Effect of DNA Methylation on the 3′→5′ Exonuclease Activity of Major Human Abasic Site Endonuclease APEX1

Abstract

Abstract: Apurinic/apyrimidinic (AP) endonucleases are the key enzymes in the DNA base excision repair, as they hydrolyze the phosphodiester bond in the AP site formed after removal of the damaged base. Major human AP endonuclease APEX1 also possesses the 3′-phosphodiesterase and 3′→5′ exonuclease activities. The biological role of the latter has not been established yet; it is assumed that it corrects DNA synthesis errors during DNA repair. If DNA is damaged at the 3′-side of 5-methylcytosine (mC) residue, the 3′→5′ exonuclease activity can change the epigenetic methylation status of the CpG dinucleotide. It remains unclear whether the 3′→5′ exonuclease activity of APEX1 contributes to the active epigenetic demethylation or, on the contrary, is limited in the case of methylated CpG dinucleotides in order to preserve the epigenetic status upon repair of accidental DNA damage. Here, we report the results of the first systematic study on the efficiency of removal of 3′-terminal nucleotides from the substrates modeling DNA repair intermediates in the CpG dinucleotides. The best substrates for the 3′→5′ exonuclease activity of APEX1 were oligonucleotides with the 3′-terminal bases non-complementary to the template, while the worst substrates contained mC. The presence of mC in the complementary strand significantly reduced the reaction rate even for the non-complementary 3′-ends. Therefore, the efficiency of the 3′→5′ exonuclease reaction catalyzed by APEX1 is limited in the case of the methylated CpG dinucleotides, which likely reflects the need to preserve the epigenetic status during DNA repair.