Fig. 3

YTHDF1 is required for maintaining cancer stem cell properties of GBM cell line. a Sphere- formation assay performed on DBTRG-05MG cells transduced with control (pLKO.1) and two YTHDF1-targeting shRNA constructs (shYTHDF#1 and shYTHDF1#2). Top panel: the first generation spheres formed on day 7. Bottom panel: quantification of sphere area. The values represent the means ± SD error bars from three independent experiments. **P < 0.01 (Student’s t-test) vs. pLKO.1. b Western blotting analysis of expression of YTHDF1 and stemness markers, CD133, NANOG, SOX2, OCT4 and REX1, in sphere cells. GAPDH used as a loading control. c Transwell migration assay performed on DBTRG-05MG cells transfected with control (siCtl) and YTHDF-targeting siRNAs (siYTHDF1#1 and siYTHDF1#2). Cells that migrated from the top to the underside of transwell filters in 24 h were fixed and stained with PI. d Top panel: quantification of the PI-stained migrated cells. Mean numbers of cells from three independent experiments are shown with SD error bars. **P < 0.01, ***P < 0.005 (Student’s t-test) vs. siCtl. Bottom panel: western blot showing expression of YTHDF1 in control (siCtl) and YTHDF1-targeting siRNA (siYTHDF1#1, siYTHDF1#2) transfected cells subjected to transwell migration assay. GAPDH used as a loading control