Abstract
Non-homologous end joining (NHEJ) is the principal mechanism in human cells for repairing DNA double strand breaks (DSBs). The aim of this study is to clarify the biological roles of NHEJ-related genes on DNA damage induced by ionizing radiation (IR) through the generation and characterization of NHEJ-related gene deficient human cell lines. We have generated 3 cell lines that defective in XRCC4, Artemis and MDC1, respectively, by gene targeting in human colon cancer cell line HCT116. Chromosomal aberrations induced by X-ray irradiation were significantly higher in all deficient cell lines than those in parental HCT116. Survival rate after X-irradiation was the highest in the parental HCT116 and the lowest in XRCC4-/- cells, while MDC1-/- and Artemis-/- cells exhibited intermediated radio-sensitivities between the parental and XRCC4-/- cells. Generation of g-H2AX foci increased in a dose-dependent manner of X-rays and the number of foci reached the maximum at 30 ~ 60 min after X-irradiation in all cell lines. The foci then disappeared gradually and returned to the basal level within 4 hr in the parental HCT116, while a number of foci still remained at 4 hr after X-irradiation in XRCC4-/- and MDC1-/- cells. MDC1 might affect the formation of 53BP1 foci, since many g-H2AX foci did not colocalized to 53BP1 foci in the MDC1-/- cells after X-irradiation. These rersults suggest that defects in NHEJ-related genes cause a deterioration of g-H2AX foci-associated event, probably a DNA DSB repair process and in turn become a serious reason for the radio-sensitivity of theses cells.
