The Rev1-Polζ pathway is believed to be the major mechanism of translesion DNA synthesis and base damage-induced mutagenesis in eukaryotes. While it is widely believed that Rev1 plays a non-catalytic function in translesion synthesis, the role of its dCMP transferase activity remains uncertain. To determine the relevance of its catalytic function in translesion synthesis, we separated the Rev1 dCMP transferase activity from its non-catalytic function in yeast. This was achieved by mutating two conserved amino acid residues in the catalytic domain of Rev1, i.e. D467A/E468A, where its catalytic function was abolished but its non-catalytic function remained intact. In this mutant strain, whereas translesion synthesis and mutagenesis of UV radiation were fully functional, those of a site-specific 1,N6-ethenoadenine were severely deficient. Specifically, the predominant A→G mutations resulting from C insertion opposite the lesion were abolished. Therefore, translesion synthesis and mutagenesis of 1,N6-ethenoadenine require the catalytic function of the Rev1 dCMP transferase, in contrast to those of UV lesions, which only require the non-catalytic function of Rev1. These results show that the catalytic function of the Rev1 dCMP transferase is required in a lesion-specific manner for translesion synthesis and base damage-induced mutagenesis.
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