scholarly journals p21 Inhibits Cdk1 in the Absence of Cdk2 to Maintain the G1/S Phase DNA Damage Checkpoint

2008 ◽  
Vol 19 (1) ◽  
pp. 65-77 ◽  
Author(s):  
Ande Satyanarayana ◽  
Mary Beth Hilton ◽  
Philipp Kaldis
Keyword(s):  
Dna Damage ◽  
Liver Cells ◽  
S Phase ◽  
Dna Damage Checkpoint ◽  
Regenerating Liver ◽  
Γ Irradiation ◽  
Embryonic Fibroblasts ◽  
Repair Activity ◽  
Lethal Irradiation

Cdk1 was proposed to compensate for the loss of Cdk2. Here we present evidence that this is possible due to premature translocation of Cdk1 from the cytoplasm to the nucleus in the absence of Cdk2. We also investigated the consequence of loss of Cdk2 on the maintenance of the G1/S DNA damage checkpoint. Cdk2−/− mouse embryonic fibroblasts in vitro as well as regenerating liver cells after partial hepatectomy (PH) in Cdk2−/− mice, arrest promptly at the G1/S checkpoint in response to γ-irradiation due to activation of p53 and p21 inhibiting Cdk1. Furthermore re-entry into S phase after irradiation was delayed in Cdk2−/− cells due to prolonged and impaired DNA repair activity. In addition, Cdk2−/− mice were more sensitive to lethal irradiation compared to wild-type and displayed delayed resumption of DNA replication in regenerating liver cells. Our results suggest that the G1/S DNA damage checkpoint is intact in the absence of Cdk2, but Cdk2 is important for proper repair of the damaged DNA.

scholarly journals The Schizosaccharomyces pombe S-Phase Checkpoint Differentiates Between Different Types of DNA Damage

Genetics ◽  
1998 ◽  
Vol 149 (4) ◽  
pp. 1729-1737 ◽  
Author(s):  
Nicholas Rhind ◽  
Paul Russell
Keyword(s):  
Dna Damage ◽  
Schizosaccharomyces Pombe ◽  
S Phase ◽  
Dna Damage Checkpoint ◽  
Haploid Cell ◽  
Γ Irradiation ◽  
Replication Checkpoint ◽  
Checkpoint Proteins ◽  
Base Modification ◽  
Different Types

Abstract We have identified an S-phase DNA damage checkpoint in Schizosaccharomyces pombe. This checkpoint is dependent on Rad3, the S. pombe homolog of the mammalian ATM/ATR checkpoint proteins, and Cds1. Cds1 had previously been believed to be involved only in the replication checkpoint. The requirement of Cds1 in the DNA damage checkpoint suggests that Cds1 may be a general target of S-phase checkpoints. Unlike other checkpoints, the S. pombe S-phase DNA damage checkpoint discriminates between different types of damage. UV-irradiation, which causes base modification that can be repaired during G1 and S-phase, invokes the checkpoint, while γ-irradiation, which causes double-stranded breaks that cannot be repaired by a haploid cell if induced before replication, does not invoke the checkpoint. Because the same genes are required to respond to UV- and γ-irradiation during G2, this discrimination may represent an active suppression of the γ response during S-phase.

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.

scholarly journals Hemoglobin switching in sheep: characteristics of BFU-E-derived colonies from fetal liver

Blood ◽  
1980 ◽  
Vol 56 (3) ◽  
pp. 495-500
Author(s):  
JE Barker
Keyword(s):  
Stem Cells ◽  
Fetal Liver ◽  
Fetal Hemoglobin ◽  
Liver Cells ◽  
S Phase ◽  
Hemoglobin Switching ◽  
Fetal Liver Cells ◽  
Beta 2 ◽  
Number Of Cells

Two types of erythroid colonies were generated in vitro from sheep fetal liver cells. The first type consisted of single colonies of 8–256 cells that were well hemoglobinized by 4 days; these are thought to originate from CFU-E. The second type consisted of macroscopic colonies composed of several subcolonies that matured between days 3 and 8 in vitro. At maturity, each contained 256 to > 1000 cells that formed a discrete macroscopic cluster. The macroscopic colonies, not previously described in sheep, are thought to be derived from BFU-E. The characteristics of sheep BFU-E were defined and the production of fetal hemoglobin (HbF, alpha 1, gamma 2) and HbC (alpha 2 beta 2) was compared in colonies derived from CFU-E or BFU-E. Bursts developed at erythropoietin (epo) concentrations as low as 0.1 U/ml, although the number observed increased with epo concentration up to 10 U/ml. The number of bursts observed was approximately proportional to the number of cells plated. As shown by thymidine suicide, approximately 50% of both the BFU e and CFU-E were in S-phase when obtained from the fetus. BFU-E were smaller and partially separable from CFU-E after sedimentation at unit gravity. The beta c/gamma synthetic ratio in colonies derived from BFU-E was greater than in CFU-E-derived colonies. These data suggest that the capacity for generation of erythroblasts making HbC is greater in the earlier or more primitive erythroid stem cells in fetal liver.

MDC1 is required for the intra-S-phase DNA damage checkpoint

Nature ◽  
2003 ◽  
Vol 421 (6926) ◽  
pp. 952-956 ◽  
Author(s):  
Michal Goldberg ◽  
Manuel Stucki ◽  
Jacob Falck ◽  
Damien D'Amours ◽  
Dinah Rahman ◽  
...  
Keyword(s):  
Dna Damage ◽  
S Phase ◽  
Dna Damage Checkpoint

scholarly journals Direct Kinase-to-Kinase Signaling Mediated by the FHA Phosphoprotein Recognition Domain of the Dun1 DNA Damage Checkpoint Kinase

2003 ◽  
Vol 23 (4) ◽  
pp. 1441-1452 ◽  
Author(s):  
Vladimir I. Bashkirov ◽  
Elena V. Bashkirova ◽  
Edwin Haghnazari ◽  
Wolf-Dietrich Heyer
Keyword(s):  
Dna Damage ◽  
Target Genes ◽  
Genotoxic Stress ◽  
Dna Damage Checkpoint ◽  
Kinase Signaling ◽  
M Cell ◽  
Fha Domain ◽  
Checkpoint Pathway

ABSTRACT The serine-threonine kinase Dun1 contains a forkhead-associated (FHA) domain and functions in the DNA damage checkpoint pathway of Saccharomyces cerevisiae. It belongs to the Chk2 family of checkpoint kinases, which includes S. cerevisiae Rad53 and Mek1, Schizosaccharomyces pombe Cds1, and human Chk2. Dun1 is required for DNA damage-induced transcription of certain target genes, transient G2/M arrest after DNA damage, and DNA damage-induced phosphorylation of the DNA repair protein Rad55. Here we report that the FHA phosphoprotein recognition domain of Dun1 is required for direct phosphorylation of Dun1 by Rad53 kinase in vitro and in vivo. trans phosphorylation by Rad53 does not require the Dun1 kinase activity and is likely to involve only a transient interaction between the two kinases. The checkpoint functions of Dun1 kinase in DNA damage-induced transcription, G2/M cell cycle arrest, and Rad55 phosphorylation are severely compromised in an FHA domain mutant of Dun1. As a consequence, the Dun1 FHA domain mutant displays enhanced sensitivity to genotoxic stress induced by UV, methyl methanesulfonate, and the replication inhibitor hydroxyurea. We show that the Dun1 FHA domain is critical for direct kinase-to-kinase signaling from Rad53 to Dun1 in the DNA damage checkpoint pathway.

scholarly journals DNA Damage Checkpoint Responses in the S Phase of Synchronized Diploid Human Fibroblasts

2014 ◽  
Vol 91 (1) ◽  
pp. 109-116 ◽  
Author(s):  
Paul D. Chastain ◽  
Bruna P. Brylawski ◽  
Yingchun C. Zhou ◽  
Shangbang Rao ◽  
Haitao Chu ◽  
...  
Keyword(s):  
Dna Damage ◽  
Human Fibroblasts ◽  
S Phase ◽  
Dna Damage Checkpoint

scholarly journals Critical Role for Mouse Hus1 in an S-Phase DNA Damage Cell Cycle Checkpoint

2003 ◽  
Vol 23 (3) ◽  
pp. 791-803 ◽  
Author(s):  
Robert S. Weiss ◽  
Philip Leder ◽  
Cyrus Vaziri
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Synthesis ◽  
Critical Role ◽  
S Phase ◽  
Cell Cycle Checkpoint ◽  
Dna Damage Checkpoint ◽  
Null Mouse ◽  
Cycle Checkpoint ◽  
S Phase Checkpoint

ABSTRACT Mouse Hus1 encodes an evolutionarily conserved DNA damage response protein. In this study we examined how targeted deletion of Hus1 affects cell cycle checkpoint responses to genotoxic stress. Unlike hus1− fission yeast (Schizosaccharomyces pombe) cells, which are defective for the G2/M DNA damage checkpoint, Hus1-null mouse cells did not inappropriately enter mitosis following genotoxin treatment. However, Hus1-deficient cells displayed a striking S-phase DNA damage checkpoint defect. Whereas wild-type cells transiently repressed DNA replication in response to benzo(a)pyrene dihydrodiol epoxide (BPDE), a genotoxin that causes bulky DNA adducts, Hus1-null cells maintained relatively high levels of DNA synthesis following treatment with this agent. However, when treated with DNA strand break-inducing agents such as ionizing radiation (IR), Hus1-deficient cells showed intact S-phase checkpoint responses. Conversely, checkpoint-mediated inhibition of DNA synthesis in response to BPDE did not require NBS1, a component of the IR-responsive S-phase checkpoint pathway. Taken together, these results demonstrate that Hus1 is required specifically for one of two separable mammalian checkpoint pathways that respond to distinct forms of genome damage during S phase.

scholarly journals A Role for Saccharomyces cerevisiae Chk1p in the Response to Replication Blocks

2004 ◽  
Vol 15 (9) ◽  
pp. 4051-4063 ◽  
Author(s):  
Kaila L. Schollaert ◽  
Julie M. Poisson ◽  
Jennifer S. Searle ◽  
Jennifer A. Schwanekamp ◽  
Craig R. Tomlinson ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Damage ◽  
Transcriptional Response ◽  
S Phase ◽  
Yeast Cells ◽  
Dna Damage Checkpoint ◽  
Checkpoint Kinases ◽  
Checkpoint Pathway ◽  
Dna Replication And Repair ◽  
S Phase Checkpoint

Replication blocks and DNA damage incurred during S phase activate the S-phase and intra-S-phase checkpoint responses, respectively, regulated by the Atrp and Chk1p checkpoint kinases in metazoans. In Saccharomyces cerevisiae, these checkpoints are regulated by the Atrp homologue Mec1p and the kinase Rad53p. A conserved role of these checkpoints is to block mitotic progression until DNA replication and repair are completed. In S. cerevisiae, these checkpoints include a transcriptional response regulated by the kinase Dun1p; however, dun1Δ cells are proficient for the S-phase-checkpoint-induced anaphase block. Yeast Chk1p kinase regulates the metaphase-to-anaphase transition in the DNA-damage checkpoint pathway via securin (Pds1p) phosphorylation. However, like Dun1p, yeast Chk1p is not required for the S-phase-checkpoint-induced anaphase block. Here we report that Chk1p has a role in the intra-S-phase checkpoint activated when yeast cells replicate their DNA in the presence of low concentrations of hydroxyurea (HU). Chk1p was modified and Pds1p was transiently phosphorylated in this response. Cells lacking Dun1p were dependent on Chk1p for survival in HU, and chk1Δ dun1Δ cells were defective in the recovery from replication interference caused by transient HU exposure. These studies establish a relationship between the S-phase and DNA-damage checkpoint pathways in S. cerevisiae and suggest that at least in some genetic backgrounds, the Chk1p/securin pathway is required for the recovery from stalled or collapsed replication forks.

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