Involvement of DNA polymerase β in protection against the cytotoxicity of oxidative DNA damage

Abstract

We had shown previously that DNA polymerase β (β-pol) null mouse fibroblasts, deficient in base excision repair (BER), are hypersensitive to monofunctional methylating agents but not to hydrogen peroxide (H 2O 2). This is surprising because β-pol is thought to be involved in BER of oxidative as well as methylated DNA damage. We confirm these findings here in early-passage cells. However, with time in culture, β-pol null cells become hypersensitive to H 2O 2 and other reactive oxygen species-generating agents. Analysis of in vitro BER reveals a strong deficiency in single-nucleotide BER of 8-oxoguanine (8-oxoG) by both early- and late-passage β-pol null cell extracts. Therefore, in early-passage wild-type and β-pol null cells, the capacity for single-nucleotide BER of 8-oxoG does not correlate with cellular sensitivity to H 2O 2. Expression of β-pol protein in the late-passage null cells almost completely reverses the H 2O 2-hypersensitivity phenotype. Methoxyamine (MX) treatment sensitizes late-passage wild-type cells to H 2O 2 as expected for β-pol-mediated single-nucleotide BER; however in β-pol null cells, MX has no effect. The data indicate a role(s) of β-pol-dependent repair in protection against the cytotoxicity of oxidative DNA damage in wild-type cells.