Efficiency of radiation-induced base lesion excision and the order of enzymatic treatment

Abstract

Purpose: To clarify whether initial base excision repair processes at clustered DNA damage sites comprising multiple base lesions affect subsequent excision processes via the formation of additional strand breaks by glycosylase and apurinic/apyrimidinic (AP) endonuclease base excision enzymes.

Materials and methods: Plasmid DNA (pUC18) as a model DNA molecule was exposed to high-linear-energy-transfer (LET) ionizing radiation (He²⁺or C⁶⁺ions) or low-LET ionizing radiation (X-rays) under various conditions to produce varied radical-scavenging effects. pUC18 was then treated sequentially or simultaneously with two bacterial base excision enzymes (glycosylases), namely, endonuclease III and formamidopyrimidine-DNA glycosylase, which convert pyrimidine (or abasic [AP] site) and purine (or AP site) lesions to single-strand breaks (SSB), respectively. Yields of additional SSB or double-strand breaks (DSB) as digestion products were examined after changing the order of enzymatic treatment.

Results: There were few differences among the enzymatic treatments, indicating that treatment order did not affect the final yields of additional SSB or DSB formed by glycosylase activity. This suggests that of the total damage, the fraction of clustered damage sites with a persistent base lesion dependent on the order of glycosylase treatment was insignificant if present.

Conclusion: Base lesion clusters induced by high- or low-LET radiation appear three or more base pairs apart, and are promptly converted to a DSB by glycosylase, regardless of the order of enzymatic treatment.

Keywords:

• Base excision repair
• base lesions
• clustered DNA damage
• high-LET radiation

Acknowledgements

We thank Professor P. O’Neill at the Gray Institute for Radiation Oncology & Biology at the University of Oxford for the useful suggestions throughout this work.

Disclosure statement

The authors have no declarations of interest to report. The authors alone are responsible for the content and writing of the paper.

Funding

This study was supported by a Grant-in-Aid for Science Research from the Japan Society for the Promotion of Science (JSPS) KAKENHI (grant No. 15H02823), and by the Research Project at NIRS-HIMAC (project Nos. 16B446, 19B446) and at JAEA-TIARA (project No. 121007).