scholarly journals Studying S-Phase DNA Damage Checkpoints Using the Fission Yeast Schizosaccharomyces pombe

2011 ◽  
pp. 13-21 ◽  
Author(s):  
Nicholas Willis ◽  
Nicholas Rhind
Keyword(s):  
Dna Damage ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
S Phase ◽  
Dna Damage Checkpoints

scholarly journals Mechanism of Caffeine-Induced Checkpoint Override in Fission Yeast

2000 ◽  
Vol 20 (12) ◽  
pp. 4288-4294 ◽  
Author(s):  
Bettina A. Moser ◽  
Jean-Marc Brondello ◽  
Beth Baber-Furnari ◽  
Paul Russell
Keyword(s):  
Dna Damage ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Protein Kinases ◽  
Molecular Target ◽  
S Phase ◽  
Dna Damage Checkpoint ◽  
Mitotic Checkpoints ◽  
Direct Activation

ABSTRACT Mitotic checkpoints restrain the onset of mitosis (M) when DNA is incompletely replicated or damaged. These checkpoints are conserved between the fission yeast Schizosaccharomyces pombe and mammals. In both types of organisms, the methylxanthine caffeine overrides the synthesis (S)-M checkpoint that couples mitosis to completion of DNA S phase. The molecular target of caffeine was sought in fission yeast. Caffeine prevented activation of Cds1 and phosphorylation of Chk1, two protein kinases that enforce the S-M checkpoint triggered by hydroxyurea. Caffeine did not inhibit these kinases in vitro but did inhibit Rad3, a kinase that regulates Cds1 and Chk1. In accordance with this finding, caffeine also overrode the G2-M DNA damage checkpoint that requires Rad3 function. Rad3 coprecipitated with Cds1 expressed at endogenous amounts, a finding that supports the hypothesis that Rad3 is involved in direct activation of Cds1.

Quantitative PCR for detection of DNA damage in mitochondrial DNA of the fission yeast Schizosaccharomyces pombe

2016 ◽  
Vol 127 ◽  
pp. 77-81 ◽  
Author(s):  
Takanori Senoo ◽  
Mayumi Yamanaka ◽  
Atori Nakamura ◽  
Tomoki Terashita ◽  
Shinji Kawano ◽  
...  
Keyword(s):  
Dna Damage ◽  
Mitochondrial Dna ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Quantitative Pcr

scholarly journals Schizosaccharomyces pombe Checkpoint Response to DNA Interstrand Cross-Links

2003 ◽  
Vol 23 (13) ◽  
pp. 4728-4737 ◽  
Author(s):  
Sarah Lambert ◽  
Sarah J. Mason ◽  
Louise J. Barber ◽  
John A. Hartley ◽  
Jackie A. Pearce ◽  
...  
Keyword(s):  
Dna Damage ◽  
Schizosaccharomyces Pombe ◽  
Mammalian Cells ◽  
Excision Repair ◽  
S Phase ◽  
Repair Gene ◽  
Phase Arrest ◽  
S Phase Arrest ◽  
Interstrand Cross Links ◽  
Cross Links

ABSTRACT Drugs that produce covalent interstrand cross-links (ICLs) in DNA remain central to the treatment of cancer, but the cell cycle checkpoints activated by ICLs have received little attention. We have used the fission yeast, Schizosaccharomyces pombe, to elucidate the checkpoint responses to the ICL-inducing anticancer drugs nitrogen mustard and mitomycin C. First we confirmed that the repair pathways acting on ICLs in this yeast are similar to those in the main organisms studied to date (Escherichia coli, budding yeast, and mammalian cells), principally nucleotide excision repair and homologous recombination. We also identified and disrupted the S. pombe homologue of the Saccharomyces cerevisiae SNM1/PSO2 ICL repair gene and found that this activity is required for normal resistance to cross-linking agents, but not other forms of DNA damage. Survival and biochemical analysis indicated a key role for the “checkpoint Rad” family acting through the chk1-dependent DNA damage checkpoint in the ICL response. Rhp9-dependent phosphorylation of Chk1 correlates with G2 arrest following ICL induction. In cells able to bypass the G2 block, a second-cycle (S-phase) arrest was observed. Only a transient activation of the Cds1 DNA replication checkpoint factor occurs following ICL formation in wild-type cells, but this is increased and persists in G2 arrest-deficient mutants. This likely reflects the fraction of cells escaping the G2 damage checkpoint and arresting in the subsequent S phase due to ICL replication blocks. Disruption of cds1 confers increased resistance to ICLs, suggesting that this second-cycle S-phase arrest might be a lethal event.

scholarly journals The Fission Yeast Rad32 (Mre11)-Rad50-Nbs1 Complex Is Required for the S-Phase DNA Damage Checkpoint

2003 ◽  
Vol 23 (18) ◽  
pp. 6564-6573 ◽  
Author(s):  
Charly Chahwan ◽  
Toru M. Nakamura ◽  
Sasirekha Sivakumar ◽  
Paul Russell ◽  
Nicholas Rhind
Keyword(s):  
Dna Damage ◽  
Fission Yeast ◽  
Chromosome Instability ◽  
S Phase ◽  
Comparative Genomic ◽  
Dna Damage Checkpoint ◽  
Checkpoint Signaling ◽  
Damage Repair ◽  
Genomic Approach ◽  
Telomere Regulation

ABSTRACT Mre11, Rad50, and Nbs1 form a conserved heterotrimeric complex that is involved in recombination and DNA damage checkpoints. Mutations in this complex disrupt the S-phase DNA damage checkpoint, the checkpoint which slows replication in response to DNA damage, and cause chromosome instability and cancer in humans. However, how these proteins function and specifically where they act in the checkpoint signaling pathway remain crucial questions. We identified fission yeast Nbs1 by using a comparative genomic approach and showed that the genes for human Nbs1 and fission yeast Nbs1 and that for their budding yeast counterpart, Xrs2, are members of an evolutionarily related but rapidly diverging gene family. Fission yeast Nbs1, Rad32 (the homolog of Mre11), and Rad50 are involved in DNA damage repair, telomere regulation, and the S-phase DNA damage checkpoint. However, they are not required for G2 DNA damage checkpoint. Our results suggest that a complex of Rad32, Rad50, and Nbs1 acts specifically in the S-phase branch of the DNA damage checkpoint and is not involved in general DNA damage recognition or signaling.

scholarly journals Identification of S-phase DNA damage-response targets in fission yeast reveals conservation of damage-response networks

2016 ◽  
Vol 113 (26) ◽  
pp. E3676-E3685 ◽  
Author(s):  
Nicholas A. Willis ◽  
Chunshui Zhou ◽  
Andrew E. H. Elia ◽  
Johanne M. Murray ◽  
Antony M. Carr ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Damage ◽  
Dna Replication ◽  
Fission Yeast ◽  
Dna Damage Response ◽  
Cellular Response ◽  
Genome Stability ◽  
Biological Significance ◽  
S Phase ◽  
Damage Response

The cellular response to DNA damage during S-phase regulates a complicated network of processes, including cell-cycle progression, gene expression, DNA replication kinetics, and DNA repair. In fission yeast, this S-phase DNA damage response (DDR) is coordinated by two protein kinases: Rad3, the ortholog of mammalian ATR, and Cds1, the ortholog of mammalian Chk2. Although several critical downstream targets of Rad3 and Cds1 have been identified, most of their presumed targets are unknown, including the targets responsible for regulating replication kinetics and coordinating replication and repair. To characterize targets of the S-phase DDR, we identified proteins phosphorylated in response to methyl methanesulfonate (MMS)-induced S-phase DNA damage in wild-type, rad3∆, and cds1∆ cells by proteome-wide mass spectrometry. We found a broad range of S-phase–specific DDR targets involved in gene expression, stress response, regulation of mitosis and cytokinesis, and DNA replication and repair. These targets are highly enriched for proteins required for viability in response to MMS, indicating their biological significance. Furthermore, the regulation of these proteins is similar in fission and budding yeast, across 300 My of evolution, demonstrating a deep conservation of S-phase DDR targets and suggesting that these targets may be critical for maintaining genome stability in response to S-phase DNA damage across eukaryotes.

Genotoxicity study with special reference to DNA damage by comet assay in fission yeast, Schizosaccharomyces pombe exposed to drinking water

2008 ◽  
Vol 46 (1) ◽  
pp. 402-407 ◽  
Author(s):  
Pamela Banerjee ◽  
Soumendra N. Talapatra ◽  
Nivedita Mandal ◽  
Geetanjali Sundaram ◽  
Aniruddha Mukhopadhyay ◽  
...  
Keyword(s):  
Dna Damage ◽  
Drinking Water ◽  
Comet Assay ◽  
Special Reference ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast

scholarly journals A Fission Yeast Gene,him1+/dfp1+, Encoding a Regulatory Subunit for Hsk1 Kinase, Plays Essential Roles in S-Phase Initiation as Well as in S-Phase Checkpoint Control and Recovery from DNA Damage

1999 ◽  
Vol 19 (8) ◽  
pp. 5535-5547 ◽  
Author(s):  
Tadayuki Takeda ◽  
Keiko Ogino ◽  
Etsuko Matsui ◽  
Min Kwan Cho ◽  
Hiroyuki Kumagai ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Fission Yeast ◽  
Kinase Activity ◽  
S Phase ◽  
Regulatory Subunit ◽  
Checkpoint Control ◽  
Regulatory Subunits ◽  
Hydroxyurea Treatment ◽  
S Phase Checkpoint

ABSTRACT Saccharomyces cerevisiae CDC7 encodes a serine/threonine kinase required for G1/S transition, and its related kinases are present in fission yeast as well as in higher eukaryotes, including humans. Kinase activity of Cdc7 protein depends on the regulatory subunit, Dbf4, which also interacts with replication origins. We have identified him1+ from two-hybrid screening with Hsk1, a fission yeast homologue of Cdc7 kinase, and showed that it encodes a regulatory subunit of Hsk1. Him1, identical to Dfp1, previously identified as an associated molecule of Hsk1, binds to Hsk1 and stimulates its kinase activity, which phosphorylates both catalytic and regulatory subunits as well as recombinant MCM2 protein in vitro. him1+ is essential for DNA replication in fission yeast cells, and its transcription is cell cycle regulated, increasing at middle M to late G1. The protein level is low at START in G1, increases at the G1/S boundary, and is maintained at a high level throughout S phase. Him1 protein is hyperphosphorylated at G1/S through S during the cell cycle as well as in response to early S-phase arrest induced by nucleotide deprivation. Deletion of one of the motifs conserved in regulatory subunits for Cdc7-related kinases as well as alanine substitution of three serine and threonine residues present in the same motif resulted in a defect in checkpoint regulation normally induced by hydroxyurea treatment. The alanine mutant also showed growth retardation after UV irradiation and the addition of methylmethane sulfonate. In keeping with this result, a database search indicates that him1+ is identical to rad35+ . Our results reveal a novel function of the Cdc7/Dbf4-related kinase complex in S-phase checkpoint control as well as in growth recovery from DNA damage in addition to its predicted essential function in S-phase initiation.

scholarly journals DNA damage checkpoint dynamics drive cell cycle phase transitions

2017 ◽  
Author(s):  
Hui Xiao Chao ◽  
Cere E. Poovey ◽  
Ashley A. Privette ◽  
Gavin D. Grant ◽  
Hui Yan Chao ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Phase Transitions ◽  
Cell Cycle Progression ◽  
S Phase ◽  
Cell Cycle Phase ◽  
Cycle Phase ◽  
Dna Damage Checkpoint ◽  
Cycle Progression ◽  
Dna Damage Checkpoints

ABSTRACTDNA damage checkpoints are cellular mechanisms that protect the integrity of the genome during cell cycle progression. In response to genotoxic stress, these checkpoints halt cell cycle progression until the damage is repaired, allowing cells enough time to recover from damage before resuming normal proliferation. Here, we investigate the temporal dynamics of DNA damage checkpoints in individual proliferating cells by observing cell cycle phase transitions following acute DNA damage. We find that in gap phases (G1 and G2), DNA damage triggers an abrupt halt to cell cycle progression in which the duration of arrest correlates with the severity of damage. However, cells that have already progressed beyond a proposed “commitment point” within a given cell cycle phase readily transition to the next phase, revealing a relaxation of checkpoint stringency during later stages of certain cell cycle phases. In contrast to G1 and G2, cell cycle progression in S phase is significantly less sensitive to DNA damage. Instead of exhibiting a complete halt, we find that increasing DNA damage doses leads to decreased rates of S-phase progression followed by arrest in the subsequent G2. Moreover, these phase-specific differences in DNA damage checkpoint dynamics are associated with corresponding differences in the proportions of irreversibly arrested cells. Thus, the precise timing of DNA damage determines the sensitivity, rate of cell cycle progression, and functional outcomes for damaged cells. These findings should inform our understanding of cell fate decisions after treatment with common cancer therapeutics such as genotoxins or spindle poisons, which often target cells in a specific cell cycle phase.

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