Abstract:A surveillance mechanism, the S phase checkpoint, blocks progression into mitosis in response to DNA damage and replication stress. Segregation of damaged or incompletely replicated chromosomes results in genomic instability. In humans, the S phase checkpoint has been shown to constitute an anti-cancer barrier. Inhibition of mitotic cyclin dependent kinase (M-CDK) activity by Wee1 kinases is critical to block mitosis in some organisms. However, such mechanism is dispensable in the response to genotoxic stress … Show more
“…1). Deletion of the Mec1/ATR effector Swe1/Wee1 (Palou et al 2015) made no difference.Fig. 1Deletion of Mec1 is not sufficient to allow the segregation of damaged chromosomes.…”
Section: Chromosome Segregation In Response To Genotoxic Stress Is Stmentioning
confidence: 99%
“…In our recent work to better understand how cells block anaphase in response to challenged DNA replication, we showed that the S phase checkpoint prevents chromosome segregation through three independent, redundant, downstream pathways (Palou et al 2015). Mec1/ATR inhibits mitotic Cdk1 activity through downstream effector kinases, Rad53/Chk2 and Swe1/Wee1.…”
Section: Introductionmentioning
confidence: 99%
“…Mec1/ATR inhibits mitotic Cdk1 activity through downstream effector kinases, Rad53/Chk2 and Swe1/Wee1. However, deletion of Rad53/Chk2 and Swe1/Wee1 is not enough to allow cells slip into anaphase in the presence of genotoxic stress, and control on Pds1/cohesin must be abrogated as well (Palou et al 2015). Previous reports place Pds1/securin under the control of the checkpoint effector kinases Chk1 and Rad53/Chk2 (Sanchez et al 1999; Agarwal et al 2003; Kim and Burke 2008).…”
Spontaneous DNA damage poses a continuous threat to genomic integrity. If unchecked, genotoxic insults result in genomic instability, a hallmark of cancer cells. In eukaryotic cells a DNA Damage Response (DDR) detects and responds to genotoxic stress, acting as an anti-cancer barrier in humans. Among other actions, the DDR blocks the segregation of incompletely replicated or damaged chromosomes, thus preventing aneuploidy. In a work aimed at better understanding such S-M control, we recently showed that cells block anaphase through different control pathways. The S phase checkpoint kinase Mec1/ATR inhibits mitotic Cyclin Dependent Kinase activity through effector kinases Swe1/Wee1 and Rad53/Chk2. Cells also stabilize the levels of Pds1/securin to block sister chromatid segregation in response to DNA damage. We show here that Pds1/securin abundance is still secured when the S phase checkpoint response is fully abrogated in mec1/ATR tel1/ATM double null mutants. When such cells are exposed to genotoxic stress, Pds1/securin is stabilized in a spindle assembly checkpoint (SAC) dependent manner. Disruption of the SAC and the S phase checkpoint together, allows chromosome segregation in the presence of DNA damage or replication stress. Our results place the SAC as a part of the DDR, which appears to count on different, independent control layers to preserve genomic integrity when chromosome replication is challenged.
“…1). Deletion of the Mec1/ATR effector Swe1/Wee1 (Palou et al 2015) made no difference.Fig. 1Deletion of Mec1 is not sufficient to allow the segregation of damaged chromosomes.…”
Section: Chromosome Segregation In Response To Genotoxic Stress Is Stmentioning
confidence: 99%
“…In our recent work to better understand how cells block anaphase in response to challenged DNA replication, we showed that the S phase checkpoint prevents chromosome segregation through three independent, redundant, downstream pathways (Palou et al 2015). Mec1/ATR inhibits mitotic Cdk1 activity through downstream effector kinases, Rad53/Chk2 and Swe1/Wee1.…”
Section: Introductionmentioning
confidence: 99%
“…Mec1/ATR inhibits mitotic Cdk1 activity through downstream effector kinases, Rad53/Chk2 and Swe1/Wee1. However, deletion of Rad53/Chk2 and Swe1/Wee1 is not enough to allow cells slip into anaphase in the presence of genotoxic stress, and control on Pds1/cohesin must be abrogated as well (Palou et al 2015). Previous reports place Pds1/securin under the control of the checkpoint effector kinases Chk1 and Rad53/Chk2 (Sanchez et al 1999; Agarwal et al 2003; Kim and Burke 2008).…”
Spontaneous DNA damage poses a continuous threat to genomic integrity. If unchecked, genotoxic insults result in genomic instability, a hallmark of cancer cells. In eukaryotic cells a DNA Damage Response (DDR) detects and responds to genotoxic stress, acting as an anti-cancer barrier in humans. Among other actions, the DDR blocks the segregation of incompletely replicated or damaged chromosomes, thus preventing aneuploidy. In a work aimed at better understanding such S-M control, we recently showed that cells block anaphase through different control pathways. The S phase checkpoint kinase Mec1/ATR inhibits mitotic Cyclin Dependent Kinase activity through effector kinases Swe1/Wee1 and Rad53/Chk2. Cells also stabilize the levels of Pds1/securin to block sister chromatid segregation in response to DNA damage. We show here that Pds1/securin abundance is still secured when the S phase checkpoint response is fully abrogated in mec1/ATR tel1/ATM double null mutants. When such cells are exposed to genotoxic stress, Pds1/securin is stabilized in a spindle assembly checkpoint (SAC) dependent manner. Disruption of the SAC and the S phase checkpoint together, allows chromosome segregation in the presence of DNA damage or replication stress. Our results place the SAC as a part of the DDR, which appears to count on different, independent control layers to preserve genomic integrity when chromosome replication is challenged.
“…Cyclin Dependent Kinase (CDK) activity is under strict checkpoint regulation upon different genotoxic stresses (Hartwell & Weinert 1989; Furnari et al . 1997; Palou et al . 2015).…”
In eukaryotic cells a surveillance mechanism, the S phase checkpoint, detects and responds to insults that challenge chromosomal replication, arresting cell cycle progression and triggering appropriate events to prevent genomic instability. In the budding yeast Saccharomyces cerevisiae, Mec1/ATM/ATR, and its downstream kinase Rad53/Chk2, mediate the response to genotoxic stress. In this study, we place Cip1, a recently identified Cdk1 inhibitor (CKI), under the regulation of Mec1 and Rad53 in response to genotoxic stress. Cip1 accumulates dramatically in a Mec1 and Rad53 dependent manner upon replication stress. This increase requires the activity of MBF, but not the transcriptional activator kinase Dun1. At the protein level, stabilization of replication stress induced Cip1 requires continued de novo protein synthesis. In addition, Cip1 is phosphorylated at an S/TQ motif in a Mec1 dependent manner. Deletion of Cip1 affects proliferation in hydroxyurea containing plates. Significantly, the sensitivity is increased when the dosage of the G1 cyclin CLN2 is increased, compatible to a role of Cip1 as a G1-CDK inhibitor. In all, our results place Cip1 under the S phase checkpoint response to genotoxic stress. Furthermore, Cip1 plays a significant role to preserve viability in response to insults that threaten chromosome replication.
“…Early work in mitotic cells indicated that Swe1-catalyzed Cdk1 phosphorylation regulates the morphogenesis checkpoint (Lew and Reed 1995). However, it is now known that Swe1 is also a component of one of three Mec1-dependent mechanisms that operate in the S phase checkpoint to prevent cell cycle progression into mitosis (Palou et al 2015). …”
In the budding yeast Saccharomyces cerevisiae, unnatural stabilization of the cyclin-dependent kinase inhibitor Sic1 during meiosis can trigger extra rounds of DNA replication. When programmed DNA double-strand breaks (DSBs) are generated but not repaired due to absence of DMC1, a pathway involving the checkpoint gene RAD17 prevents this DNA rereplication. Further genetic analysis has now revealed that prevention of DNA rereplication also requires MEC1, which encodes a protein kinase that serves as a central checkpoint regulator in several pathways including the meiotic recombination checkpoint response. Downstream of MEC1, MEK1 is required through its function to inhibit repair between sister chromatids. By contrast, meiotic recombination checkpoint effectors that regulate gene expression and cyclin-dependent kinase activity are not necessary. Phosphorylation of histone H2A, which is catalyzed by Mec1 and the related Tel1 protein kinase in response to DSBs, and can help coordinate activation of the Rad53 checkpoint protein kinase in the mitotic cell cycle, is required for the full checkpoint response. Phosphorylation sites that are targeted by Rad53 in a mitotic S phase checkpoint response are also involved, based on the behavior of cells containing mutations in the DBF4 and SLD3 DNA replication genes. However, RAD53 does not appear to be required, nor does RAD9, which encodes a mediator of Rad53, consistent with their lack of function in the recombination checkpoint pathway that prevents meiotic progression. While this response is similar to a checkpoint mechanism that inhibits initiation of DNA replication in the mitotic cell cycle, the evidence points to a new variation on DNA replication control.
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