Fig. 3: TCOF1 depletion attenuates stemness of TNBC CSCs.

a MDA-MB-468 and HCC1806 cells expressing tet-on TCOF1 or CTL gRNA were treated with dox (100 ng/ml) for 5 days. Cells were then seeded for first- and second-generation mammosphere-formation assay. Left panel, representative pictures of mammosphere. Right panel, bar graphs depict the mammosphere number. Error bars, mean ± SEM of three independent experiments. *p < 0.05; **p < 0.01; ***p < 0.001. b MDA-MB-468 cells expressing TCOF1 mut or control vector were infected with tet-on TCOF1 or CTL gRNA. Cells were treated with dox (100 ng/ml) for 5 days and then subjected to mammosphere-formation assay. Error bars, mean ± SEM of three independent experiments. *p < 0.05; ***p < 0.001. c Ratio of secondary to primary mammosphere number of HCC1806 and MDA-MB-468 lines. Error bars, mean ± SEM of three independent experiments. *p < 0.05; **p < 0.01. d mRNA level of ALDH1A1 in MDA-MB-468 with or without TCOF1 knockout. Error bars, mean ± SEM of three independent experiments. **p < 0.01. e ALDH activity of cells derived from HCC1806 and MDA-MB-468 spheroids with or without TCOF1 knockout, measured by AldeRed ALDH detection assay. f, g Limiting dilution analysis of tumour-initiating cell frequency of HCC1806 and MDA-MB-468 cells with or without TCOF1 knockout. Tumour incidence was shown as number of tumour formed/number of injected MFP. Tumour-initiating cell frequency was calculated using the ELDA software. Cl confidence interval. p values were calculated by two-sided Student’s t test in a–d.