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Updated: Jun 28, 2026
Examination of Mitotic and Meiotic Fission Yeast Nuclear Dynamics by Fluorescence Live-cell Microscopy
Published on: June 24, 2019
Checkpoint-dependent regulation of origin firing and replication fork movement in response to DNA damage in fission
Sanjay Kumar1, Joel A Huberman
1Department of Cancer Biology, Roswell Park Cancer Institute, Elm & Carlton Streets, Buffalo, New York 14263-0001, USA.
Abstract:
To elucidate the checkpoint mechanism responsible for slowing passage through S phase when fission yeast cells are treated with the DNA-damaging agent methyl methanesulfonate (MMS), we carried out two-dimensional gel analyses of replication intermediates in cells synchronized by cdc10 block (in G(1)) followed by release into synchronous S phase. The results indicated that under these conditions early-firing centromeric origins were partially delayed but late-firing telomeric origins were not delayed. Replication intermediates persisted in MMS-treated cells, suggesting that replication fork movement was inhibited. These effects were dependent on the Cds1 checkpoint kinase and were abolished in cells overexpressing the Cdc25 phosphatase, suggesting a role for the Cdc2 cyclin-dependent kinase. We conclude that both partial inhibition of the firing of a subset of origins and inhibition of replication fork movement contribute to the slowing of S phase in MMS-treated fission yeast cells.
Insights
Fission yeast cells slow S phase progression when exposed to DNA damage via methyl methanesulfonate (MMS). This response involves delayed origin firing and inhibited replication fork movement, regulated by the Cds1 checkpoint kinase.
Area of Science:
- Cellular biology
- Molecular genetics
- DNA replication and repair
Background:
- Cell cycle checkpoints are crucial for maintaining genomic stability.
- DNA-damaging agents like methyl methanesulfonate (MMS) trigger cellular responses to prevent mutations.
- Understanding the S phase checkpoint is vital for comprehending DNA damage tolerance.
Purpose of the Study:
- To investigate the molecular mechanisms by which fission yeast cells slow S phase progression upon methyl methanesulfonate (MMS) treatment.
- To identify the key proteins and pathways involved in the DNA damage-induced S phase checkpoint.
Main Methods:
- Synchronized fission yeast cells (cdc10 block) were treated with methyl methanesulfonate (MMS).
- Two-dimensional gel electrophoresis was used to analyze replication intermediates.
- Genetic analysis involving checkpoint-deficient mutants (Cds1) and overexpression strains (Cdc25) was performed.
Main Results:
- Methyl methanesulfonate (MMS) treatment caused a partial delay in the firing of early-replicating centromeric origins but not late-replicating telomeric origins.
- Replication intermediates accumulated, indicating inhibition of replication fork movement.
- These effects were dependent on the Cds1 checkpoint kinase and were influenced by Cdc25 phosphatase levels, implicating Cdc2 kinase.
Conclusions:
- The S phase slowing in response to methyl methanesulfonate (MMS) is a complex process involving both delayed origin firing and inhibited replication fork progression.
- The Cds1 checkpoint kinase plays a critical role in mediating this response.
- The findings highlight the intricate regulation of DNA replication under genotoxic stress in fission yeast.
Related Concept Videos
02:37Restarting Stalled Replication Forks
02:36DNA Damage can Stall the Cell Cycle
02:36DNA Damage Can Stall the Cell Cycle
01:02The DNA Replication Fork
01:02The DNA Replication Fork
02:38S-Cdk Initiates DNA Replication
Two states at the origin of replication
In eukaryotes, the initiation of replication occurs at many sites on the chromosomes, called the origins of replication.