Journal
NUCLEIC ACIDS RESEARCH
Volume 36, Issue 9, Pages 2895-2905Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkn126
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Funding
- Intramural NIH HHS Funding Source: Medline
- NCI NIH HHS [CA40615, R01 CA040615] Funding Source: Medline
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Ionizing radiation induces various clustered DNA lesions, including double-strand breaks (DSBs) accompanied by nearby oxidative base damage. Previous work showed that, in HeLa nuclear extracts, DSBs with partially complementary 3 overhangs and a one-base gap in each strand are accurately rejoined, with the gaps being filled by DNA polymerase . To determine the possible effect of oxidative base damage on this process, plasmid substrates were constructed containing overhangs with 8-oxoguanine or thymine glycol in base-pairing positions of 3-base (-ACG or -GTA) 3 overhangs. In this context, 8-oxoguanine was well tolerated by the end-joining machinery when present at one end of the break, but not when present at both ends. Thymine glycol was less well tolerated than 8-oxoguanine, reducing gap filling and accurate rejoining by at least 10-fold. The results suggest that complex DSBs can be accurately rejoined despite the presence of accompanying base damage, but that nonplanar bases constitute a major barrier to this process and promote error-prone joining. A chimeric DNA polymerase, in which the catalytic domain of polymerase was replaced with that of polymerase , could not substitute for polymerase in these assays, suggesting that this domain is specifically adapted for gap filling on aligned DSB ends.
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