Figure 1
Expression of RUNX1 or ZNF238 leads to terminal differentiation of RMS cells. (A) qPCR for RUNX1 was performed in RD cells either infected with a control virus, or the forced MyoD ~ E dimer (MD ~ E) as well as control (0 h) human fibroblasts and fibroblasts differentiated into myotubes (96 h). (B) RT-PCR for the two isoforms of ZNF238 in RD cells and fibroblasts as in 1A. (C) Myosin heavy chain (MHC) immunostains in RD cells either not infected (no infection), infected with a control GFP-expressing lentivirus (GFP control) or RUNX1 or ZNF238 expressing lentivirus. All cells were infected at approximately equivalent MOIs, and cells were allowed to differentiate for 72 h in low-serum media before staining. GFP was detected directly, without the use of an antibody. (D) qPCR for muscle-specific creatine kinase (CKM) in RD cells infected with either ZNF238 or RUNX1 viruses compared to cells with control retroviruses. (E) After 24 h of differentiation, RD cells were pulsed for a further 24 h with EdU-containing differentiation media, before fixation and quantification of the percentage of EdU-positive cells. (F) qPCR for ID2 and ID3 in control and ZNF238-expressing RD cells. All qPCR data are normalized to TIMM17b expression, and the level in control cells is set to 1. All bar graphs represent the mean ± SEM of at least three independent experiments. *: P < 0.05; **: P < 0.01; ***: P < 0.001; ****: P < 1 × 10-4.