Abstract
Oxidative stress is a very frequent source of DNA damage. Many cellular DNA polymerases (Pols) can incorporate ribonucleotides (rNMPs) during DNA synthesis. However, whether oxidative stress-triggered DNA repair synthesis contributes to genomic rNMPs incorporation is so far not fully understood. Human specialized Pols β and λ are the important enzymes involved in the oxidative stress tolerance, acting both in base excision repair and in translesion synthesis past the very frequent oxidative lesion 7,8-dihydro-8-oxoguanine (8-oxo-G). We found that Pol β, to a greater extent than Pol λ can incorporate rNMPs opposite normal bases or 8-oxo-G, and with a different fidelity. Further, the incorporation of rNMPs opposite 8-oxo-G delays repair by DNA glycosylases. Studies in Pol β- and λ-deficient cell extracts suggest that Pol β levels can greatly affect rNMP incorporation opposite oxidative DNA lesions.
Highlights
Oxidative stress is a very frequent source of DNA damage
Since the products resulting from ribonucleoside monophosphates (rNMPs) incorporation were migrating slower than those arising from dNMP incorporation, they could be distinguished on the sequencing gels
In addition to providing the necessary gap-filling synthesis, we have previously shown that both Pols operate in a specialized type of translesion synthesis (TLS) in the context of MutY homologue (MutYH)-initiated repair of 8-oxo-G:A mismatches[25]
Summary
Oxidative stress is a very frequent source of DNA damage. Many cellular DNA polymerases (Pols) can incorporate ribonucleotides (rNMPs) during DNA synthesis. During DNA replication 8-oxo-G lesions in the template strand lead to high frequency of misincorporation, generating A:8-oxo-G mismatches The repair of these mismatches requires the sequential action of the MutY homologue (MutYH) and 8-oxo-G DNA glycosylase (OGG1), as well as of a specialized TLS Pol. We have previously shown that Pol l is the most accurate in MutYH-initiated pathway, ensuring correct dCMP incorporation opposite 8-oxo-G, while Pol b can substitute for Pol l, but at the expense of a reduced fidelity, leading to frequent misincorporation of dAMP opposite the lesion[25]. We have previously shown that Pol l is the most accurate in MutYH-initiated pathway, ensuring correct dCMP incorporation opposite 8-oxo-G, while Pol b can substitute for Pol l, but at the expense of a reduced fidelity, leading to frequent misincorporation of dAMP opposite the lesion[25] It is not known whether DNA repair synthesis by Pols b and l, either during BER or 8-oxo-G bypass, could contribute to genomic rNMPs accumulation. Our results suggest that Pol b, to a greater extent than Pol l, plays a role in rNMP incorporation opposite oxidative DNA damage
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