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Do replication forks control late origin firing in Saccharomyces cerevisiae?

Ma E, Hyrien O, Goldar A - Nucleic Acids Res. (2011)

Bottom Line: However, measurements of I(t) are based on population averages, which may bias the shape of the I(t) because of imperfect cell synchrony and cell-to-cell variability.The single cell I(t) correlates with fork density in wild-type cells, which is specifically loosened in late S phase in the clb5Δ mutant.Overall, these results suggest that the replication forks emanating from early fired origins facilitate origin firing in later-replicating regions.

View Article: PubMed Central - PubMed

Affiliation: Commissariat à l'Energie Atomique (CEA), iBiTec-S, 91191 Gif-sur-Yvette, France.

ABSTRACT
Recent studies of eukaryotic DNA replication timing profiles suggest that the time-dependent rate of origin firing, I(t), has a universal shape, which ensures a reproducible replication completion time. However, measurements of I(t) are based on population averages, which may bias the shape of the I(t) because of imperfect cell synchrony and cell-to-cell variability. Here, we measure the population-averaged I(t) profile from synchronized Saccharomyces cerevisiae cells using DNA combing and we extract the single-cell I(t) profile using numerical deconvolution. The single cell I(t) and the population-averaged I(t) extracted from DNA combing and replication timing profiles are similar, indicating a genome scale invariance of the replication process, and excluding cell-to-cell variability in replication time as an explanation for the shape of I(t). The single cell I(t) correlates with fork density in wild-type cells, which is specifically loosened in late S phase in the clb5Δ mutant. A previously proposed numerical model that reproduces the wild-type I(t) profile, could also describe the clb5Δ mutant I(t) once modified to incorporate the decline in CDK activity and the looser dependency of initiation on fork density in the absence of Clb5p. Overall, these results suggest that the replication forks emanating from early fired origins facilitate origin firing in later-replicating regions.

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Normalized rate of origin firing versus normalized fork density extracted from DNA combing data in cell populations (A) and in single cells (B). The open black squares (open square) and the open grey circles (open circle) represent the increasing and decreasing part of I(t), respectively. The solid black and dashed grey lines are the spline smoothed curves in panel A, and the five points smoothed curves in panel (B).
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gkr982-F5: Normalized rate of origin firing versus normalized fork density extracted from DNA combing data in cell populations (A) and in single cells (B). The open black squares (open square) and the open grey circles (open circle) represent the increasing and decreasing part of I(t), respectively. The solid black and dashed grey lines are the spline smoothed curves in panel A, and the five points smoothed curves in panel (B).

Mentions: The shape similarity of the I(t) and Nf(t) profiles, which both peak at 30 min in S phase, demonstrate a correlation between fork density and rate of origin firing. This is further exemplified by plotting the normalized rate of origin firing against the normalized replication fork density for the DNA combing experiment (Figure 5A). Removing temporal dispersion as described above strikingly improved the observed correlation (Figure 5B).Figure 5.


Do replication forks control late origin firing in Saccharomyces cerevisiae?

Ma E, Hyrien O, Goldar A - Nucleic Acids Res. (2011)

Normalized rate of origin firing versus normalized fork density extracted from DNA combing data in cell populations (A) and in single cells (B). The open black squares (open square) and the open grey circles (open circle) represent the increasing and decreasing part of I(t), respectively. The solid black and dashed grey lines are the spline smoothed curves in panel A, and the five points smoothed curves in panel (B).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3300028&req=5

gkr982-F5: Normalized rate of origin firing versus normalized fork density extracted from DNA combing data in cell populations (A) and in single cells (B). The open black squares (open square) and the open grey circles (open circle) represent the increasing and decreasing part of I(t), respectively. The solid black and dashed grey lines are the spline smoothed curves in panel A, and the five points smoothed curves in panel (B).
Mentions: The shape similarity of the I(t) and Nf(t) profiles, which both peak at 30 min in S phase, demonstrate a correlation between fork density and rate of origin firing. This is further exemplified by plotting the normalized rate of origin firing against the normalized replication fork density for the DNA combing experiment (Figure 5A). Removing temporal dispersion as described above strikingly improved the observed correlation (Figure 5B).Figure 5.

Bottom Line: However, measurements of I(t) are based on population averages, which may bias the shape of the I(t) because of imperfect cell synchrony and cell-to-cell variability.The single cell I(t) correlates with fork density in wild-type cells, which is specifically loosened in late S phase in the clb5Δ mutant.Overall, these results suggest that the replication forks emanating from early fired origins facilitate origin firing in later-replicating regions.

View Article: PubMed Central - PubMed

Affiliation: Commissariat à l'Energie Atomique (CEA), iBiTec-S, 91191 Gif-sur-Yvette, France.

ABSTRACT
Recent studies of eukaryotic DNA replication timing profiles suggest that the time-dependent rate of origin firing, I(t), has a universal shape, which ensures a reproducible replication completion time. However, measurements of I(t) are based on population averages, which may bias the shape of the I(t) because of imperfect cell synchrony and cell-to-cell variability. Here, we measure the population-averaged I(t) profile from synchronized Saccharomyces cerevisiae cells using DNA combing and we extract the single-cell I(t) profile using numerical deconvolution. The single cell I(t) and the population-averaged I(t) extracted from DNA combing and replication timing profiles are similar, indicating a genome scale invariance of the replication process, and excluding cell-to-cell variability in replication time as an explanation for the shape of I(t). The single cell I(t) correlates with fork density in wild-type cells, which is specifically loosened in late S phase in the clb5Δ mutant. A previously proposed numerical model that reproduces the wild-type I(t) profile, could also describe the clb5Δ mutant I(t) once modified to incorporate the decline in CDK activity and the looser dependency of initiation on fork density in the absence of Clb5p. Overall, these results suggest that the replication forks emanating from early fired origins facilitate origin firing in later-replicating regions.

Show MeSH
Related in: MedlinePlus