MiR-4796 enhances the sensitivity of breast cancer cells to ionising radiation by impairing the DNA repair pathway.

Abstract

MicroRNAs (miRNAs) are important regulators of DNA damage response (DDR) through post-transcriptional regulation on their target genes, which are implicated in DDR and DNA repair (DR). In this study, we investigated the functional roles and target genes of miR-4796 and miR-1287 in breast cancer cells in response to radiation. The molecular mechanism of miR-4796 in regulating the radiosensitivity of breast cancer cells was also elucidated. Real-time polymerase chain reaction detected miR-4796 and miR-1287 expression; colony formation assay and irradiation therapy tumour xenograft in vivo examined radiosensitising effect; comet assay assessed DNA damage; immunofluorescence imaging determined the formation of γ-H2AX foci; targetscan and RegRNA predicted target mRNAs; luciferase reporter and mutation assays validated target genes; western blotting detected the expression of genes at the protein level; and flow cytometry quantified the activities of nonhomologous end-joining (NHEJ) and homologous recombination (HR). The expressions of miR-4796 and miR-1287 were acutely fluctuated in response to ionising radiation. In the absence of radiation, overexpression of miR-1287 dramatically promoted growth of breast cancer cells in vitro and in vivo, whereas overexpression of miR-4796 did not affect cell growth. When under the treatment with radiation, overexpression of miR-4796 suppressed DR and sensitised cancer cells to radiation both in vitro and in vivo. However, such effect was only observed in cell assays in the overexpressed miR-1287 group, and not confirmed in vivo. We therefore further explored the molecular mechanism of action of miR-4796, and found that miR-4796 targeted multiple components of DDR and DR, including ATM, BRCA1, PARP and RAD51. Moreover, overexpression of miR-4796 inhibited the expression of these DDR components at the protein level. In addition, miR-4796 inhibited HR and NHEJ repair pathways and aggravated radiation-induced DNA damage. The findings here suggest that miR-4796 can enhance radiation-induced cell death by directly targeting multiple DDR components, and repress NHEJ and HR DNA repair pathways. miR-4796 can act as an effective radiation sensitising agent.