Ionizing radiation or hydroxyurea (HU) induce cell cycle delays mediated by checkpoint controls. Caffeine treatment can override checkpoint controls and permit cell cycle progression into mitosis, resulting in sensitivity to radiation and HU. Radiosensitivity is especially enhanced in p53-defective cells. The mechanism by which caffeine can override checkpoint controls will be examined in the fission yeast Schizosaccharomyces pombe. Mutants have been isolated which cannot be sensitized to radiation or HU by caffeine. Two are altered in rhp6 and retain checkpoint functions in the presence of caffeine. Based on comparisons with the budding yeast system, it i suggested that rhp6 requires fission yeast homologues of Ubr1 and Rad18. To test this model, fission yeast homologues of these genes will be isolated, and corresponding mutants will be constructed and subsequently tested for mutant phenotypes predicted by the model. The additional five previously identified fission yeast mutations that can override caffeine sensitization will be characterized. Roles in repair and cell cycle progression will be tested to support or modify the model.
