The effect of subcellular localization of tumor BRCA1 on the sensitivity of cell radiation and PARP inhibitor / 临床与实验病理学杂志

ABSTRACT

Purpose To investigate the effect of subcellular location of tumor BRCA1 on the sensitivity to ionizing radiation (IR) and PARP inhibitor.Methods siRNA of BRCA1 were first used to inhibit endougenous BRCA1 expression in MCF7 cells.Then,plasmids of pCMV-3xFlag-WT-BRCA1,pCMV-3xFlag-NES-BRCA1 and pCMV-3xFlag-NLS-BRCA1 were transfected in MCF7 cells.Immunofluorescence staining was used to detect BRCA1 subcellular location as well as the formation of Rad51 and γ-H2AX foci.Apoptotic cells were analyzed by flow cytometry,and colony formation assay was performed to evaluate the survival of cells.Results There were 47％ cells with nuclear BRCA1,23％ cells with cytoplasmic BRCA1 and 30％ cell with mixed nuclear and cytoplasmic BRCA1 expression in WT-BRCA1 transfected cell.There were 87％ cells with nuclear BRCA1 in NES-BRCA1 transfected cell,and 82％ cells with cytoplasmic BRCA1 in NLS-BRCA1 transfected cell.There were 87％,84％ and 13％ Rad51 foci positive cells at 2 hours after 4 Gy radiation treatment and 22％,25％ and 59％ γ-H2AX foci positive cells at 24 hours after 4Gy radiation treatment in WT-BRCA1,NES-BRCA1 mutant and NLS-BRCA1 mutant transfected cell respectively.ABT-888 and radiation treatment induced more apoptosis and fewer colonies in NLS-BRCA1 transfected cell than WT-BRCA1,NES-BRCA1 mutant transfected cell.Conclusion Subcellular location of BRCA1 might affect homologous recombination repair of DNA double strand breaks and can be used to predict sensitivity to IR and PARP inhibitor.