Replication Dynamics of the Yeast Genome

Raghuraman, M. K.; Winzeler, Elizabeth A.; Collingwood, David H.; Hunt, Sonia Y.; Wodicka, Lisa; Conway, Andrew; Lockhart, David J.; Davis, Ronald W.; Brewer, Bonita J.; Fangman, Walton L.

doi:10.1126/science.294.5540.115

Cited by 752 publications

(1,006 citation statements)

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“…The distribution of early origins is uneven, with an enrichment in pericentromeric regions, and a depletion in subtelomeric regions. The genome‐wide pattern of ARS activation timing defines a population‐average replication timing program (Raghuraman et al , 2001). To investigate the link between genome organization and replication timing, the read coverage of the Hi‐C libraries was used to compute the replication timing profile of the cell population for each of the time point, and follow their progression through S phase.…”

Section: Resultsmentioning

confidence: 99%

Cohesins and condensins orchestrate the 4D dynamics of yeast chromosomes during the cell cycle

et al. 2017

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Duplication and segregation of chromosomes involves dynamic reorganization of their internal structure by conserved architectural proteins, including the structural maintenance of chromosomes (SMC) complexes cohesin and condensin. Despite active investigation of the roles of these factors, a genome‐wide view of dynamic chromosome architecture at both small and large scale during cell division is still missing. Here, we report the first comprehensive 4D analysis of the higher‐order organization of the Saccharomyces cerevisiae genome throughout the cell cycle and investigate the roles of SMC complexes in controlling structural transitions. During replication, cohesion establishment promotes numerous long‐range intra‐chromosomal contacts and correlates with the individualization of chromosomes, which culminates at metaphase. In anaphase, mitotic chromosomes are abruptly reorganized depending on mechanical forces exerted by the mitotic spindle. Formation of a condensin‐dependent loop bridging the centromere cluster with the rDNA loci suggests that condensin‐mediated forces may also directly facilitate segregation. This work therefore comprehensively recapitulates cell cycle‐dependent chromosome dynamics in a unicellular eukaryote, but also unveils new features of chromosome structural reorganization during highly conserved stages of cell division.

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Section: Resultsmentioning

confidence: 99%

Cohesins and condensins orchestrate the 4D dynamics of yeast chromosomes during the cell cycle

et al. 2017

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“…We hypothesized that break resection would normally be chased and terminated by DNA re‐synthesis which would have a much faster rate than the processing [140 kb/h for conventional replication (Raghuraman et al , 2001), but could be slower during repair synthesis, vs. 4 kb/h for resection (Fishman‐Lobell et al , 1992)]. In fact, by comparing the reconstitution of fragments S2 and L (Fig 6B), we could estimate the rate of dsDNA restoration in srs2 Δ.…”

Section: Resultsmentioning

confidence: 99%

Unloading of homologous recombination factors is required for restoring double‐stranded DNA at damage repair loci

et al. 2017

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Cells use homology‐dependent DNA repair to mend chromosome breaks and restore broken replication forks, thereby ensuring genome stability and cell survival. DNA break repair via homology‐based mechanisms involves nuclease‐dependent DNA end resection, which generates long tracts of single‐stranded DNA required for checkpoint activation and loading of homologous recombination proteins Rad52/51/55/57. While recruitment of the homologous recombination machinery is well characterized, it is not known how its presence at repair loci is coordinated with downstream re‐synthesis of resected DNA. We show that Rad51 inhibits recruitment of proliferating cell nuclear antigen (PCNA), the platform for assembly of the DNA replication machinery, and that unloading of Rad51 by Srs2 helicase is required for efficient PCNA loading and restoration of resected DNA. As a result, srs2Δ mutants are deficient in DNA repair correlating with extensive DNA processing, but this defect in srs2Δ mutants can be suppressed by inactivation of the resection nuclease Exo1. We propose a model in which during re‐synthesis of resected DNA, the replication machinery must catch up with the preceding processing nucleases, in order to close the single‐stranded gap and terminate further resection.

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“…This increase in origin efficiency would, in turn, ensure that random gaps would be efficiently closed by new origin firing. Both the constant number of replication forks and the increasing density of origin firing predicted by this model have been observed in budding yeast and frog embryo extracts 3,9,12,25 . Moreover, in the case of frog embryo extracts, the polymerase-loading and replication-fork protein Cdc45 seems to be limiting for replication, suggesting that, in this system, regulation may involve polymerase recycling 29 .…”

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confidence: 92%

“…Budding yeast has well-defined, site-specific origins, many of which are efficient and fire in as many as 90% of S phases 10,11 . Theses characteristics lead to fairly homogeneous replication kinetics 12 . The fact that budding yeast more closely fits the replicon model has made it much easier to understand replication in budding yeast, and has supported application of the replicon paradigm to eukaryotes in general.…”

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confidence: 99%

DNA replication timing: random thoughts about origin firing

2006

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Regions of metazoan genomes replicate at defined times within S phase. This observation suggests that replication origins fire with a defined timing pattern that remains the same from cycle to cycle. However, an alterative model based on the stochastic firing of origins may also explain replication timing. This model assumes varying origin efficiency instead of a strict origin-timing programme. Here, we discuss the evidence for both models.Models for the organization of eukaryotic DNA replication have to account for two diametrically opposed conceptions of replication. On one side of the spectrum is the replicon paradigm, exemplified by the mechanism of bacterial replication, in which origins are welldefined sites that fire once in each cell cycle, leading to uniform replication that is identical in every cell 1 . At the opposite end of the spectrum is the mechanism of replication in frog and fly embryos, in which replication initiates asynchronously throughout S phase at indiscriminate sites, presumably leading to a different, random pattern of replication in each cell 2,3 . It is clear that neither of these extreme mechanisms reflect the reality of replication in most eukaryotic somatic cells, but it is also clear that both models contain a kernel of truth. So how do we reconcile these apparently competing views of replication? Recent work suggests a way to bridge the gap and to reconcile stochastic origin firing with defined patterns of genome replication.It is useful to first establish which parts of each model generally apply to eukaryotic replication. The replicon paradigm, which serves as the foundation for most textbook models of eukaryotic replication, describes eukaryotic replication reasonably well, with two major exceptions: first, most eukaryotes do not seem to have well-defined, sequence-specific origins4 , 5. Consequently, it has been difficult to identify metazoan replication origins, and in the few cases in which they have been identified, they seem to have no particular sequence specificity. However, too few metazoan origins have been defined to exclude the possibility of origin specific sequences. The question of what constitutes a eukaryotic replication origin remains an active field of research, and as knowledge of the exact location of origins is not necessary for the ideas presented here, we will not pursue it further. Second, eukaryotic origins are inefficient and fire in only a subset of S phases 6 -only a fraction of the origins established in G1 are fired in S phase and it is a different set in each cell. In fission yeast, most origins fire about 30% of the time, whereas in mammals, the well-studied dihydrofolate reductase (DHFR) origins fire approximately 20% of the time 7, 8.In terms of these two exceptions to the replicon paradigm, eukaryotic replication looks more like the example of frog and fly embryos. Frog embryos go to the extreme of establishing origins randomly across the genome, without regard to DNA sequence 2,9 . This is not the case for eukaryotes in general, whi...

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Replication Dynamics of the Yeast Genome

Cited by 752 publications

References 31 publications

Cohesins and condensins orchestrate the 4D dynamics of yeast chromosomes during the cell cycle

Cohesins and condensins orchestrate the 4D dynamics of yeast chromosomes during the cell cycle

Unloading of homologous recombination factors is required for restoring double‐stranded DNA at damage repair loci

DNA replication timing: random thoughts about origin firing

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