8-Oxo-2'-deoxyguanosine Replication in Mutational Hot Spot Sequences of the p53 Gene in Human Cells Is Less Mutagenic than That of the Corresponding Formamidopyrimidine

7,8-Dihydro-8-oxo-2 '-deoxyguanosine (8-OxodGuo) is a ubiquitous DNA damage formed by oxidation of 2 '-deoxyguanosine. In this study, plasmid DNA containing 8-OxodGuo located in three mutational hot spots of human cancers, codons 248, 249, and 273 of the Tp53 tumor suppressor gene, was replicated in HEK 293T cells. 8-OxodGuo was only a weak block of replication, and the bypass was largely error-free. The mutations (1-5%) were primarily G-* T transversions, and the mutation frequency was generally lower than that of the chemically related Fapy center dot dG. A unique 8-OxodGuo mutation spectrum was observed at each site, as reflected by replication in translesion synthesis (TLS) polymerase-or hPol A-deficient cells. In codon 248 (CG*G) and 249 (AG*G), where G* denotes 8-OxodGuo, hPol ? and hPol C carried out largely error-free bypass of the lesion, whereas hPol lc and hPol t were involved mostly in error-prone TLS, resulting in G-* T mutations. 8-OxodGuo bypass in codon 273 (CG*T) was unlike the other two sites, as hPol lc participated in the mostly error-free bypass of the lesion. Yet, in all three sites, including codon 273, simultaneous deficiency of hpol lc and hPol t resulted in reduction of G-* T transversions. This indicates a convincing role of these two TLS polymerases in error-prone bypass of 8-OxodGuo. Although the dominant mutation was G-* T in each site, in codon 249, and to a lesser extent in codon 248, significant semi-targeted single-base deletions also occurred, which suggests that 8-OxodGuo can initiate slippage of a base near the lesion site. This study underscores the importance of sequence context in 8-OxodGuo mutagenesis in human cells. It also provides a more comprehensive comparison between 8-OxodGuo and the sister lesion, Fapy center dot dG. The greater mutagenicity of the latter in the same sequence contexts indicates that Fapy center dot dG is a biologically significant lesion and biomarker on par with 8-OxodGuo.

Automated summary

This study found that 8-OxodGuo is a common DNA damage and can be located in three mutational hotspots in human cancer cells. The study also found that 8-OxodGuo has a low replication-blocking effect and low error rate. Different positions of 8-OxodGuo have different mutation spectra, but hPol lc and hPol t play important roles in error-prone bypass of 8-OxodGuo at all positions.