Host: The Japan Radiation Research Society, Chairman of the 52nd Annual Meeting, Toshiteru Okubo (Radiation Effects Research Foundation)
ATM-dependent foci formation of checkpoint factors is important for the induction of DNA damage response. Previously, we developed the index to quantify the signals sufficient for initiating the G2/M checkpoint, which is the Sum Of Integrated Density (SOID). The SOID is calculated based on the intensity of the fluorescence for histone H2AX phosphorylation. It integrates foci number, foci area and density, reflecting the molecular numbers of checkpoint factors. In the present study, we determined the SOID value in cells that overcame the G2 checkpoint, and compared the SOID value in those, in which G2/M checkpoint was inactivated by ATM inhibitor KU55933 (KU). We also analyzed chromosomal aberrations to estimate the type of damage which was detected as phosphorylated H2AX foci.
Normal human diploid cells were exposed to low dose of X-irradiation. We found that mitotic cells disappeared two hours after 0.4 Gy of X-rays, whereas no such G2 arrest was observed with 0.1 Gy. Analysis of phosphorylated H2AX foci in the G1 cells, which progressed after irradiation of 0.4 Gy, showed SOID of less than 5000. We also examined chromosomal aberrations in 0.4 Gy-irradiated cells. The average number of chromosomal aberrations in irradiated cells was 2, but it increased to 5 with KU treatment. Most of the chromosomal aberrations were gaps and breaks.
These results indicated that there is a threshold in the DNA damage signal essential for the execution of G2/M checkpoint. For 0.4 Gy of X-rays, it is about 5000 SOID, which corresponds to 3~4 of chromosomal aberrations.
