Interplay between Ku and replication protein A in the restriction of Exo1-mediated DNA break end resection

Danielle S. Krasner, James M. Daley, Patrick Sung, Hengyao Niu

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

DNA double-strand breaks can be eliminated via non-homologous end joining or homologous recombination. Non-homologous end joining is initiated by the association of Ku with DNA ends. In contrast, homologous recombination entails nucleolytic resection of the 5′-strands, forming 3′-ssDNA tails that become coated with replication protein A (RPA). Ku restricts end access by the resection nuclease Exo1. It is unclear how partial resection might affect Ku engagement and Exo1 restriction. Here, we addressed these questions in a reconstituted system with yeast proteins. With blunt-ended DNA, Ku protected against Exo1 in a manner that required its DNA end-binding activity. Despite binding poorly to ssDNA, Ku could nonetheless engage a 5′-recessed DNA end with a 40-nucleotide (nt) ssDNA overhang, where it localized to the ssDNA-dsDNA junction and efficiently blocked resection by Exo1. Interestingly, RPA could exclude Ku from a partially resected structure with a 22-nt ssDNA tail and thus restored processing by Exo1. However, at a 40-nt tail, Ku remained stably associated at the ssDNA-dsDNA junction, and RPA simultaneously engaged the ssDNA region. We discuss a model in which the dynamic equilibrium between Ku and RPA binding to a partially resected DNA end influences the timing and efficiency of the resection process.

Original languageEnglish (US)
Pages (from-to)18806-18816
Number of pages11
JournalJournal of Biological Chemistry
Volume290
Issue number30
DOIs
StatePublished - Jul 24 2015

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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