Pulsed-field gel electrophoresis analysis of the repair of psoralen plus UVA induced DNA photoadducts in Saccharomyces cerevisiae.

In the yeast Saccharomyces cerevisiae, double-strand breaks (DSB) have been observed during the DNA repair of psoralen plus UVA induced lesions. In the present paper, we analyzed this repair step in some detail using pulsed-field gel electrophoresis (CHEF) to get a better understanding of this phenomenon with regard to the type of lesions induced and the repair pathways involved. The results confirm that, during post-treatment incubation of Saccharomyces cerevisiae cells, DSB are formed. Their appearance is dose-dependent and the rate of induction is comparable in large (chromosome IV) and small (chromosome III) chromosomes. The formation of DSB is evidenced by the breakage of linear chromosomes III and IV, but also, after high doses, by the linearization of a circular form of chromosome III. The induction of DSB appears to be highly dependent on the induction of interstrand cross-links since they are clearly present after treatments with 8-MOP plus 365 nm radiation (inducing monoadducts and cross-linking in DNA), but practically absent after treatment with 8-MOP plus 405 nm radiation (inducing predominantly monoadducts) at comparable levels of photoadducts. The occurrence of DSB is dependent on the RAD2 and RAD52, but not on the RAD6 gene. It is likely that the specific processing of DNA lesions involving DSB is related to the genotoxic consequences observed.