Dark Recovery Phenomena in Yeast: II. Conditions That Modify the Recovery Process

Abstract

The dark recovery kinetics of X- or UV-irradiated diploid yeast (Saccharomyces cerevisiae) are dose-dependent, since both the initial lag and the time required to attain maximum viable titer are increasing functions of dose. The recovery process exhibits temperature and pH optima close to 30[degree]C and pH 6.0, respectively; addition of certain monovalent and divalent cations to the irradiated suspensions can partially inhibit re -covery, but for other ions enhancement of recovery is seen. These results support the view that dark recovery of radiation damage in yeast is due to the action of an enzyme or enzyme system. Although the optimal conditions for dark recovery are in many ways similar to those for growth, the 2 processes appear to be mutually antagonistic. The reduction in recovery observed on addition of nutrients to the storage medium suggests that, once a cell is committed to postirradia-tion multiplication, it is incapable of recovery. Storage of irradiated cells either under N, or in the presence of metabolic antagonists such as 2,4-dinitrophenol, potassium cyanide, or sodium azide, can significantly reduce the final level of recovered viability. Conversely, storage under increased oxygen tension, or in the presence of ATP (but not AMP) or chloramphenicol, enhances recovery after X or UV irradiation. This suggests that energy is required for the repair reactions and that it must be supplied primarily through oxidative metabolism in the form of ATP. Treatment of yeast with DNA-specific agents such as ethyl methane sulfonate or acridine dyes serves to inhibit recovery after either X or UV irradiation. In addition, recovery can be inhibited by treatment with protein-specific agents, such as heat (60[degree]C) or caffeine. These effects probably reflect, on the one hand, excessive structural alteration of the DNA substrate and, on the ohter, damage to the repair enzymes involved.