS **p = 0.0048. e Cells were transfected with 20 nM ASO and two days later analyzed by western blot; n = 2, one representative image shown. f U2OS cells were transfected with 20 nM ASO and fixed two days later for flow cytometry analysis; data are presented as mean values +/− SEM, n = 3 biologically independent experiments; one-way ANOVA, df = 8, F = 11.19; ns = non-significant, control vs antiteloC *p = 0.0135, antiteloG vs antiteloC *p = 0.0169. g Cells were treated with 20 μM of ASO antiteloC without transfection reagent and three days later analyzed by western blot. h The ratios between cleaved caspase 3 over actin and cleaved PARP1 over total PARP1 from the experiment shown in (g) were measured; data are presented as mean values +/− SEM, n = 3 non-ALT and n = 3 ALT cell lines; two-tailed paired t-test, df = 2, t(Ccaspase-3) = 4.458, t(PARP1) = 5.811,CCaspase3 *p = 0.0468, PARP1 *p = 0.0284. i U2OS cells were transfected with the indicated antiteloC ASO at a range of concentrations and live cells were counted by Incucyte three days after transfection; data are presented as mean values +/− SEM, n = 3 biologically independent experiments. Source data are provided as a Source data file.