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. 2013 Jul 8;122(7):1293–1304. doi: 10.1182/blood-2013-05-501072

Figure 1.

Figure 1

The concept of RAD52-dependent synthetic lethality in proliferating LSCs/LPCs. (A, upper) Fold over baseline expansion of different stem and progenitor cell subpopulations from 3 normal, 6 CML-CP, and 4 CML-AP samples during a 5-day culture; (lower) number of colonies from different stem and progenitor cell subpopulations; *P < .05 in comparison with normal counterparts. N.D., not determined. (B, upper) Differential expression of genes involved in HRR in stem and progenitor populations in CML-CP and CML-AP patients, and normal donors identified by gene ontology ANOVA; (lower) heat map of HRR genes upregulated in LSCs and LPCs compared with normal HSCs. Results from HSCs and LSCs are highlighted in boxes. (C) Ingenuity pathway analysis identifies HRR as a key upregulated pathway in LSCs. (D) Percentage of the remaining colony-forming activity of Lin−CD34+ bone marrow cells from 3 healthy donors (○) and 3 CML-CP patients (●) treated with indicated micromol concentrations of B02 and RI-1. (E) Percentage of Flag-RAD51(WT) and Flag-RAD51(F259V) foci colocalizing with endogenous RAD52 in 10 cells/group; *P < .001. (F) Number of GFP+ cells representing HRR activity in parental 32Dcl3 (32D) and BCR-ABL1-positive 32Dcl3 (32D-B/A) cells expressing Flag-RAD51(WT) (black bars) and Flag-RAD51(F259V) (gray bars); *P = .003. (G) Number of colonies from parental UT7 and BCR-ABL1-positive UT7 (UT7-B/A) cells expressing Flag-RAD51(WT) (black bars) and Flag-RAD51(F259V) (gray bars); *P < .001. (H-I) RAD52-dependent synthetic lethality. (H) HSCs/HPCs usually employ the BRCA1/BRCA2/PALB2-RAD51 pathway to repair a DSB, whereas the RAD52-RAD51 axis forms an alternative mechanism. Thus, RAD51 foci often colocalize with BRCA1, but not RAD52 foci. (I) The downregulation of BRCA1 protein in CML-CP LSCs/LPCs forces them to use the RAD52-RAD51 pathway. In concordance, RAD51 nuclear foci often colocalize with RAD52, but not BRCA1. Twenty-five cells per group were analyzed; representative nuclear foci are shown (yellow color indicates colocalization).