Neddylation promotes ubiquitylation and release of Ku from DNA damage sites
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Authors
Brown, Jessica S
Lukashchuk, Natalia
Sczaniecka-Clift, Matylda
Britton, Sébastien
le, Sage Carlos
Calsou, Patrick
Beli, Petra
Publication Date
2015-04-23Journal Title
Cell Reports
ISSN
2211-1247
Publisher
Elsevier
Volume
11
Pages
704-714
Language
English
Type
Article
Metadata
Show full item recordCitation
Brown, J. S., Lukashchuk, N., Sczaniecka-Clift, M., Britton, S., le, S. C., Calsou, P., Beli, P., et al. (2015). Neddylation promotes ubiquitylation and release of Ku from DNA damage sites. Cell Reports, 11 704-714. https://doi.org/10.1016/j.celrep.2015.03.058
Abstract
The activities of many DNA-repair proteins are
controlled through reversible covalent modification
by ubiquitin and ubiquitin-like molecules. Nonhomologous
end-joining (NHEJ) is the predominant
DNA double-strand break (DSB) repair pathway in
mammalian cells and is initiated by DSB ends being
recognized by the Ku70/Ku80 (Ku) heterodimer. By
using MLN4924, an anti-cancer drug in clinical trials
that specifically inhibits conjugation of the ubiquitin-
like protein, NEDD8, to target proteins, we
demonstrate that NEDD8 accumulation at DNA-damage
sites is a highly dynamic process. In addition, we
show that depleting cells of the NEDD8 E2-conjugating
enzyme, UBE2M, yields ionizing radiation
hypersensitivity and reduced cell survival following
NHEJ. Finally, we demonstrate that neddylation
promotes Ku ubiquitylation after DNA damage and
release of Ku and Ku-associated proteins from damage
sites following repair. These studies provide insights
into how the NHEJ core complex dissociates
from repair sites and highlight its importance for
cell survival following DSB induction.
Sponsorship
We thank Thimo Kurz (University of Dundee, UK) for providing MLN4924 and Kate Dry, Rimma Berlotserkovskaya (S.P.J.’s laboratory), and Eric Lightcap (Takeda Pharmaceuticals) for critical reading of the manuscript. We thank Sylvie Urbe and Michael Clague (University of Liverpool, UK) for providing the GFP-CSN5 plasmid, the Division of Signal Transduction Therapy (University of Dundee, UK) for providing UBE2M and UBE2F plasmids, Matthew Petroski (Sanford-Burnham Medical Research Institute, US) for providing FLAG-UBA3 wild-type (WT) and FLAG-UBA3-A171T constructs, and Nico Dantuma (Karolinska Institute, Sweden) and Changshun Shao (Rutgers University) for providing CUL4A and CUL4B plasmids, respectively. We also thank Nicola Lawrence, Alex Sossick, and Richard Butler (Gurdon Institute, Cambridge, UK) for help with microscopy, Volocity, and Fiji. Research in the S.P.J.’s laboratory is funded by Cancer Research UK programme grant C6/A11224, the European Research Council, and the European Community Seventh Framework Programme grant agreement no. HEALTH-F2-2010-259893 (DDResponse). Core funding is provided by CRUK (C6946/A14492) and the Wellcome Trust (WT092096). S.P.J. receives his salary from the University of Cambridge, UK, supplemented by CRUK. N.L. is funded by CRUK programme grant C6/A11224, J.S.B. is funded by a Wellcome Trust Clinical Fellowship (WT083416), and Y.G. and M.S.-C. are funded by European Research Council grant DDREAM. S.B. was funded by an EMBO long-term fellowship ALTF 93-2010, Cancer Research UK, and a post-doctoral grant from Ligue Nationale Contre le Cancer. P.B. is supported by the Emmy Noether Programme of the German Research Foundation (DFG, BE 5342/1-1).
Funder references
Cancer Research UK (11224)
Wellcome Trust (092096/Z/10/Z)
Cancer Research UK (A14492)
Identifiers
External DOI: https://doi.org/10.1016/j.celrep.2015.03.058
This record's URL: https://www.repository.cam.ac.uk/handle/1810/247966
Rights
Attribution 2.0 UK: England & Wales
Licence URL: http://creativecommons.org/licenses/by/2.0/uk/
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