A Link between ORC-origin binding mechanisms and origin activation time revealed in budding yeast.
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.
Department of Biomolecular Chemistry, School of Medicine and Public Health, University of Wisconsin-Madison, Madison, Wisconsin, United States of America ; Program in Cellular and Molecular Biology, College of Agriculture and Life Sciences, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
Timothy
T
Hoggard
Erika
E
Shor
Carolin A
CA
Müller
Conrad A
CA
Nieduszynski
Catherine A
CA
Fox
D001665:Binding Sites; D002843:Chromatin / Q000235:genetics*; D002875:Chromosomes / Q000235:genetics; D004247:DNA / Q000235:genetics; D004261:DNA Replication / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics; D016193:G1 Phase / Q000235:genetics; D009707:Nucleosomes / Q000235:genetics; D051738:Origin Recognition Complex / Q000235:genetics*; D011485:Protein Binding; D018741:Replication Origin / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D002843:Chromatin; D004268:DNA-Binding Proteins; D009707:Nucleosomes; D051738:Origin Recognition Complex; D004247:DNA