Figure 4. APEX2 is synthetic lethal with BRCA1 and BRCA2 deficiency.

(A-D) MCA assays were performed using the indicated gRNAs in the indicated cell backgrounds. GFP-labeled B2MUT cells were mixed with isogenic E2-Crimson-labeled B2WT cells, and the cell mixture was co-infected with the indicated individual gRNAs. After selection, the change in percent GFP⁺ cells was quantified by FACS before and after 7 d of culture, and normalized to negative control gRNAs. Error bars reflect the variability of biological triplicates, and the number of asterisks indicates the statistical significance for the corresponding experiment calculated by t-test (*=p<0.05,**=p<0.01,***=p<0.001). (E-F) MCA assays (as described above) in which colonic B2MUT and B2WT cells were mixed and treated for 12 d with the Ape nuclease inhibitor APEIII or the Ape1 redox inhibitor E3330. Error bars reflect the variability of biological triplicates, and asterisks indicated statistical significance as described above. (G) List of ORFs tested for complementation in cells expressing Cas9 and a gRNA to APEX2. (H) Examination of the ability of the ORFs from (F) to rescue the growth defect caused by expression of Cas9 and an APEX2 gRNA in ovarian B2MUT cells. Cells were co-infected with lentivirus expressing an APEX2 gRNA and the indicated gRNAresistant APEX2 ORF or negative control peptide. Growth was quantified after selection and growth for 8 d. (I) List of ORFs tested for complementation in cells expressing Cas9 and a gRNA to APEX2. (J) Examination of the ability of the ORFs from (I) to rescue the growth defect caused by expression of Cas9 and an APEX2 gRNA in ovarian B2MUT cells. Cells were co-infected with lentivirus expressing an APEX2 gRNA and the indicated gRNA-resistant APEX2 ORF or negative control peptide. Growth was quantified after selection and growth for 2 d.