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Papers In Press, published online ahead of print June 4, 2001
J. Biol. Chem, 10.1074/jbc.M103588200
Submitted on April 23, 2001
Revised on June 4, 2001
Accepted on June 4, 2001

Recombinational and mutagenic repair of psoralen interstrand crosslinks in Saccharomyces cerevisiae

Ross B. Greenberg, Marie Alberti, John E. Hearst, Mark A. Chua, and Wilma A. Saffran

Chemistry and Biochemistry, Queens College, City University of New York, Flushing, NY 11367

Corresponding Author: wilma_saffran{at}qc.edu

Psoralen photoreacts with DNA to form interstrand crosslinks, which can be repaired by both non-mutagenic nucleotide excision repair and recombinational repair pathways and by mutagenic pathways. In the yeast Saccharomyces cerevisiae psoralen crosslinks are processed by nucleotide excision repair to form double strand breaks (DSBs). In yeast, DSBs are repaired primarily by homologous recombination, predicting that crosslink and DSB repair should induce similar recombination endpoints. We compared psoralen crosslink, psoralen monoadduct and DSB repair using plasmid substrates with site-specific lesions, and measured the patterns of gene conversion, crossing over and targeted mutation. Psoralen crosslinks induced both recombination and mutations, while DSBs induced only recombination, and monoadducts were neither recombinagenic nor mutagenic. Although the crosslink- and DSB-induced patterns of plasmid integration and gene conversion were similar in most respects, they showed opposite asymmetries in their unidirectional conversion tracts: primarily upstream from the damage site for crosslinks, but downstream for DSBs. Crosslinks induced targeted mutations in 5% of the repaired plasmids; all were base substitutions, primarily T-A to C-G transitions. The major pathway of psoralen crosslink repair in yeast is error-free, and involves formation of DSB intermediates, followed by homologous recombination. A fraction of the crosslinks enter an error-prone pathway, resulting in mutations at the damage site.


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