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Reduced Rif2 and lack of Mec1 target short telomeres for elongation rather than double-strand break repair

Nature Structural & Molecular Biology volume 17, pages 1438–1445 (2010)Cite this article

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Abstract

Telomerase in Saccharomyces cerevisiae binds and preferentially elongates short telomeres, and this process requires the checkpoint kinase Tel1. Here we show that the Mre11 complex bound preferentially to short telomeres, which could explain the preferential binding of Tel1 to these ends. Compared to wild-type length telomeres, short telomeres generated by incomplete replication had low levels of the telomerase inhibitory protein Rif2. Moreover, in the absence of Rif2, Tel1 bound equally well to short and wild-type length telomeres, suggesting that low Rif2 content marks short telomeres for preferential elongation. In congenic strains, a double-strand break bound at least 140 times as much Mec1 in the first cell cycle after breakage as did a short telomere in the same time frame. Binding of replication protein A was also much lower at short telomeres. The absence of Mec1 at short telomeres could explain why they do not trigger a checkpoint-mediated cell-cycle arrest.

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Figure 1: MRX binds preferentially to short telomeres.
Figure 2: Mec1-HA does not bind preferentially to short telomeres, even in tel1Δ cells.
Figure 3: Mec1 and Rfa1 bind DSBs, even when the break is adjacent to telomeric DNA.
Figure 4: Rfa1 binding and H2A phosphorylation are similar at short and wild-type length telomeres.
Figure 5: Rif2 (and Rap1) but not Rif1 or Yku80 occupancy are reduced at short telomeres.
Figure 6: Preferential binding of Tel1 to short telomeres is lost in rif2Δ cells.

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Acknowledgements

We thank D. Shore and A. Bianchi (University of Geneva) for strains and advice on the DSB assay, K. Runge for advice on telomere PCR, T. Petes (Duke University) for strains, M. Jayaram for discussions about FLP-induced DSBs and C. Webb and Y. Wu for comments on the manuscript. This work was supported by grants from the US National Institutes of Health (NIH; GM43265 to V.A.Z.), postdoctoral fellowships from the NIH (M.S.), Deutsche Forschungsgemeinschaft (K.P.) and NJCCR (K.P.), and predoctoral fellowships from NJCCR (J.A.P., J.S.M.) and NIH (J.S.M.).

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  1. Jean S McGee and Jane A Phillips: These authors contributed equally to this work.

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  1. Department of Molecular Biology, Princeton University, Princeton, New Jersey, USA

    Jean S McGee, Jane A Phillips, Angela Chan, Michelle Sabourin, Katrin Paeschke & Virginia A Zakian

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Contributions

J.S.M. did the experiments in Figures 5 and 6, J.A.P. did the experiments in Figure 3, A.C. and M.S. did experiments in Figures 1, 2 and 4, and K.P. helped with analysis of H2A phosphorylation. V.A.Z. and all other authors participated in the design and interpretation of experiments and in the preparation of the manuscript.

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Correspondence to Virginia A Zakian.

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McGee, J., Phillips, J., Chan, A. et al. Reduced Rif2 and lack of Mec1 target short telomeres for elongation rather than double-strand break repair. Nat Struct Mol Biol 17, 1438–1445 (2010). https://doi.org/10.1038/nsmb.1947

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