A Link Between ORC-origin Binding Mechanisms and Origin Activation Time Revealed in Budding Yeast

PLoS Genet. 2013;9(9):e1003798. doi: 10.1371/journal.pgen.1003798. Epub 2013 Sep 12.

Abstract

Eukaryotic DNA replication origins are selected in G1-phase when the origin recognition complex (ORC) binds chromosomal positions and triggers molecular events culminating in the initiation of DNA replication (a.k.a. origin firing) during S-phase. Each chromosome uses multiple origins for its duplication, and each origin fires at a characteristic time during S-phase, creating a cell-type specific genome replication pattern relevant to differentiation and genome stability. It is unclear whether ORC-origin interactions are relevant to origin activation time. We applied a novel genome-wide strategy to classify origins in the model eukaryote Saccharomyces cerevisiae based on the types of molecular interactions used for ORC-origin binding. Specifically, origins were classified as DNA-dependent when the strength of ORC-origin binding in vivo could be explained by the affinity of ORC for origin DNA in vitro, and, conversely, as 'chromatin-dependent' when the ORC-DNA interaction in vitro was insufficient to explain the strength of ORC-origin binding in vivo. These two origin classes differed in terms of nucleosome architecture and dependence on origin-flanking sequences in plasmid replication assays, consistent with local features of chromatin promoting ORC binding at 'chromatin-dependent' origins. Finally, the 'chromatin-dependent' class was enriched for origins that fire early in S-phase, while the DNA-dependent class was enriched for later firing origins. Conversely, the latest firing origins showed a positive association with the ORC-origin DNA paradigm for normal levels of ORC binding, whereas the earliest firing origins did not. These data reveal a novel association between ORC-origin binding mechanisms and the regulation of origin activation time.

Publication types

  • Research Support, American Recovery and Reinvestment Act
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • Chromatin / genetics*
  • Chromosomes / genetics
  • DNA / genetics
  • DNA Replication / genetics*
  • DNA-Binding Proteins / genetics
  • G1 Phase / genetics
  • Nucleosomes / genetics
  • Origin Recognition Complex / genetics*
  • Protein Binding
  • Replication Origin / genetics
  • Saccharomyces cerevisiae / genetics*

Substances

  • Chromatin
  • DNA-Binding Proteins
  • Nucleosomes
  • Origin Recognition Complex
  • DNA