Identification of APN2, the Saccharomyces cerevisiae homolog of the major human AP endonuclease HAP1, and its role in the repair of abasic sites

Abstract

Abasic (AP) sites arise in DNA through spontaneous base loss and enzymatic removal of damaged bases. APN1 encodes the major AP-endonuclease of Saccharomyces cerevisiae. Human HAP1 (REF1) encodes the major AP endonuclease which, in addition to its role in DNA repair, functions as a redox regulatory protein. We identify APN2, the yeast homolog of HAP1 and provide evidence that Apn1 and Apn2 represent alternate pathways for repairing AP sites. The apn1⌬ apn2⌬ strain displays a highly elevated level of MMS-induced mutagenesis, which is dependent on the REV3, REV7, and REV1 genes. Our findings indicate that AP sites are highly cytotoxic and mutagenic in eukaryotes, and that the REV3, REV7-encoded DNA polymerase mediates the mutagenic bypass of AP sites.

Early steps in the DNA base excision/single-strand interruption repair pathway in mammalian cells

“…The structure of substrate bound active APE1 could not be solved in the presence of Mg 2+ for obvious reasons. The key residues and motifs required for catalytic activity and metal binding in all Xth family members are conserved which allowed our lab to clone the S. cerevisiae Apn2 [99]. We subsequently identified and cloned S. pombe APN2 based on sequence homology [104].…”

“…Xth and Nfo type APEs, named APN2 and APN1, respectively, are expressed in yeast [98,99]. In contrast to the situation in E. coli, APN1 is the major contributor of APE activity in this unicellular eukaryote.…”

Dynamics and diversions in base excision DNA repair of oxidized abasic lesions

“…The best studied AP endonucleases are E. coli exonuclease III and its human homolog Ape1, and also E. coli endonuclease IV (Mol et al, 2000). Less well-characterized members of this family have recently been identified in yeast (Eth1/Apn2 protein) (Bennett, 1999;Johnson et al, 1998b) and mammalian cells (Ape2 protein) (Hadi et al, 2002;Hadi and Wilson, 2000;Tsuchimoto et al, 2001). A structurally distinct family of AP endonucleases is typified by E. coli endonuclease IV and S. cerevisiae Apn1 protein.…”

“…The yeast S. cerevisiae contains one partial homolog of exonuclease III, the Eth1/Apn2 protein (Bennett, 1999;Johnson et al, 1998b). This protein consists of a *350-residue N-terminal domain homologous to exonuclease III and an C-terminal extension of *160 residues unrelated to other AP endonucleases.…”

Yeast Rev1 Protein Is a G Template-specific DNA Polymerase

“…Although the Rev1 protein is almost as indispensable for UV mutagenesis as is Pol, its C-transferase activity is not needed for this function, because C insertion occurs only rarely opposite the UV lesions in yeast (15,16) and inactivation of this biochemical activity has no impact on UV mutagenesis. 2 The mutagenic bypass of abasic sites also requires Rev1 and Pol (17), and it depends upon the sequential action of two DNA polymerases in which one inserts the nucleotide opposite the abasic site and Pol subsequently extends from the inserted nucleotide (18). Although Rev1 is able to insert a C residue opposite an abasic site in an in vitro reaction, the inactivation of Rev1 C-transferase activity causes no significant reduction in the incidence of mutations resulting from the bypass of abasic sites (18).…”