53BP1 contributes to survival of cells irradiated with X-ray during G1 without Ku70 or Artemis.

Ionizing radiation (IR) induces a variety of DNA lesions. The most significant lesion is a DNA double-strand break (DSB), which is repaired by homologous recombination or nonhomologous end joining (NHEJ) pathway. Since we previously demonstrated that IR-responsive protein 53BP1 specifically enhances activity of DNA ligase IV, a DNA ligase required for NHEJ, we investigated responses of 53BP1-deficient chicken DT40 cells to IR. 53BP1-deficient cells showed increased sensitivity to X-rays during G1 phase. Although intra-S and G2/M checkpoints were intact, the frequency of isochromatid-type chromosomal aberrations was elevated after irradiation in 53BP1-deficient cells. Furthermore, the disappearance of X-ray-induced gamma-H2AX foci, a marker of DNA DSBs, was prolonged in 53BP1-deficient cells. Thus, the elevated X-ray sensitivity in G1 phase cells was attributable to repair defect for IR-induced DNA-damage. Epistasis analysis revealed that 53BP1 plays a role in a pathway distinct from the Ku-dependent and Artemis-dependent NHEJ pathways, but requires DNA ligase IV. Strikingly, disruption of the 53BP1 gene together with inhibition of phosphatidylinositol 3-kinase family by wortmannin completely abolished colony formation by cells irradiated during G1 phase. These results demonstrate that the 53BP1-dependent repair pathway is important for survival of cells irradiated with IR during the G1 phase of the cell cycle.