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Mol Cell. 2016 Oct 6;64(1):189-198. doi: 10.1016/j.molcel.2016.08.037.

VCP/p97 Extracts Sterically Trapped Ku70/80 Rings from DNA in Double-Strand Break Repair.

Author information

1
Molecular Biology I, Faculty of Biology, University of Duisburg-Essen, 45117 Essen, Germany.
2
ICCE, Faculty of Biology, University of Duisburg-Essen, 45117 Essen, Germany.
3
Institute of Medical Radiation Biology, Medical School, University of Duisburg-Essen, 45122 Essen, Germany.
4
Chemical Proteomics, Faculty of Biology, University of Duisburg-Essen, 45117 Essen, Germany.
5
Laboratory of Chromosome and Cell Biology, The Rockefeller University, New York, NY 10065, USA.
6
Molecular Biology I, Faculty of Biology, University of Duisburg-Essen, 45117 Essen, Germany. Electronic address: hemmo.meyer@uni-due.de.

Abstract

During DNA double-strand break (DSB) repair, the ring-shaped Ku70/80 complex becomes trapped on DNA and needs to be actively extracted, but it has remained unclear what provides the required energy. By means of reconstitution of DSB repair on beads, we demonstrate here that DNA-locked Ku rings are released by the AAA-ATPase p97. To achieve this, p97 requires ATP hydrolysis, cooperates with the Ufd1-Npl4 ubiquitin-adaptor complex, and specifically targets Ku80 that is modified by K48-linked ubiquitin chains. In U2OS cells, chemical inhibition of p97 or siRNA-mediated depletion of p97 or its adapters impairs Ku80 removal after non-homologous end joining of DSBs. Moreover, this inhibition attenuates early steps in homologous recombination, consistent with p97-driven Ku release also affecting repair pathway choice. Thus, our data answer a central question regarding regulation of Ku in DSB repair and illustrate the ability of p97 to segregate even tightly bound protein complexes for release from DNA.

KEYWORDS:

DNA repair; Ku; Ku80; NHEJ; VCP; chromatin; double strand break; homologous recombination; p97

PMID:
27716483
PMCID:
PMC5161236
DOI:
10.1016/j.molcel.2016.08.037
[Indexed for MEDLINE]
Free PMC Article

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