Ino80 chromatin remodeling complex promotes recovery of stalled replication forks.
BACKGROUND
Chromatin remodeling complexes facilitate the access of enzymes that mediate transcription, replication or repair of DNA by modulating nucleosome position and/or composition. Ino80 is the DNA-dependent Snf2-like ATPase subunit of a complex whose nucleosome remodeling activity requires actin-related proteins, Arp4, Arp5 and Arp8, as well as two RuvB-like DNA helicase subunits. Budding yeast mutants deficient for Ino80 function are not only hypersensitive to reagents that induce DNA double-strand breaks, but also to those that impair replication fork progression.


RESULTS
To understand why ino80 mutants are sensitive to agents that perturb DNA replication, we used chromatin immunoprecipitation to map the binding sites of the INO80 chromatin-remodeling complex on four budding yeast chromosomes. We found that Ino80 and Arp5 binding sites coincide with origins of DNA replication and tRNA genes. In addition, Ino80 was bound at 67% of the promoters of genes that are sensitive to ino80 mutation. When replication forks were arrested near origins in the presence of hydroxyurea (HU), the amount of INO80 complex at stalled forks and at unfired origins increased selectively. Importantly, the resumption of DNA replication after release from a HU block was impaired in ino80 mutants. These cells accumulated double-strand breaks as they attempted to restart replication. Consistently, ino80-deficient cells, although proficient for checkpoint activation, delay recovery from the checkpoint response.


CONCLUSIONS
The INO80 chromatin remodeling complex is enriched at stalled replication forks, where it promotes the resumption of replication upon recovery from fork arrest.
Shimada
Kenji
K
Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, CH-4058 Basel, Switzerland.
Oma
Yukako
Y
Schleker
Thomas
T
Kugou
Kazuto
K
Ohta
Kunihiro
K
Harata
Masahiko
M
Gasser
Susan M
SM
D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins / Q000378:metabolism; D064447:Checkpoint Kinase 2; D042002:Chromatin Assembly and Disassembly / Q000502:physiology*; D053903:DNA Breaks, Double-Stranded; D004257:DNA Polymerase II / Q000378:metabolism; D004261:DNA Replication / Q000502:physiology*; D006918:Hydroxyurea / Q000378:metabolism; D009154:Mutation; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D012343:RNA, Transfer / Q000235:genetics; D051721:Rad52 DNA Repair and Recombination Protein / Q000378:metabolism; D018741:Replication Origin / Q000502:physiology*; D016196:S Phase / Q000502:physiology; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; C483404:INO80 complex, S cerevisiae; C440014:LCD1 protein, S cerevisiae; D010750:Phosphoproteins; C497197:RAD52 protein, S cerevisiae; D051721:Rad52 DNA Repair and Recombination Protein; D029701:Saccharomyces cerevisiae Proteins; D012343:RNA, Transfer; D064447:Checkpoint Kinase 2; D017346:Protein Serine-Threonine Kinases; C067022:RAD53 protein, S cerevisiae; D004257:DNA Polymerase II; D006918:Hydroxyurea