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Radiation and Environmental Biophysics

, Volume 46, Issue 4, pp 401–407 | Cite as

Roles of Saccharomyces cerevisiae RAD17 and CHK1 checkpoint genes in the repair of double-strand breaks in cycling cells

  • Nelson Bracesco
  • Ema C. Candreva
  • Deborah Keszenman
  • Ana G. Sánchez
  • Sandra Soria
  • Mercedes Dell
  • Wolfram Siede
  • Elia NunesEmail author
Original Paper

Abstract

Checkpoints are components of signalling pathways involved in genome stability. We analysed the putative dual functions of Rad17 and Chk1 as checkpoints and in DNA repair using mutant strains of Saccharomyces cerevisiae. Logarithmic populations of the diploid checkpoint-deficient mutants, chk1Δ/chk1Δ and rad17Δ/rad17Δ, and an isogenic wild-type strain were exposed to the radiomimetic agent bleomycin (BLM). DNA double-strand breaks (DSBs) determined by pulsed-field electrophoresis, surviving fractions, and proliferation kinetics were measured immediately after treatments or after incubation in nutrient medium in the presence or absence of cycloheximide (CHX). The DSBs induced by BLM were reduced in the wild-type strain as a function of incubation time after treatment, with chromosomal repair inhibited by CHX. rad17Δ/rad17Δ cells exposed to low BLM concentrations showed no DSB repair, low survival, and CHX had no effect. Conversely, rad17Δ/rad17Δ cells exposed to high BLM concentrations showed DSB repair inhibited by CHX. chk1Δ/chk1Δ cells showed DSB repair, and CHX had no effect; these cells displayed the lowest survival following high BLM concentrations. Present results indicate that Rad17 is essential for inducible DSB repair after low BLM-concentrations (low levels of oxidative damage). The observations in the chk1Δ/chk1Δ mutant strain suggest that constitutive nonhomologous end-joining is involved in the repair of BLM-induced DSBs. The differential expression of DNA repair and survival in checkpoint mutants as compared to wild-type cells suggests the presence of a regulatory switch-network that controls and channels DSB repair to alternative pathways, depending on the magnitude of the DNA damage and genetic background.

Keywords

Mutant Strain Proliferation Kinetic Liquid Nutrient Medium Solid Nutrient Medium Cell Proliferation Kinetic 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

This work was supported by NIH/Fogarty International Research Collaboration Award TW01189. We are indebted to Lourdes Blanc for excellent technical assistance, to PEDECIBA (Uruguay) for partial support, and to Dr. Anna Friedl for comments on the manuscript.

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Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Nelson Bracesco
    • 1
  • Ema C. Candreva
    • 1
  • Deborah Keszenman
    • 1
  • Ana G. Sánchez
    • 1
  • Sandra Soria
    • 1
  • Mercedes Dell
    • 1
  • Wolfram Siede
    • 2
  • Elia Nunes
    • 1
    Email author
  1. 1.Lab. Radiobiología, Departamento Biofísica, Facultad de MedicinaUniversidad de la RepúblicaMontevideoUruguay
  2. 2.Department of Cell Biology and GeneticsUniversity of North Texas Health Science CenterFort WorthUSA

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