Fig. 3

Overexpression of TMEM120B enhanced stemness of breast cancer cells. (A) GO analysis was performed to detect the biological process significantly correlated with the deletion of TMEM120B in MDA-453 cells. (B) Bioinformatics analysis for the mRNAsi Stemness score of TMEM120B (C) Immunoblotting of Myc-tag, ALDH1, OCT4, NANOG, SOX2, and GAPDH after overexpressing or deleting TMEM120B in SK-BR-3 and MDA-231 cells. Both the first (D, scale bar = 250 μm) and second round of sphere formation assays (E) were performed to examine the effects on stemness of cells after overexpressing or knocking out TMEM120B in SK-BR-3 or MDA-231 cells. (F) Immunoblotting of Myc-tag, OCT4, NANOG, ALDH1, SOX2, and GAPDH in MDA-231 cells in both non-sphere and sphere groups. (G) Flow cytometry assay detected the ratio of ALDH1⁺ cells upon overexpression or deletion of TMEM120B in SK-BR-3 or MDA-231 cells (H) Diluted injection xenograft assays to explore the effects on stemness of breast cancer cells upon depleting TMEM120B in MDA-231 cells in vivo. Quantification data are expressed as mean ± SD of three independent experiments (t-test, two-sided, *P < 0.05, **P < 0.01, ***P < 0.001)