Translesion synthesis of acetylaminofluorene-dG adducts by DNA polymerase zeta is stimulated by yeast Rev1 protein.

Translesion synthesis is an important mechanism in response to unrepaired DNA lesions during replication. The DNA polymerase zeta (Polzeta) mutagenesis pathway is a major error-prone translesion synthesis mechanism requiring Polzeta and Rev1. In addition to its dCMP transferase, a non-catalytic function of Rev1 is suspected in cellular response to certain types of DNA lesions. However, it is not well understood about the non-catalytic function of Rev1 in translesion synthesis. We have analyzed the role of Rev1 in translesion synthesis of an acetylaminofluorene (AAF)-dG DNA adduct. Purified yeast Rev1 was essentially unresponsive to a template AAF-dG DNA adduct, in contrast to its efficient C insertion opposite a template 1,N6-ethenoadenine adduct. Purified yeast Polzeta was very inefficient in the bypass of the AAF-dG adduct. Combining Rev1 and Polzeta, however, led to a synergistic effect on translesion synthesis. Rev1 protein enhanced Polzeta-catalyzed nucleotide insertion opposite the AAF-dG adduct and strongly stimulated Polzeta-catalyzed extension from opposite the lesion. Rev1 also stimulated the deficient synthesis by Polzeta at the very end of undamaged DNA templates. Deleting the C-terminal 205 aa of Rev1 did not affect its dCMP transferase activity, but abolished its stimulatory activity on Polzeta-catalyzed extension from opposite the AAF-dG adduct. These results suggest that translesion synthesis of AAF-dG adducts by Polzeta is stimulated by Rev1 protein in yeast. Consistent with the in vitro results, both Polzeta and Rev1 were found to be equally important for error-prone translesion synthesis across from AAF-dG DNA adducts in yeast cells.