Abstract
ATM kinase is activated after ionizing radiation (IR) and transduces DNA damage signal by phosphorylating various substrates, including itself. We previously demonstrated that persistent foci of Ser1981-phosphorylated ATM grow to more than 1.6 µm in diameter and the foci growth is quite relevant to G1 checkpoint induction. Moreover, our analysis revealed that the frequency of the grown foci increased as a quadratic function of IR dose. The results led us to draw hypothesis that interaction between two DNA double-strand breaks, which results in chromosome translocation, elicits the foci growth. To test the hypothesis, we performed chromosome analysis (Giemsa staining and WCP-FISH of Chr. #1-6) of the first metaphase 36-48 h after 2 Gy-gamma-irradiation on confluent normal human diploid fibroblasts (S+G2+M<2%) treated with ATM inhibitor, KU55933 (KU). We found that dicentric with fragment (a kind of chromosome translocation) was 4-fold more in KU-treated population than in DMSO-treated one in Giemsa staining. By WCP-FISH analysis, 3-7-fold more chromosome translocation was detected in KU-treated population than in DMSO-treated one. We also performed the similar examination using p53 siRNA to inhibit only G1 checkpoint function of ATM. We found that 2.5-fold more dicentric with fragment was observed in p53-siRNA-treated population than in control-siRNA-treated one. These results implicate that the foci growth-dependent G1 checkpoint is a mechanism in normal human cells, which recognizes chromosome translocation and suppresses propagation of cells with chromosome translocation.
