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The Rad50 coiled-coil domain is indispensable for Mre11 complex functions

Nature Structural & Molecular Biology volume 18, pages 1124–1131 (2011)Cite this article

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Abstract

The Mre11 complex (Mre11, Rad50 and Xrs2 in Saccharomyces cerevisiae) influences diverse functions in the DNA damage response. The complex comprises the globular DNA-binding domain and the Rad50 hook domain, which are linked by a long and extended Rad50 coiled-coil domain. In this study, we constructed rad50 alleles encoding truncations of the coiled-coil domain to determine which Mre11 complex functions required the full length of the coils. These mutations abolished telomere maintenance and meiotic double-strand break (DSB) formation, and severely impaired homologous recombination, indicating a requirement for long-range action. Nonhomologous end joining, which is probably mediated by the globular domain of the Mre11 complex, was also severely impaired by alteration of the coiled-coil and hook domains, providing the first evidence of their influence on this process. These data show that functions of Mre11 complex are integrated by the coiled coils of Rad50.

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Figure 1: Rad50 coiled-coil mutants.
Figure 2: Mre11 complex integrity and DNA association are intact without the hook and most of the coiled coil.
Figure 3: rad50 hook and coiled-coil mutants are defective in meiotic DSB formation and telomere maintenance.
Figure 4: The Rad50 hook and coiled-coil domains are required for cell survival in the presence of MMS.
Figure 5: SCR is promoted by the Rad50 hook and coiled-coil domain.
Figure 6: NHEJ in rad50coils.
Figure 7: Distal domain alterations and potential effects on the globular domain.

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Acknowledgements

We are grateful to J. Haber (Brandeis University), F. Uhlmann (London Research Institute), L. Symington (Columbia University), S. Keeney (Memorial Sloan- Kettering Cancer Center), A. Amon (Massachusetts Institute of Technology) and T. Wilson (University of Michigan) for yeast strains, reagents and technical support, to K.-P. Hopfner for structural advice, to G. Bryant and D. Spagna for help with the qPCR, and to current and former members of the Petrini laboratory for insightful comments. We thank S. Keeney for critical reading of the manuscript. This work was supported by GM56888 (J.H.J.P.), PBZH33-112756 and PA0033-117484 from the Swiss National Science Foundation and the Eugen and Elisabeth Schellenberg Foundation (M.H.), BFU2006-05260 and Consolider Ingenio 2010 CSD2007-015 from the Spanish Ministry of Science and Innovation (A.A.) and ES07061 (P.S.).

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Authors and Affiliations

  1. Laboratory of Chromosome Biology, Memorial Sloan‐Kettering Cancer Center, New York, New York, USA

    Marcel Hohl & John H J Petrini

  2. Yale University School of Medicine, New Haven, Connecticut, USA

    Youngho Kwon, Xiaoyu Xue & Patrick Sung

  3. Centro Andaluz de Biología Molecular y Medicina Regenerativa, Universidad de Sevilla, Sevilla, Spain

    Sandra Muñoz Galván, Cristina Tous & Andrés Aguilera

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  1. Marcel Hohl
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  3. Sandra Muñoz Galván
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Contributions

M.H., Y.K., X.X., S.M.G. and C.T. carried out experiments. J.H.J.P. designed research in consultation with P.S. and A.A. J.H.J.P. and M.H. wrote the manuscript.

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Correspondence to John H J Petrini.

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Hohl, M., Kwon, Y., Galván, S. et al. The Rad50 coiled-coil domain is indispensable for Mre11 complex functions. Nat Struct Mol Biol 18, 1124–1131 (2011). https://doi.org/10.1038/nsmb.2116

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