Published online 13 January 2006
Article |
Pol
, Pol
and Rev1 together are required for G to T transversion mutations induced by the (+)- and (–)-trans-anti-BPDE-N2-dG DNA adducts in yeast cells
Graduate Center for Toxicology, University of Kentucky Lexington, KY 40536, USA 1Department of Chemistry, New York University New York, NY 10003, USA
*To whom correspondence should be addressed. Tel: +1 859 323 5784; Fax: +1 859 323 1059; Email: zwang{at}uky.edu
Received November 11, 2005. Revised January 2, 2006. Accepted January 2, 2006.
Benzo[a]pyrene is an important environmental mutagen and carcinogen. Its metabolism in cells yields the mutagenic, key ultimate carcinogen 7R,8S,9S,10R-anti-benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide, (+)-anti-BPDE, which reacts via its 10-position with N2-dG in DNA to form the adduct (+)-trans-anti-BPDE-N2-dG. To gain molecular insights into BPDE-induced mutagenesis, we examined in vivo translesion synthesis and mutagenesis in yeast cells of a site-specific 10S (+)-trans-anti-BPDE-N2-dG adduct and the stereoisomeric 10R (–)-trans-anti-BPDE-N2-dG adduct. In wild-type cells, bypass products consisted of 76% C, 14% A and 7% G insertions opposite (+)-trans-anti-BPDE-N2-dG; and 89% C, 4% A and 4% G insertions opposite (–)-trans-anti-BPDE-N2-dG. Translesion synthesis was reduced by 
26–37% in rad30 mutant cells lacking Pol
, but more deficient in rev1 and almost totally deficient in rev3 (lacking Pol
) mutants. C insertion opposite the lesion was reduced by 24–33% in rad30 mutant cells, further reduced in rev1 mutant, and mostly disappeared in the rev3 mutant strain. The insertion of A was largely abolished in cells lacking either Pol, Pol or Rev1. The insertion of G was not detected in either rev1 or rev3 mutant cells. The rad30 rev3 double mutant exhibited a similar phenotype as the single rev3 mutant with respect to translesion synthesis and mutagenesis. These results show that while the Pol pathway is generally required for translesion synthesis and mutagenesis of the (+)- and (–)-trans-anti-BPDE-N2-dG DNA adducts, Pol, Pol and Rev1 together are required for G
T transversion mutations, a major type of mutagenesis induced by these lesions. Based on biochemical and genetic results, we present mechanistic models of translesion synthesis of these two DNA adducts, involving both the one-polymerase one-step and two-polymerase two-step models.
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