Fast Kinetics of the Oxygen Effect for DNA Double-strand Breakage and Cell Killing in Irradiated Yeast

Summary

Lifetimes of oxygen-dependent precursors of DNA double-strand breaks (dsb), as determined by applying the gas explosion technique, were found to be dose-dependent. The analysis of data for dsb induction obtained in diploid 211*B cells by the neutral sucrose sedimentation technique exhibits a half-life of 2.18 ms at a pulse dose of 500 Gy. Lifetimes are also obtained indirectly by analysing the inactivation of diploid rad54–3 cells, which are defective in the rejoining of dsb when incubated at 36°C. The half-life at a pulse dose of 40 Gy is only 0·25 ms. Since the cell size of both strains is very similar, the nine-fold longer lifetime determined in cells of strain 211*B may be caused by radiolytic depletion of glutathione due to the 12-fold higher dose applied to these cells. Therefore, the lifetime measured by the inactivation of rad54-3 cells (36°C) is considered to be more relevant than that obtained by direct measurements of dsb. The influence of dsb rejoining on the fast kinetics of the oxygen effect was studied using rad54-3 cells, which are capable of rejoining dsb when incubated at 23°C. When allowance for dsb rejoining was made, two components become detectable with half-lives of 0·75 and 29 ms. Haploid yeast cells in stationary phase are not capable of rejoining dsb. Using such cells proficient or deficient in the synthesis of glutathione (GSH), the lifetimes were found to be 0·37 or 0·49 ms for GSH-proficient (100 per cent GSH) and -deficient (2·1 per cent GSH) cells, which is in agreement with the view that chemical restitution of oxygen-dependent precursors is impaired in gsh⁻ cells.