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Nucleic Acids Research, 2001, Vol. 29, No. 13 2875-2883
© 2001 Oxford University Press

Translesion synthesis by yeast DNA polymerase {zeta} from templates containing lesions of ultraviolet radiation and acetylaminofluorene

Dongyu Guo, Xiaohua Wu, Deepak K. Rajpal, John-Stephen Taylor1 and Zhigang Wang* Graduate Center for Toxicology, 306 Health Sciences Research Building, University of Kentucky, Lexington, KY 40536, USA and 1Department of Chemistry, Washington University, St Louis, MO 63130, USA

In the yeast Saccharomyces cerevisiae, DNA polymerase {zeta} (Pol{zeta}) is required in a major lesion bypass pathway. To help understand the role of Pol{zeta} in lesion bypass, we have performed in vitro biochemical analyses of this polymerase in response to several DNA lesions. Purified yeast Pol{zeta} performed limited translesion synthesis opposite a template TT (6-4) photoproduct, incorporating A or T with similar efficiencies (and less frequently G) opposite the 3' T, and predominantly A opposite the 5' T. Purified yeast Pol{zeta} predominantly incorporated a G opposite an acetylaminofluorene (AAF)-adducted guanine. The lesion, however, significantly inhibited subsequent extension. Furthermore, yeast Pol{zeta} catalyzed extension DNA synthesis from primers annealed opposite the AAF-guanine and the 3' T of the TT (6-4) photoproduct with varying efficiencies. Extension synthesis was more efficient when A or C was opposite the AAF-guanine, and when G was opposite the 3' T of the TT (6-4) photoproduct. In contrast, the 3' T of a cis–syn TT dimer completely blocked purified yeast Pol{zeta}, whereas the 5' T was readily bypassed. These results support the following dual-function model of Pol{zeta}. First, Pol{zeta} catalyzes nucleotide incorporation opposite AAF-guanine and TT (6-4) photoproduct with a limited efficiency. Secondly, more efficient bypass of these lesions may require nucleotide incorporation by other DNA polymerases followed by extension DNA synthesis by Pol{zeta}.

* To whom correspondence should be addressed. Tel: +1 859 323 5784; Fax: +1 859 323 1059; Email: zwang{at}pop.uky.edu


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