Fig 2: sin-lncRNA rewires metabolic activity in senescent cells(A) Left, representative β-galactosidase images of untreated control cells and control, sin-lncRNA-depleted or DLST-depleted IMR90 cells induced to senescence for 5 days. Scale bars: 50 μm. Right, quantification of the percentage of β-galactosidase-positive cells in each condition normalized to control untreated cells set as 1. The graph represents the mean ± SD (n = 4). Two-tailed Student’s t test, ∗∗p < 0.01.(B) Absorbance levels of crystal violet solution measured at 560 nm in the conditions described in (E). The graph represents the mean ± SD (n = 4). Two-tailed Student’s t test, ∗p < 0.05.(C) Lactate levels measured from the supernatant of IMR90 ER:RAS fibroblasts transfected with the indicated siRNA and induced with 4OHT for 3 or 6 days. The graph represents the mean ± SD (n = 3). Two-tailed Student’s t test, ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001.(D) Left, schematic showing alternative metabolic pathways. Right, RT-qPCR of the corresponding transcripts in sin-lncRNA knockdown cells relative to control (n = 3). The graph represents the mean ± SD (n = 4). Two-tailed Student’s t test, ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001.(E) RT-qPCR of metabolic transcripts in the cells in stable senescent IMR90 ER:RAS cells expressing a doxycycline-inducible wild-type (WT) or NLS mutant (Mut) (R224A/K226E) DLST construct in the absence (−Dox) or presence (+Dox) of 100 ng/mL of doxycycline for 72 h (n = 3).(F) ChIP-qPCR analysis of H3K79 succinylation in senescent IMR90 ER:RAS cells upon 6 days of 4OHT treatment transfected with siRNA control or siRNA against sin-lncRNA at the promoters of different metabolic genes. Regions located at intergenic regions (Neg1 and Neg2) served as negative controls. The graph shows a representative experiment (n = 2).