Fig. 5: CXorf56 protein interacts with Ku70, suppressing Ku70-mediated NHEJ repair to enhance Olaparib resistance.

CXorf56 protein (A) and Ku70 (B) complexes were co-immunoprecipitated with CXorf56 protein and Ku70 antibodies and immunoblotted with the indicated antibodies. C, D MDA-MB-231 cells were infected with indicated lentiviral plasmids and treated with Olaparib. The indicated protein levels (C) and the sensitivity to Olaparib were assessed (D). Data were analyzed by a two-tailed t test. E, F NTC and corresponding knockdown MDA-MB-231 cells were treated with Olaparib (1.5 μM), and γ-H2AX foci before or 48 hours after Olaparib treatment were detected by immunofluorescence. Representative images of γ-H2AX foci are shown in E; Scale bars = 10 μm. Quantification of focus signals is shown in F. Data were analyzed by a two-tailed t test. G, J NTC and corresponding knockdown MDA-MB-231 cells were transfected with NHEJ reporter or HR reporter along with pCBA-I-SceI and mCherry. 48 hours later, cells were harvested and subjected to flow cytometric analysis. Data were analyzed by a two-tailed t test. I–K The tumor growth of the indicated MDA-MB-231 cells was examined in xenografts under the treatment of Olaparib (n = 6). Mouse xenograft tumors (I), the elevation of tumor size for 40 days (J), and the weights of the xenograft tumors (K) were presented. Data were analyzed by a two-tailed t test. L Representative IHC images of CXorf56 protein, Ku70, and γ-H2AX expression in mouse xenograft tumor tissues; Scale bars = 100 μm. M Differences in the γ-H2AX protein levels in xenograft tumor tissues were presented in the violin plot (n = 6). Data were analyzed by a two-tailed t test.Remarks: ^nsp ≥ 0.05, *p < 0.05, **p < 0.01, ***p < 0.001; Data are presented as mean ± SD.