CML SPCs show regulation of the hsa-mir183/EGR1 axis. (A) Genome-wide miRNA expression for healthy and CML SPCs (N = 5 biological replicates). (Bi) Heatmap for the OncoMir MiRNA Q-PCR Array in SPCs showing statistically significant regulation of miRNAs between healthy and CML SPCs. (ii) Q-PCR for hsa-mir183 regulation in CML vs healthy SPCs. (C) CML SPCs treated for 24 hours with dasatinib (DAS; 150 nM) and nilotinib (NIL; 1 µM) and analysis by Q-PCR for hsa-mir183. (Di) mRNA level of the hsa-mir183 target gene EGR1 in healthy and CML CD34⁺38⁻ cells. (ii) CML CD34⁺ cells treated for 7 days with imatinib (IM; 5 µM) and live cells analyzed by Q-PCR for EGR1 expression (N = 6). (E) hsa-mir183 knockdown GFP⁺ CML SPCs sorted and analyzed for EGR1 mRNA level by Q-PCR. Mir zip-scrambled vector was used as negative control. (F) Representative plot for cell divisions analyzed in hsa-mir183 knockdown GFP⁺ CML SPCs using Cell Trace Violet staining. Colcemid treatment used to visualize undivided cells. (G) CFC analysis carried out in hsa-mir183 knockdown CML SPCs. (H) Luciferase assay showing binding between of hsa-mir183 and EGR1 in KCL22 cells. Cells transfected with oligos containing the EGR1 3′UTR (along with the binding site for hsa-mir183, WT) or a mutant version (MUT) together with hsa-mir183 mimic or scrambled negative control. Each experiment had N = 3 biological replicates; *P < .05; **P < .01; *** P < .001. ND, no drug.