CHARACTERIZATION OF THE ALTERNATIVE EXCISION-REPAIR PATHWAY OF UV-DAMAGED DNA IN SCHIZOSACCHAROMYCES-POMBE

Schizosaccharomyces pombe cells deficient in nucleotide excision repai r (NER) are still able to remove photoproducts from cellular DNA, show ing that there is a second pathway for repair of UV damage in this org anism. We have characterized this repair pathway by cloning and disrup tion of the genomic gene encoding UV damage endonuclease (UVDE). Altho ugh uvde gene disruptant cells are only mildly UV sensitive, a double disruptant of uvde and rad13 (a S.pombe mutant defective in NER) was s ynergistically more sensitive than either single disruptant and was un able to remove any photoproducts from cellular DNA. Analysis of the ki netics of photoproduct removal in different mutants showed that the UV DE-mediated pathway operates much more rapidly than NER. In contrast t o a previous report, our genetic analysis showed that rad12 and uvde a re not the same gene. Disruption of the rad2 gene encoding a structure -specific flap endonuclease makes cells UV sensitive, but much of this sensitivity is not observed if the uvde gene is also disrupted, Furth er genetic and immunochemical analyses suggest that DNA incised by UVD E is processed by two separate mechanisms, one dependent and one indep endent of flap endonuclease.