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Limiting the persistence of a chromosome break diminishes its mutagenic potential.
- Source :
- PLoS Genetics, Vol 5, Iss 10, p e1000683 (2009)
- Publication Year :
- 2009
- Publisher :
- Public Library of Science (PLoS), 2009.
-
Abstract
- To characterize the repair pathways of chromosome double-strand breaks (DSBs), one approach involves monitoring the repair of site-specific DSBs generated by rare-cutting endonucleases, such as I-SceI. Using this method, we first describe the roles of Ercc1, Msh2, Nbs1, Xrcc4, and Brca1 in a set of distinct repair events. Subsequently, we considered that the outcome of such assays could be influenced by the persistent nature of I-SceI-induced DSBs, in that end-joining (EJ) products that restore the I-SceI site are prone to repeated cutting. To address this aspect of repair, we modified I-SceI-induced DSBs by co-expressing I-SceI with a non-processive 3' exonuclease, Trex2, which we predicted would cause partial degradation of I-SceI 3' overhangs. We find that Trex2 expression facilitates the formation of I-SceI-resistant EJ products, which reduces the potential for repeated cutting by I-SceI and, hence, limits the persistence of I-SceI-induced DSBs. Using this approach, we find that Trex2 expression causes a significant reduction in the frequency of repair pathways that result in substantial deletion mutations: EJ between distal ends of two tandem DSBs, single-strand annealing, and alternative-NHEJ. In contrast, Trex2 expression does not inhibit homology-directed repair. These results indicate that limiting the persistence of a DSB causes a reduction in the frequency of repair pathways that lead to significant genetic loss. Furthermore, we find that individual genetic factors play distinct roles during repair of non-cohesive DSB ends that are generated via co-expression of I-SceI with Trex2.
Details
- Language :
- English
- ISSN :
- 15537390 and 15537404
- Volume :
- 5
- Issue :
- 10
- Database :
- Directory of Open Access Journals
- Journal :
- PLoS Genetics
- Publication Type :
- Academic Journal
- Accession number :
- edsdoj.37f94376d0514eff8af9d0448fd73e77
- Document Type :
- article
- Full Text :
- https://doi.org/10.1371/journal.pgen.1000683