scholarly journals Mcm2 Is a Direct Substrate of ATM and ATR during DNA Damage and DNA Replication Checkpoint Responses

2004 ◽  
Vol 279 (51) ◽  
pp. 53353-53364 ◽  
Author(s):  
Hae Yong Yoo ◽  
Anna Shevchenko ◽  
Andrej Shevchenko ◽  
William G. Dunphy
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Replication Checkpoint ◽  
Dna Replication Checkpoint

Overlapping Roles of the Spindle Assembly and DNA Damage Checkpoints in the Cell-Cycle Response to Altered Chromosomes in Saccharomyces cerevisiae

Genetics ◽  
2002 ◽  
Vol 161 (2) ◽  
pp. 521-534
Author(s):  
Peter M Garber ◽  
Jasper Rine
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Spindle Checkpoint ◽  
Dna Lesions ◽  
Replication Proteins ◽  
Replication Checkpoint ◽  
Dna Damage Checkpoints ◽  
Dna Replication Checkpoint ◽  
Mcm Proteins ◽  
Stalled Replication Forks

Abstract The MAD2-dependent spindle checkpoint blocks anaphase until all chromosomes have achieved successful bipolar attachment to the mitotic spindle. The DNA damage and DNA replication checkpoints block anaphase in response to DNA lesions that may include single-stranded DNA and stalled replication forks. Many of the same conditions that activate the DNA damage and DNA replication checkpoints also activated the spindle checkpoint. The mad2Δ mutation partially relieved the arrest responses of cells to mutations affecting the replication proteins Mcm3p and Pol1p. Thus a previously unrecognized aspect of spindle checkpoint function may be to protect cells from defects in DNA replication. Furthermore, in cells lacking either the DNA damage or the DNA replication checkpoints, the spindle checkpoint contributed to the arrest responses of cells to the DNA-damaging agent methyl methanesulfonate, the replication inhibitor hydroxyurea, and mutations affecting Mcm2p and Orc2p. Thus the spindle checkpoint was sensitive to a wider range of chromosomal perturbations than previously recognized. Finally, the DNA replication checkpoint did not contribute to the arrests of cells in response to mutations affecting ORC, Mcm proteins, or DNA polymerase δ. Thus the specificity of this checkpoint may be more limited than previously recognized.

A method for assaying the sensitivity of Drosophila replication checkpoint mutants to anti-cancer and DNA-damaging drugs.

Genome ◽  
2002 ◽  
Vol 45 (5) ◽  
pp. 881-889 ◽  
Author(s):  
Colleen M Radcliffe ◽  
Elizabeth A Silva ◽  
Shelagh D Campbell
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Cell Cycle Progression ◽  
Tumor Formation ◽  
Cycle Progression ◽  
Replication Checkpoint ◽  
Anti Cancer ◽  
Dna Replication Checkpoint ◽  
Checkpoint Genes ◽  
Regulate Cell Cycle Progression

In multi-cellular organisms, failure to properly regulate cell-cycle progression can result in inappropriate cell death or uncontrolled cell division leading to tumor formation. To guard against such events, conserved regulatory mechanisms called "checkpoints" block progression into mitosis in response to DNA damage and incomplete replication, as well as in response to other signals. Checkpoint mutants in organisms as diverse as yeast and humans are sensitive to various chemical agents that inhibit DNA replication or cause DNA damage. This phenomenon is the primary rationale for chemotherapy, which uses drugs that preferentially target tumor cells with compromised checkpoints. In this study, we demonstrate the use of Drosophila checkpoint mutants as a system for assaying the effects of various DNA-damaging and anti-cancer agents in a developing multicellular organism. Dwee1, grp and mei-41 are genes that encode kinases that function in the DNA replication checkpoint. We tested zygotic mutants of each gene for sensitivity to the DNA replication inhibitor hydroxyurea (HU), methyl methanosulfonate (MMS), ara-C, cisplatin, and the oxygen radical generating compound paraquat. The mutants show distinct differences in their sensitivity to each of the drugs tested, suggesting an underlying complexity in the responses of individual checkpoint genes to genotoxic stress.Key words: hydroxyurea (HU), ara-C, cisplatin, methyl methane sulfonate (MMS), paraquat.

scholarly journals The DNA damage and the DNA replication checkpoints converge at the MBF transcription factor

2013 ◽  
Vol 24 (21) ◽  
pp. 3350-3357 ◽  
Author(s):  
Tsvetomira Ivanova ◽  
Isabel Alves-Rodrigues ◽  
Blanca Gómez-Escoda ◽  
Chaitali Dutta ◽  
James A. DeCaprio ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Fission Yeast ◽  
Yeast Cells ◽  
Replication Checkpoint ◽  
Dna Damaging Agents ◽  
Dna Replication Checkpoint ◽  
Transcriptional Complex ◽  
Carboxy Terminal ◽  
Cycle Regulation

In fission yeast cells, Cds1 is the effector kinase of the DNA replication checkpoint. We previously showed that when the DNA replication checkpoint is activated, the repressor Yox1 is phosphorylated and inactivated by Cds1, resulting in activation of MluI-binding factor (MBF)–dependent transcription. This is essential to reinitiate DNA synthesis and for correct G1-to-S transition. Here we show that Cdc10, which is an essential part of the MBF core, is the target of the DNA damage checkpoint. When fission yeast cells are treated with DNA-damaging agents, Chk1 is activated and phosphorylates Cdc10 at its carboxy-terminal domain. This modification is responsible for the repression of MBF-dependent transcription through induced release of MBF from chromatin. This inactivation of MBF is important for survival of cells challenged with DNA-damaging agents. Thus Yox1 and Cdc10 couple normal cell cycle regulation in unperturbed conditions and the DNA replication and DNA damage checkpoints into a single transcriptional complex.

scholarly journals Increased Genome Instability and Telomere Length in the elg1-Deficient Saccharomyces cerevisiae Mutant Are Regulated by S-Phase Checkpoints

2004 ◽  
Vol 3 (6) ◽  
pp. 1557-1566 ◽  
Author(s):  
Soma Banerjee ◽  
Kyungjae Myung
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Damage ◽  
Dna Replication ◽  
Telomere Length ◽  
Genome Instability ◽  
Dna Recombination ◽  
Cell Cycle Checkpoints ◽  
Telomere Maintenance ◽  
Replication Checkpoint ◽  
Dna Replication Checkpoint

ABSTRACT Gross chromosomal rearrangements (GCRs) are frequently observed in cancer cells. Abnormalities in different DNA metabolism including DNA replication, cell cycle checkpoints, chromatin remodeling, telomere maintenance, and DNA recombination and repair cause GCRs in Saccharomyces cerevisiae. Recently, we used genome-wide screening to identify several genes the deletion of which increases GCRs in S. cerevisiae. Elg1, which was discovered during this screening, functions in DNA replication by participating in an alternative replication factor complex. Here we further characterize the GCR suppression mechanisms observed in the elg1Δ mutant strain in conjunction with the telomere maintenance role of Elg1. The elg1Δ mutation enhanced spontaneous DNA damage and resulted in GCR formation. However, DNA damage due to inactivation of Elg1 activates the intra-S checkpoints, which suppress further GCR formation. The intra-S checkpoints activated by the elg1Δ mutation also suppress GCR formation in strains defective in the DNA replication checkpoint. Lastly, the elg1Δ mutation increases telomere size independently of other previously known telomere maintenance proteins such as the telomerase inhibitor Pif1 or the telomere size regulator Rif1. The increase in telomere length caused by the elg1Δ mutation was suppressed by a defect in the DNA replication checkpoint, which suggests that DNA replication surveillance by Dpb11-Mec1/Tel1-Dun1 also has an important role in telomere length regulation.

scholarly journals The Drosophila chk2 Gene loki Is Essential for Embryonic DNA Double-Strand-Break Checkpoints Induced in S Phase or G2

Genetics ◽  
2003 ◽  
Vol 163 (3) ◽  
pp. 973-982 ◽  
Author(s):  
Nisrine Masrouha ◽  
Long Yang ◽  
Sirine Hijal ◽  
Stéphane Larochelle ◽  
Beat Suter
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Replication ◽  
Strand Break ◽  
Cell Cycle Checkpoints ◽  
Phylogenetic Study ◽  
Embryonic Cells ◽  
Replication Checkpoint ◽  
C Elegans ◽  
Dna Replication Checkpoint

Abstract Cell cycle checkpoints are signal transduction pathways that control the order and timing of cell cycle transitions, ensuring that critical events are completed before the occurrence of the next cell cycle transition. The Chk2 family of kinases is known to play a central role in mediating the cellular responses to DNA damage or DNA replication blocks in various organisms. Here we show through a phylogenetic study that the Drosophila melanogaster serine/threonine kinase Loki is the homolog of the yeast Mek1p, Rad53p, Dun1p, and Cds1 proteins as well as the human Chk2. Functional analyses allowed us to conclude that, in flies, chk2 is involved in monitoring double-strand breaks (DSBs) caused by irradiation during S and G2 phases. In this process it plays an essential role in inducing a cell cycle arrest in embryonic cells. Our results also show that, in contrast to C. elegans chk2, Drosophila chk2 is not essential for normal meiosis and recombination, and it also appears to be dispensable for the MMS-induced DNA damage checkpoint and the HU-induced DNA replication checkpoint during larval development. In addition, Drosophila chk2 does not act at the same cell cycle phases as its yeast homologs, but seems rather to be involved in a pathway similar to the mammalian one, which involves signaling through the ATM/Chk2 pathway in response to genotoxic insults. As mutations in human chk2 were linked to several cancers, these similarities point to the usefulness of the Drosophila model system.

scholarly journals The Direct Binding of Mrc1, a Checkpoint Mediator, to Mcm6, a Replication Helicase, Is Essential for the Replication Checkpoint against Methyl Methanesulfonate-Induced Stress

2009 ◽  
Vol 29 (18) ◽  
pp. 5008-5019 ◽  
Author(s):  
Makiko Komata ◽  
Masashige Bando ◽  
Hiroyuki Araki ◽  
Katsuhiko Shirahige
Keyword(s):  
Amino Acid ◽  
Dna Replication ◽  
Terminal Region ◽  
Methyl Methanesulfonate ◽  
Checkpoint Activation ◽  
Replication Checkpoint ◽  
Hydroxyurea Treatment ◽  
Dna Replication Checkpoint ◽  
Direct Binding

ABSTRACT Mrc1 plays a role in mediating the DNA replication checkpoint. We surveyed replication elongation proteins that interact directly with Mrc1 and identified a replicative helicase, Mcm6, as a specific Mrc1-binding protein. The central portion of Mrc1, containing a conserved coiled-coil region, was found to be essential for interaction with the 168-amino-acid C-terminal region of Mcm6, and introduction of two amino acid substitutions in this C-terminal region abolished the interaction with Mrc1 in vivo. An mcm6 mutant bearing these substitutions showed a severe defect in DNA replication checkpoint activation in response to stress caused by methyl methanesulfonate. Interestingly, the mutant did not show any defect in DNA replication checkpoint activation in response to hydroxyurea treatment. The phenotype of the mcm6 mutant was suppressed when the mutant protein was physically fused with Mrc1. These results strongly suggest for the first time that an Mcm helicase acts as a checkpoint sensor for methyl methanesulfonate-induced DNA damage through direct binding to the replication checkpoint mediator Mrc1.

scholarly journals Stockpiling of Cdc25 during a DNA replication checkpoint arrest in Schizosaccharomyces pombe.

1996 ◽  
Vol 16 (1) ◽  
pp. 86-93 ◽  
Author(s):  
R Kovelman ◽  
P Russell
Keyword(s):  
Dna Replication ◽  
Schizosaccharomyces Pombe ◽  
Replication Checkpoint ◽  
Activation Assay ◽  
M Phase ◽  
Dna Replication Checkpoint ◽  
High Level ◽  
Tyrosyl Phosphorylation ◽  
In Cells

The DNA replication checkpoint couples the onset of mitosis with the completion of S phase. It is clear that in the fission yeast Schizosaccharomyces pombe, operation of this checkpoint requires maintenance of the inhibitory tyrosyl phosphorylation of Cdc2. Cdc25 phosphatase induces mitosis by dephosphorylating tyrosine 15 of Cdc2. In this report, Cdc25 is shown to accumulate to a very high level in cells arrested in S. This shows that mechanisms which modulate the abundance of Cdc25 are unconnected to the DNA replication checkpoint. Using a Cdc2/cyclin B activation assay, we found that Cdc25 activity increased approximately 10-fold during transit through M phase. Cdc25 was activated by phosphorylations that were dependent on Cdc2 activity in vivo. Cdc25 activation was suppressed in cells arrested in G1 and S. However, Cdc25 was more highly modified and appeared to be somewhat more active in S than in G1. This finding might be connected to the fact that progression from G1 to S increases the likelihood that constitutive Cdc25 overproduction will cause inappropriate mitosis.

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