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SRS2 and SGS1 prevent chromosomal breaks and stabilize triplet repeats by restraining recombination

Nature Structural & Molecular Biology volume 16, pages 159–167 (2009)Cite this article

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Abstract

Several molecular mechanisms have been proposed to explain trinucleotide repeat expansions. Here we show that in yeast srs2Δ cells, CTG repeats undergo both expansions and contractions, and they show increased chromosomal fragility. Deletion of RAD52 or RAD51 suppresses these phenotypes, suggesting that recombination triggers trinucleotide repeat instability in srs2Δ cells. In sgs1Δ cells, CTG repeats undergo contractions and increased fragility by a mechanism partially dependent on RAD52 and RAD51. Analysis of replication intermediates revealed abundant joint molecules at the CTG repeats during S phase. These molecules migrate similarly to reversed replication forks, and their presence is dependent on SRS2 and SGS1 but not RAD51. Our results suggest that Srs2 promotes fork reversal in repetitive sequences, preventing repeat instability and fragility. In the absence of Srs2 or Sgs1, DNA damage accumulates and is processed by homologous recombination, triggering repeat rearrangements.

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Figure 1: CAG•CTG repeats show increased fragility in the absence of Srs2 or Sgs1 helicases.
Figure 2: Effect of the trinucleotide repeat tract orientation on stability.
Figure 3: Analysis of replication intermediates at the ARG2 locus by two-dimensional gel electrophoresis.
Figure 4: Analysis of replication intermediates at ARG2 by two-dimensional gel electrophoresis in wild-type (WT) and mutant strains.
Figure 5: A model showing different pathways to repair replication fork damage due to structure-forming sequences.

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  • 26 January 2009

    In the version of this article initially published online, the labels in Figure 1c, in the key to Figure 4 and in Table 2 were incorrect. The error has been corrected for the print, PDF and HTML versions of this article.

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Acknowledgements

A.K. is grateful to G. Maffioletti for teaching her the two-dimensional gel electrophoresis technique and helping with her first successful experiments. A.K. and G.-F.R. thank the people who gave them advice concerning two-dimensional gel electrophoresis: M. Foiani, C. Maric, A. Ceschia and A. Kaykov. They also gratefully acknowledge the help of G. Millot for advice concerning the various statistical tests used in this manuscript. They and B.D. also thank their colleagues of the Unité de Génétique Moléculaire des Levures for many fruitful discussions and G. Fischer for careful reading of the manuscript. R.P.A. would like to acknowledge the help of K. Suryanarayanan in installing the SALVADOR program. A.K. was funded by the Ministère de la Recherche and the Fondation pour la Recherche Médicale (FRM). This work was supported by grant 3738 from the Association pour la Recherche contre le Cancer (ARC), grant ANR-05-BLAN-0331 from the Agence Nationale de la Recherche, US National Institutes of Health grant GM063066 to C.H.F., Tufts University FRAC award to C.H.F. and GSC Research Award to R.P.A. B.D. is a member of the Institut Universitaire de France.

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Author notes

  1. Alix Kerrest and Ranjith P Anand: These authors contributed equally to this work.

Authors and Affiliations

  1. Institut Pasteur, Unité de Génétique Moléculaire des Levures, CNRS, URA2171, Université Pierre et Marie Curie, UFR 927, 25 rue du Dr Roux, Paris, F-75015, France

    Alix Kerrest, Bernard Dujon & Guy-Franck Richard

  2. Department of Biology, Tufts University, Medford, 02155, Massachusetts, USA

    Ranjith P Anand, Rangapriya Sundararajan & Catherine H Freudenreich

  3. Instituto FIRC di Oncologia Molecolare, Via Amadello 16, Milano, 20141, Italy

    Rodrigo Bermejo & Giordano Liberi

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  1. Alix Kerrest
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  2. Ranjith P Anand
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  3. Rangapriya Sundararajan
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  4. Rodrigo Bermejo
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  6. Bernard Dujon
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  8. Guy-Franck Richard
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Contributions

A.K. and G.-F.R. conceived of and performed the instability and two-dimensional studies on yeast chromosome X; C.H.F. and R.P.A. conceived of and performed the instability and fragility studies on the YAC, with R.S. contributing the rad51Δ (CAG)0 and (CAG)70 fragility analyses; R.B. and G.L. gave expert assistance with two-dimensional gel electrophoresis; A.K., B.D., G.-F.R., C.H.F. and R.P.A. analyzed the data and wrote the manuscript.

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Correspondence to Guy-Franck Richard.

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Kerrest, A., Anand, R., Sundararajan, R. et al. SRS2 and SGS1 prevent chromosomal breaks and stabilize triplet repeats by restraining recombination. Nat Struct Mol Biol 16, 159–167 (2009). https://doi.org/10.1038/nsmb.1544

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