Checkpoint inhibition of origin firing prevents inappropriate replication outside of S-phase
- Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, United Kingdom
Abstract
Checkpoints maintain the order of cell cycle events during DNA damage or incomplete replication. How the checkpoint response is tailored to different phases of the cell cycle remains poorly understood. The S-phase checkpoint for example results in the slowing of replication, which in budding yeast occurs by Rad53-dependent inhibition of the initiation factors Sld3 and Dbf4. Despite this, we show here that Rad53 phosphorylates both of these substrates throughout the cell cycle at the same sites as in S-phase, suggesting roles for this pathway beyond S-phase. Indeed, we show that Rad53-dependent inhibition of Sld3 and Dbf4 limits re-replication in G2/M, preventing gene amplification. In addition, we show that inhibition of Sld3 and Dbf4 in G1 prevents premature initiation at all origins at the G1/S transition. This study redefines the scope of the 'S-phase checkpoint' with implications for understanding checkpoint function in cancers that lack cell cycle controls.
Data availability
Sequencing data has been deposited in GEO under the accession code GSE159122 and GSE163571
-
Checkpoint inhibition of origin firing prevents inappropriate replication outside of S-phaseNCBI Gene Expression Omnibus, GSE159122.
-
Checkpoint inhibition of origin firing prevents inappropriate replication outside of S-phaseNCBI Gene Expression Omnibus, GSE163571.
Article and author information
Author details
Miguel Monteiro Santos
Funding
Worlwide Cancer Research (AICR 10-0908)
- Mark C Johnson
- Geylani Can
- Miguel Monteiro Santos
- Diana Alexander
- Philip Zegerman
Wellcome Trust (107056/Z/15/Z)
- Mark C Johnson
- Geylani Can
- Miguel Monteiro Santos
- Diana Alexander
- Philip Zegerman
Biotechnology and Biological Sciences Research Council (BB/M011194/1)
- Miguel Monteiro Santos
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2021, Johnson et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 2,323
- views
-
- 304
- downloads
-
- 7
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Checkpoint inhibition of origin firing prevents inappropriate replication outside of S-phase
eLife 10:e63589.
https://doi.org/10.7554/eLife.63589
Further reading
-
- Chromosomes and Gene Expression
Specialized magnetic beads that bind target proteins to a cryogenic electron microscopy grid make it possible to study the structure of protein complexes from dilute samples.
-
- Chromosomes and Gene Expression
The association between late replication timing and low transcription rates in eukaryotic heterochromatin is well known, yet the specific mechanisms underlying this link remain uncertain. In Saccharomyces cerevisiae, the histone deacetylase Sir2 is required for both transcriptional silencing and late replication at the repetitive ribosomal DNA (rDNA) arrays. We have previously reported that in the absence of SIR2, a de-repressed RNA PolII repositions MCM replicative helicases from their loading site at the ribosomal origin, where they abut well-positioned, high-occupancy nucleosomes, to an adjacent region with lower nucleosome occupancy. By developing a method that can distinguish activation of closely spaced MCM complexes, here we show that the displaced MCMs at rDNA origins have increased firing propensity compared to the nondisplaced MCMs. Furthermore, we found that both activation of the repositioned MCMs and low occupancy of the adjacent nucleosomes critically depend on the chromatin remodeling activity of FUN30. Our study elucidates the mechanism by which Sir2 delays replication timing, and it demonstrates, for the first time, that activation of a specific replication origin in vivo relies on the nucleosome context shaped by a single chromatin remodeler.
