Fig. 5
STAT5A/B are essential for unrestricted proliferation. A Western blot showing protein levels of phospho (p) STAT5 and total STAT5 in fl/fl Pdgfrb following CRSIPR/Cas9 mediated knockout of either STAT5A, STAT5B or both genes. GAPDH serves as the loading control. The molecular weight of analyzed proteins in kiloDaltons (KDa) is shown on the left. B Heatmap depicting cell viability of fl/fl Pdgfrb primary tumor cells following CRSIPR/Cas9 deletion of either STAT5A, STAT5B or both genes. Cells were seeded in a 96-well plate in limiting dilutions and arbitrary fluorescence units were measured using a resazurin assay. C Representative FACS plots of Stat5aΔCRISPR, Stat5bΔCRISPR, Stat5a/bΔCRISPR, Stat3ΔCRISPR or MybΔCRISPR cells 39 days post-transduction. The left graphs represent ‘Count vs. mCherry’ and the right graphs represent ‘GFP (FITC-A channel) vs. mCherry (ECD-A channel)’. D Heatmap representing the survival of Lenti-EF1As-Cas9-P2A-GFP and U6-IT-mPgk-mCherry vector expressing cells over time. Viability was calculated as the percentage of mCherry+ cells relative to the negative non-targeting control (RosaΔCRISPR) for each condition on day 9. MybΔCRISPR was used as a positive control. E Cell viability of Stat5a/bΔCRISPR double knock out cells normalized to individual Stat5aΔCRISPR cells over time. Data is plotted as the percentage of mCherry+ cells relative to Day 9 post-transduction. F Cell viability of Stat5a/bΔCRISPR double knock out cells normalized to individual Stat5bΔCRISPR cells over time. Data is plotted as the percentage of mCherry+ cells relative to Day 9 post-transduction. B, D, E and F Data are shown as means ± SD