Human and rodent cells proficient and deficient in non-homologous end joining (NHEJ) were irradiated with X rays, 70 keV/microm carbon ions, and 200 keV/microm iron ions, and the biological effects on these cells were compared. For wild-type CHO and normal human fibroblast (HFL III) cells, exposure to iron ions yielded the lowest cell survival, followed by carbon ions and then X rays. NHEJ-deficient xrs6 (a Ku80 mutant of CHO) and 180BR human fibroblast (DNA ligase IV mutant) cells showed similar cell survival for X and carbon-ion irradiation (RBE = approximately 1.0). This phenotype is likely to result from a defective NHEJ protein because xrs6-hamKu80 cells (xrs6 cells corrected with the wild-type KU80 gene) exhibited the wild-type response. At doses higher than 1 Gy, NHEJ-defective cells showed a lower level of survival with iron ions than with carbon ions or X rays, possibly due to inactivation of a radioresistant subpopulation. The G(1) premature chromosome condensation (PCC) assay with HFL III cells revealed LET-dependent impairment of repair of chromosome breaks. Additionally, iron-ion radiation induced non-repairable chromosome breaks not observed with carbon ions or X rays. PCC studies with 180BR cells indicated that the repair kinetics after exposure to carbon and iron ions behaved similarly for the first 6 h, but after 24 h the curve for carbon ions approached that for X rays, while the curve for iron ions remained high. These chromosome data reflect the existence of a slow NHEJ repair phase and severe biological damage induced by iron ions. The auto-phosphorylation of DNA-dependent protein kinase catalytic subunits (DNA-PKcs), an essential NHEJ step, was delayed significantly by high-LET carbon- and iron-ion radiation compared to X rays. This delay was further emphasized in NHEJ-defective 180BR cells. Our results indicate that high-LET radiation induces complex DNA damage that is not easily repaired or is not repaired by NHEJ even at low radiation doses such as 2 Gy.

Selective activation of p53 target genes in response to various cellular stresses is a critical step in determining the ability to induce cell-cycle arrest or apoptosis. Here we report the identification of the microRNA miR-22 as a p53 target gene that selectively determines the induction of p53-dependent apoptosis by repressing p21. Combinatorial analyses of the AGO2 immunocomplex and gene expression profiles identified p21 as a direct target of miR-22. Induction of p21 was inhibited by miR-22 after exposure to the genotoxic agent Adriamycin (doxorubicin; Bedford Laboratories), sensitizing cells to p53-dependent apoptosis. Interestingly, the activation of miR-22 depended on the intensity of the stresses that induced cells to undergo apoptosis in the presence of p21 suppression. Our findings define an intrinsic molecular switch that determines p53-dependent cellular fate through post-transcriptional regulation of p21. Cancer Res; 71(13); 4628-39. Ó2011 AACR.