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Tel1ATM and Rad3ATR kinases promote Ccq1-Est1 interaction to maintain telomeres in fission yeast

Abstract

The evolutionarily conserved shelterin complex has been shown to play both positive and negative roles in telomerase regulation in mammals and fission yeast. Although shelterin prevents the checkpoint kinases ATM and ATR from fully activating DNA damage responses at telomeres in mammalian cells, those kinases also promote telomere maintenance. In fission yeast, cells lacking both Tel1 (ATM ortholog) and Rad3 (ATR ortholog) fail to recruit telomerase to telomeres and survive by circularizing chromosomes. However, the critical telomere substrate(s) of Tel1ATM and Rad3ATR was unknown. Here we show that phosphorylation of the shelterin subunit Ccq1 on Thr93, redundantly mediated by Tel1ATM and/or Rad3ATR, is essential for telomerase association with telomeres. In addition, we show that the telomerase subunit Est1 interacts directly with the phosphorylated Thr93 of Ccq1 to ensure telomere maintenance. The shelterin subunits Taz1, Rap1 and Poz1 (previously established inhibitors of telomerase) were also found to negatively regulate Ccq1 phosphorylation. These findings establish Tel1ATM/Rad3ATR-dependent Ccq1 Thr93 phosphorylation as a critical regulator of telomere maintenance in fission yeast.

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Figure 1: Ccq1 interacts with both Tpz1 and Est1.
Figure 2: The phosphopeptide-binding motif of Est1 is important for telomere maintenance.
Figure 3: Thr93 of Ccq1 is essential for telomere maintenance.
Figure 4: Ccq1-Est1 interaction is dependent on phosphorylated Ccq1 Thr93.

References

  1. Lansdorp, P.M. Telomeres and disease. EMBO J. 28, 2532–2540 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Palm, W. & de Lange, T. How shelterin protects mammalian telomeres. Annu. Rev. Genet. 42, 301–334 (2008).

    Article  CAS  PubMed  Google Scholar 

  3. Miyoshi, T., Kanoh, J., Saito, M. & Ishikawa, F. Fission yeast Pot1-Tpp1 protects telomeres and regulates telomere length. Science 320, 1341–1344 (2008).

    Article  CAS  PubMed  Google Scholar 

  4. Flory, M.R., Carson, A.R., Muller, E.G. & Aebersold, R. An SMC-domain protein in fission yeast links telomeres to the meiotic centrosome. Mol. Cell 16, 619–630 (2004).

    Article  CAS  PubMed  Google Scholar 

  5. Tomita, K. & Cooper, J.P. Fission yeast Ccq1 is telomerase recruiter and local checkpoint controller. Genes Dev. 22, 3461–3474 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Denchi, E.L. & de Lange, T. Protection of telomeres through independent control of ATM and ATR by TRF2 and POT1. Nature 448, 1068–1071 (2007).

    Article  CAS  PubMed  Google Scholar 

  7. Verdun, R.E. & Karlseder, J. The DNA damage machinery and homologous recombination pathway act consecutively to protect human telomeres. Cell 127, 709–720 (2006).

    Article  CAS  PubMed  Google Scholar 

  8. Moser, B.A. et al. Differential arrival of leading and lagging strand DNA polymerases at fission yeast telomeres. EMBO J. 28, 810–820 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Sabourin, M. & Zakian, V.A. ATM-like kinases and regulation of telomerase: lessons from yeast and mammals. Trends Cell Biol. 18, 337–346 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Naito, T., Matsuura, A. & Ishikawa, F. Circular chromosome formation in a fission yeast mutant defective in two ATM homologues. Nat. Genet. 20, 203–206 (1998).

    Article  CAS  PubMed  Google Scholar 

  11. Moser, B.A., Subramanian, L., Khair, L., Chang, Y.T. & Nakamura, T.M. Fission yeast Tel1ATM and Rad3ATR promote telomere protection and telomerase recruitment. PLoS Genet. 5, e1000622 (2009).

    Article  PubMed  PubMed Central  Google Scholar 

  12. Beernink, H.T., Miller, K., Deshpande, A., Bucher, P. & Cooper, J.P. Telomere maintenance in fission yeast requires an Est1 ortholog. Curr. Biol. 13, 575–580 (2003).

    Article  CAS  PubMed  Google Scholar 

  13. Fukuhara, N. et al. SMG7 is a 14-3-3-like adaptor in the nonsense-mediated mRNA decay pathway. Mol. Cell 17, 537–547 (2005).

    Article  CAS  PubMed  Google Scholar 

  14. Cooper, J.P., Nimmo, E.R., Allshire, R.C. & Cech, T.R. Regulation of telomere length and function by a Myb-domain protein in fission yeast. Nature 385, 744–747 (1997).

    Article  CAS  PubMed  Google Scholar 

  15. Kanoh, J. & Ishikawa, F. spRap1 and spRif1, recruited to telomeres by Taz1, are essential for telomere function in fission yeast. Curr. Biol. 11, 1624–1630 (2001).

    Article  CAS  PubMed  Google Scholar 

  16. Chikashige, Y. & Hiraoka, Y. Telomere binding of the Rap1 protein is required for meiosis in fission yeast. Curr. Biol. 11, 1618–1623 (2001).

    Article  CAS  PubMed  Google Scholar 

  17. Snow, B.E. et al. Functional conservation of the telomerase protein Est1p in humans. Curr. Biol. 13, 698–704 (2003).

    Article  CAS  PubMed  Google Scholar 

  18. Reichenbach, P. et al. A human homolog of yeast Est1 associates with telomerase and uncaps chromosome ends when overexpressed. Curr. Biol. 13, 568–574 (2003).

    Article  CAS  PubMed  Google Scholar 

  19. Redon, S., Reichenbach, P. & Lingner, J. Protein RNA and protein protein interactions mediate association of human EST1A/SMG6 with telomerase. Nucleic Acids Res. 35, 7011–7022 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Nakamura, T.M., Cooper, J.P. & Cech, T.R. Two modes of survival of fission yeast without telomerase. Science 282, 493–496 (1998).

    Article  CAS  PubMed  Google Scholar 

  21. Matsuoka, S. et al. ATM and ATR substrate analysis reveals extensive protein networks responsive to DNA damage. Science 316, 1160–1166 (2007).

    Article  CAS  PubMed  Google Scholar 

  22. Sugiyama, T. et al. SHREC, an effector complex for heterochromatic transcriptional silencing. Cell 128, 491–504 (2007).

    Article  CAS  PubMed  Google Scholar 

  23. Tomaska, L., Willcox, S., Slezakova, J., Nosek, J. & Griffith, J.D. Taz1 binding to a fission yeast model telomere: formation of telomeric loops and higher order structures. J. Biol. Chem. 279, 50764–50772 (2004).

    Article  CAS  PubMed  Google Scholar 

  24. Miller, K.M., Rog, O. & Cooper, J.P. Semi-conservative DNA replication through telomeres requires Taz1. Nature 440, 824–828 (2006).

    Article  CAS  PubMed  Google Scholar 

  25. Carneiro, T. et al. Telomeres avoid end detection by severing the checkpoint signal transduction pathway. Nature 467, 228–232 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Bianchi, A. & Shore, D. Increased association of telomerase with short telomeres in yeast. Genes Dev. 21, 1726–1730 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Hector, R.E. et al. Tel1p preferentially associates with short telomeres to stimulate their elongation. Mol. Cell 27, 851–858 (2007).

    Article  CAS  PubMed  Google Scholar 

  28. Sabourin, M., Tuzon, C.T. & Zakian, V.A. Telomerase and Tel1p preferentially associate with short telomeres in S. cerevisiae. Mol. Cell 27, 550–561 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Nakamura, T.M., Moser, B.A. & Russell, P. Telomere binding of checkpoint sensor and DNA repair proteins contributes to maintenance of functional fission yeast telomeres. Genetics 161, 1437–1452 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Tseng, S.F., Lin, J.J. & Teng, S.C. The telomerase-recruitment domain of the telomere binding protein Cdc13 is regulated by Mec1p/Tel1p-dependent phosphorylation. Nucleic Acids Res. 34, 6327–6336 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Gao, H. et al. Telomerase recruitment in Saccharomyces cerevisiae is not dependent on Tel1-mediated phosphorylation of Cdc13. Genetics 186, 1147–1159 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Sfeir, A. et al. Mammalian telomeres resemble fragile sites and require TRF1 for efficient replication. Cell 138, 90–103 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Martinez, P. et al. Increased telomere fragility and fusions resulting from TRF1 deficiency lead to degenerative pathologies and increased cancer in mice. Genes Dev. 23, 2060–2075 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Xin, H. et al. TPP1 is a homologue of ciliate TEBP-β and interacts with POT1 to recruit telomerase. Nature 445, 559–562 (2007).

    Article  CAS  PubMed  Google Scholar 

  35. Abreu, E. et al. TIN2-tethered TPP1 recruits human telomerase to telomeres in vivo. Mol. Cell. Biol. 30, 2971–2982 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Tejera, A.M. et al. TPP1 is required for TERT recruitment, telomere elongation during nuclear reprogramming, and normal skin development in mice. Dev. Cell 18, 775–789 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Alfa, C., Fantes, P., Hyams, J., McLoed, M. & Warbrick, E. Experiments with Fission Yeast: A Laboratory Course Manual (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, USA, 1993).

  38. Amberg, D.C., Burke, D.J. & Strathern, J.N. Methods in Yeast Genetics: A Cold Spring Harbor Laboratory Course Manual (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, USA, 2005).

  39. Bähler, J. et al. Heterologous modules for efficient and versatile PCR-based gene targeting in Schizosaccharomyces pombe. Yeast 14, 943–951 (1998).

    Article  PubMed  Google Scholar 

  40. Krawchuk, M.D. & Wahls, W.P. High-efficiency gene targeting in Schizosaccharomyces pombe using a modular, PCR-based approach with long tracts of flanking homology. Yeast 15, 1419–1427 (1999).

    Article  CAS  PubMed  Google Scholar 

  41. Khair, L., Subramanian, L., Moser, B.A. & Nakamura, T.M. Roles of heterochromatin and telomere proteins in regulation of fission yeast telomere recombination and telomerase recruitment. J. Biol. Chem. 285, 5327–5337 (2009).

    Article  PubMed  PubMed Central  Google Scholar 

  42. Nimmo, E.R., Pidoux, A.L., Perry, P.E. & Allshire, R.C. Defective meiosis in telomere-silencing mutants of Schizosaccharomyces pombe. Nature 392, 825–828 (1998).

    Article  CAS  PubMed  Google Scholar 

  43. Lopez-Girona, A. et al. Serine-345 is required for Rad3-dependent phosphorylation and function of checkpoint kinase Chk1 in fission yeast. Proc. Natl. Acad. Sci. USA 98, 11289–11294 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We thank F. Ishikawa (Kyoto University), J.P. Cooper (Cancer Research UK), M.R. Flory (Mendel Biotechnology, Inc.), V.A. Zakian (Princeton University), A.M. Carr (University of Sussex) and P. Russell (The Scripps Research Institute) for sharing published strains and plasmids, L. Khair for her initial efforts to generate yeast strains and reagents used in this study, and P. Baumann (Stowers Institute) for generously sharing his unpublished anti-Ccq1 antibody. We also thank F. Ishikawa for communicating unpublished results. J.K. was supported in part by the Federal Work-Study program. This work was supported by US National Institutes of Health grant GM078253 (T.M.N.).

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B.A.M. designed, performed and analyzed most of the experiments in this study and wrote the paper. Y.-T.C. performed the ChIP experiments illustrated in Figure 3a and initially observed Ccq1 hyperphosphorylation. J.K. assisted B.A.M. in construction of various yeast two-hybrid plasmids. T.M.N. conceived the study, designed and performed experiments, analyzed data and wrote the paper.

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Correspondence to Toru M Nakamura.

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Moser, B., Chang, YT., Kosti, J. et al. Tel1ATM and Rad3ATR kinases promote Ccq1-Est1 interaction to maintain telomeres in fission yeast. Nat Struct Mol Biol 18, 1408–1413 (2011). https://doi.org/10.1038/nsmb.2187

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