Limiting the persistence of a chromosome break diminishes its mutagenic potential

Nicole Bennardo, Amanda Gunn, Anita Cheng, Paul Hasty, Jeremy M. Stark

Producción científica: Articlerevisión exhaustiva

79 Citas (Scopus)

Resumen

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 coexpressing I-SceI with a non-processive 39 exonuclease, Trex2, which we predicted would cause partial degradation of I-SceI 39 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.

Idioma originalEnglish (US)
Número de artículoe1000683
PublicaciónPLoS Genetics
Volumen5
N.º10
DOI
EstadoPublished - oct 2009

ASJC Scopus subject areas

  • Genetics(clinical)
  • Genetics
  • Ecology, Evolution, Behavior and Systematics
  • Molecular Biology
  • Cancer Research

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