期刊
DNA REPAIR
卷 12, 期 6, 页码 422-432出版社
ELSEVIER
DOI: 10.1016/j.dnarep.2013.03.005
关键词
Nonhomologous end joining; Free radicals; Oxidative damage; Clustered DNA damage
资金
- National Institutes of Health, US Public Health Service [CA40615, CA151367, CA84442]
- US Department of Energy Office of Science [DE-AC02-05CH11231]
Both Metnase and Artemis possess endonuclease activities that trim 3' overhangs of duplex DNA. To assess the potential of these enzymes for facilitating resolution of damaged ends during double-strand break rejoining, substrates bearing a variety of normal and structurally modified 3' overhangs were constructed, and treated either with Metnase or with Artemis plus DNA-dependent protein kinase (DNA-PK). Unlike Artemis, which trims long overhangs to 4-5 bases, cleavage by Metnase was more evenly distributed over the length of the overhang, but with significant sequence dependence. In many substrates, Metnase also induced marked cleavage in the double-stranded region within a few bases of the overhang. Like Artemis, Metnase efficiently trimmed overhangs terminated in 3'-phosphoglycolates (PGs), and in some cases the presence of 3'-PG stimulated cleavage and altered its specificity. The nonplanar base thymine glycol in a 3' overhang severely inhibited cleavage by Metnase in the vicinity of the modified base, while Artemis was less affected. Nevertheless, thymine glycol moieties could be removed by Metnase- or Artemis-mediated cleavage at sites farther from the terminus than the lesion itself. In in vitro end-joining systems based on human cell extracts, addition of Artemis, but not Metnase, effected robust trimming of an unligatable 3'-PG overhang, resulting in a dramatic stimulation of ligase IV- and XLF-dependent end joining. Thus, while both Metnase and Artemis are biochemically capable of resolving a variety of damaged DNA ends for the repair of complex double-strand breaks, Artemis appears to act more efficiently in the context of other nonhomologous end joining proteins. (c) 2013 Elsevier B.V. All rights reserved.
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