Fig. 4

B3GNT5 promoted mammosphere formation and enhances CSC population. A, B Mammosphere formation of BT549 and MDA-MB231 cells with or without B3GNT5 knockout as well as stably B3GNT5-expressing KO cells (A), and representative images are shown (B). Scale bars, 50 μm. (C) Populations of CSCs (CD44^high/CD24^low) were measured by flow cytometry in MDA-MB231 and BT549 cells with or without B3GNT5 knockout as well as stably B3GNT5-expressing KO cells. D Expression of B3GNT5 was analyzed by Western blotting in MCF7 cells with stable empty vector or B3GNT5 expression. E, F Mammosphere formation (E) and population of CSCs (CD44^high/CD24^low) (F) were examined in MCF7 cells with stable empty vector or B3GNT5-WT-Flag cDNA. G Sequential mammosphere formation were measured in MDA-MB231 and BT549 cells with or without B3GNT5 knockout as well as stably B3GNT5-expressing KO cells. P1, P2 and P3 indicated primary, secondary and tertiary passage of mammosphere, respectively. H Limiting dilution sphere forming assay was performed by fluorescence-activated cell sorting in MDA-MB231 KO and BT549 KO cells with B3GNT5-WT-mcherry overexpression. Cells with high (mcherry^high) and low (mcherry^low) B3GNT5 expression were sorted separately. Data are presented as the percentage of wild-type or vector cell lines. *p < 0.05 by Student’s t test. Data are shown as mean ± SD based on three independent experiments