Gene expression profiling in pGBM cells. FACS-purified CD133+ and CD133− cells were transduced with Lenti-BMI1-693 for 48 hr in vitro and harvested for RNA extraction. Tumor cells receiving no or non-target lenti-shRNA were included as control. (A) Down-regulation of BMI1 mRNA in CD133+ (+) and CD133- (−) cells transduced by Lenti-BMI1-693. For IC-2305GBM, the CD133− cells were not included due to low cell content. (B) List of gene numbers differentially induced by lentin-BMI1-693 in CD133+ and CD133− cells derived from two PDOX models. Only a small fraction of genes were shared. (C) Heatmaps of gene expression showing changes of known genes previously associated with the over-expressed BMI1 (left panel) and the new targets associated with the silencing of BMI1 (right panel). In the known gene panel, those genes originally up-regulated by over-expressed BMI1 are indicated with up-arrows (↑), and those inhibited by the over-expressed BMI1 are unmarked. No reversal of expression levels were observed in these genes. In the new target panel, genes consistently altered by Lenti-BMI1-693 in the CD133+ cells from all 3 models are highlighted in the blue shaded area. Arrows indicate the 4 genes that were validated with qRT-PCR in (D) where the mRNA levels in cells transduced with Lenti-BMI1-693 were normalized with those transduced with Lenti-non-target shRNA and plotted as fold changes (log2).