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Mus81-Mms4 functions as a single heterodimer to cleave nicked intermediates in recombinational DNA repair.

The formation of crossovers is a fundamental genetic process. The XPF-family endonuclease Mus81-Mms4 (Eme1) contributes significantly to crossing over in eukaryotes. A key question is whether Mus81-Mms4 can process Holliday junctions that contain four uninterrupted strands. Holliday junction cleavage requires the coordination of two active sites, necessitating the assembly of two Mus81-Mms4 heterodimers. Contrary to this expectation, we show that Saccharomyces cerevisiae Mus81-Mms4 exists as a single heterodimer both in solution and when bound to DNA substrates in vitro. Consistently, immunoprecipitation experiments demonstrate that Mus81-Mms4 does not multimerize in vivo. Moreover, chromatin-bound Mus81-Mms4 does not detectably form higher-order multimers. We show that Cdc5 kinase activates Mus81-Mms4 nuclease activity on 3' flaps and Holliday junctions in vitro but that activation does not induce a preference for Holliday junctions and does not induce multimerization of the Mus81-Mms4 heterodimer. These data support a model in which Mus81-Mms4 cleaves nicked recombination intermediates such as displacement loops (D-loops), nicked Holliday junctions, or 3' flaps but not intact Holliday junctions with four uninterrupted strands. We infer that Mus81-dependent crossing over occurs in a noncanonical manner that does not involve the coordinated cleavage of classic Holliday junctions.

Pubmed ID: 22645308

Authors

  • Schwartz EK
  • Wright WD
  • Ehmsen KT
  • Evans JE
  • Stahlberg H
  • Heyer WD

Journal

Molecular and cellular biology

Publication Data

August 19, 2012

Associated Grants

  • Agency: NIGMS NIH HHS, Id: GM58015
  • Agency: NIGMS NIH HHS, Id: U54GM74929

Mesh Terms

  • Cell Cycle Proteins
  • Chromatin
  • DNA
  • DNA, Cruciform
  • DNA-Binding Proteins
  • Endonucleases
  • Flap Endonucleases
  • Protein Multimerization
  • Protein-Serine-Threonine Kinases
  • Recombination, Genetic
  • Recombinational DNA Repair
  • Saccharomyces cerevisiae
  • Saccharomyces cerevisiae Proteins