Fig. 2

Knocking out TOP1MT restrains cell proliferation and sensitizes cells to glucose starvation. a Representative Ki67 immunofluorescence staining of WT and TOP1MT-KO xenograft tumors. Scale bar, 50 μm. b, c Quantification of the fraction of Ki67-positive cells (b) and nuclei count per field (c) measured by ZEN software (6 images per animal, 5 animals per genotype). d Heat map showing significant changes in gene expression profiles analyzed with the nCounter PanCancer Progression panel in four TOP1MT-deficient vs. four WT tumors (89 genes, p < 0.05). e Transcript levels of selected genes of the PI3K/AKT pathway determined by RT-qPCR (n = 4, each performed in triplicates). f Kinetics of cell growth under standard culture conditions and under glucose withdrawal (1 g L⁻¹ glucose, dashed lines) (four independent experiments performed in quadruplets). g Growth of WT and TOP1MT-KO multicellular tumor spheroids (MCTS) formed by 10,000 HCT116 cells in six independent experiments, each performed in quintuplets. Day 0 corresponds to 48 h after cell seeding (spheroid maturation). Dashed and solid lines represent growth in the absence and presence of glucose, respectively. Data represent the means ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001, Student’s t-test