Fig. 1: Kdm5c-knockdown is associated with a competitive advantage and decreased latency in AML.
A Schematic outline of the in vivo shRNA screen targeting chromatin associated factors. B Relative proportions of chromatin-associated factors targeted in the shRNA-library based on Gene Ontology analysis. The library consists of 849 shRNAs targeting 315 genes. C Schematic outline of the competitive assay in vivo (top panel) and in vitro (bottom panel). The in vivo competitive assay was performed by BMT of Lp30 cells transduced with target-shRNA (GFP-selection) or competitive non-targeting shRNA (YFP-selection) in a 1:1 GFP/YFP ratio. The competitive advantage was assayed by flow cytometry of the BM 3 weeks later. For the in vitro competitive assay, a fraction of the cell culture was analyzed by flow cytometry at each passage. D Representative flow cytometry profiles of input and output (4 weeks post-BMT) of shControl and Kdm5c-KD groups. Target-shRNA-transduced cells are GFP-positive versus YFP-positive BM cells transduced with competitive non-targeting control-shRNA. E Performance of Lp30 Kdm5c-KD cells in BMT normalized to input ratio of target/competitor cells (n = 4 per group). F Survival analysis of mice with transplanted shControl or Kdm5c-KD Lp30 cells (n = 8 per group). G Relative expression of Kdm5a, Kdm5b, and Kdm5c in shControl and Kdm5c-KD groups assayed by RT-qPCR and normalized to Actb expression. H Survival analysis of mice transplanted with Inv(16)/Tet2−/− cells transduced with shControl or shKdm5c. Insert: Relative expression of Kdm5a in shControl and Kdm5c-KD groups assayed by RT-qPCR and normalized to Actb expression. Relative expression of Kdm5c in shControl and Kdm5c-KD cells was assayed by RT-qPCR and normalized to Actb expression. These data are a representative example of two independent experiments. I Competitive culture of control and Kdm5c-KD Inv(16)/Tet2−/− cells.
