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HDF1 and RAD17 Genes are Involved in DNA Double-strand Break Repair in Stationary Phase Saccharomyces cerevisiae

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Abstract

DNA repair, checkpoint pathways and protection mechanisms against different types of perturbations are critical factors for the prevention of genomic instability. The aim of the present work was to analyze the roles of RAD17 and HDF1 gene products during the late stationary phase, in haploid and diploid yeast cells upon gamma irradiation. The checkpoint protein, Rad17, is a component of a PCNA-like complex—the Rad17/Mec3/Ddc1 clamp—acting as a damage sensor; this protein is also involved in double-strand break (DBS) repair in cycling cells. The HDF1 gene product is a key component of the non-homologous end-joining pathway (NHEJ). Diploid and haploid rad17Δ/rad17Δ, and hdf1Δ Saccharomyces cerevisiae mutant strains and corresponding isogenic wild types were used in the present study. Yeast cells were grown in standard liquid nutrient medium, and maintained at 30°C for 21 days in the stationary phase, without added nutrients. Cell samples were irradiated with 60Co γ rays at 5 Gy/s, 50 Gy ≤ Dabs ≤ 200 Gy. Thereafter, cells were incubated in PBS (liquid holding: LH, 0 ≤ t ≤ 24 h). DNA chromosomal analysis (by pulsed-field electrophoresis), and surviving fractions were determined as a function of absorbed doses, either immediately after irradiation or after LH. Our results demonstrated that the proteins Rad17, as well as Hdf1, play essential roles in DBS repair and survival after gamma irradiation in the late stationary phase and upon nutrient stress (LH after irradiation). In haploid cells, the main pathway is NHEJ. In the diploid state, the induction of LH recovery requires the function of Rad17. Results are compatible with the action of a network of DBS repair pathways expressed upon different ploidies, and different magnitudes of DNA damage.

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Acknowledgements

We are grateful to Lourdes Blanc for expert technical assistance. This work was supported in part by PEDECIBA and Universidad de la República, Uruguay.

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Authors and Affiliations

  1. Lab. Radiobiología. Dpto. Biofísica. Facultad de Medicina y Facultad de Ciencias, Universidad de la República, Montevideo, Uruguay

    Elia Nunes, Ema Candreva, Nelson Bracesco, Ana Sánchez & Mercedes Dell

  2. Dpto. de Biofisica, Facultad de Medicina, Gral. Flores 2125.CP 11800., Montevideo, Uruguay

    Elia Nunes

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  1. Elia Nunes
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  2. Ema Candreva
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  3. Nelson Bracesco
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  4. Ana Sánchez
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  5. Mercedes Dell
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Correspondence to Elia Nunes.

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Nunes, E., Candreva, E., Bracesco, N. et al. HDF1 and RAD17 Genes are Involved in DNA Double-strand Break Repair in Stationary Phase Saccharomyces cerevisiae . J Biol Phys 34, 63–71 (2008). https://doi.org/10.1007/s10867-008-9105-0

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  • DOI: https://doi.org/10.1007/s10867-008-9105-0

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