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N Rhind

Publications and source records attributed to N Rhind.

12 recordsLinked to original sources
PubMed2001

Roles of the mitotic inhibitors Wee1 and Mik1 in the G(2) DNA damage and replication checkpoints.

The G(2) DNA damage and DNA replication checkpoints in many organisms act through the inhibitory phosphorylation of Cdc2 on tyrosine-15. This phosphorylation is catalyzed by the Wee1/Mik1 family of kinases. However, the in vivo role of these kinases in checkpoint regulation has been unclear. We show that, in the fission yeast Schizosaccharomyces pombe, Mik1 is a target of both checkpoints and that the regulation of Mik1 is, on its own, sufficient to delay mitosis in response to the checkpoints. Mik1 appears to have two roles in the DNA damage checkpoint; one in the establishment of the checkpoint and another in its maintenance. In contrast, Wee1 does not appear to be involved in the establishment of either checkpoint.

Alleles↗
PubMed2000

Regulation of mitotic inhibitor Mik1 helps to enforce the DNA damage checkpoint.

The protein kinase Chk1 enforces the DNA damage checkpoint. This checkpoint delays mitosis until damaged DNA is repaired. Chk1 regulates the activity and localization of Cdc25, the tyrosine phosphatase that activates the cdk Cdc2. Here we report that Mik1, a tyrosine kinase that inhibits Cdc2, is positively regulated by the DNA damage checkpoint. Mik1 is required for checkpoint response in strains that lack Cdc25. Long-term DNA damage checkpoint arrest fails in Deltamik1 cells. DNA damage increases Mik1 abundance in a Chk1-dependent manner. Ubiquitinated Mik1 accumulates in a proteasome mutant, which indicates that Mik1 normally has a short half-life. Thus, the DNA damage checkpoint might regulate Mik1 degradation. Mik1 protein and mRNA oscillate during the unperturbed cell cycle, with peak amounts detected around S phase. These data indicate that regulation of Mik1 abundance helps to couple mitotic onset to the completion of DNA replication and repair. Coordinated negative regulation of Cdc25 and positive regulation of Mik1 ensure the effective operation of the DNA damage checkpoint.

CDC2 Protein Kinase↗
PubMed2000

Chk1 and Cds1: linchpins of the DNA damage and replication checkpoint pathways.

Recent work on the mechanisms of DNA damage and replication cell cycle checkpoints has revealed great similarity between the checkpoint pathways of organisms as diverse as yeasts, flies and humans. However, there are differences in the ways these organisms regulate their cell cycles. To connect the conserved checkpoint pathways with various cell cycle targets requires an adaptable link that can target different cell cycle components in different organisms. The Chk1 and Cds1 protein kinases, downstream effectors in the checkpoint pathways, seem to play just such roles. Perhaps more surprisingly, the two kinases not only have different targets in different organisms but also seem to respond to different signals in different organisms. So, whereas in fission yeast Chk1 is required for the DNA damage checkpoint and Cds1 is specifically involved in the replication checkpoint, their roles seem to be shuffled in metazoans.

Animals↗
PubMed1998

Mitotic DNA damage and replication checkpoints in yeast.

Studies of the genetics of G2/M checkpoints in budding and fission yeasts have produced many of the defining concepts of checkpoint biology. Recent progress in the biochemistry of the checkpoint gene products is adding a mechanistic understanding to our models and identifying the components of the normal cell cycle machinery that are targeted by checkpoints.

DNA Damage↗
PubMed1998

The Schizosaccharomyces pombe S-phase checkpoint differentiates between different types of DNA damage.

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 gamma-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 gamma-irradiation during G2, this discrimination may represent an active suppression of the gamma response during S-phase.

Adenosine Triphosphatases↗
PubMed1998

Tyrosine phosphorylation of cdc2 is required for the replication checkpoint in Schizosaccharomyces pombe.

The DNA replication checkpoint inhibits mitosis in cells that are unable to replicate their DNA, as when nucleotide biosynthesis is inhibited by hydroxyurea. In the fission yeast Schizosaccharomyces pombe, genetic evidence suggests that this checkpoint involves the inhibition of Cdc2 activity through the phosphorylation of tyrosine-15. On the contrary, a recent biochemical study indicated that Cdc2 is in an activated state during a replication checkpoint, suggesting that phosphorylation of Cdc2 on tyrosine-15 is not part of the replication checkpoint mechanism. We have undertaken biochemical and genetic studies to resolve this controversy. We report that the DNA replication checkpoint in S. pombe is abrogated in cells that carry the allele cdc2-Y15F, expressing an unphosphorylatable form of Cdc2. Furthermore, Cdc2 isolated from replication checkpoint-arrested cells can be activated in vitro by Cdc25, the tyrosine phosphatase responsible for dephosphorylating Cdc2 in vivo, to the same extent as Cdc2 isolated from cdc25ts-blocked cells, indicating that hydroxyurea treatment causes Cdc2 activity to be maintained at a low level that is insufficient to induce mitosis. These studies show that inhibitory tyrosine-15 phosphorylation of Cdc2 is essential for the DNA replication checkpoint and suggests that Cdc25, and/or one or both of Wee1 and Mik1, the tyrosine kinases that phosphorylate Cdc2, are regulated by the replication checkpoint.

CDC2 Protein Kinase↗
PubMed1997

Cdc25 mitotic inducer targeted by chk1 DNA damage checkpoint kinase.

Arrest of the cell cycle at the G2 checkpoint, induced by DNA damage, requires inhibitory phosphorylation of the kinase Cdc2 in both fission yeast and human cells. The kinase Wee1 and the phosphatase Cdc25, which regulate Cdc2 phosphorylation, were evaluated as targets of Chk1, a kinase essential for the checkpoint. Fission yeast cdc2-3w Deltacdc25 cells, which express activated Cdc2 and lack Cdc25, were responsive to Wee1 but insensitive to Chk1 and irradiation. Expression of large amounts of Chk1 produced the same phenotype as did loss of the cdc25 gene in cdc2-3w cells. Cdc25 associated with Chk1 in vivo and was phosphorylated when copurified in Chk1 complexes. These findings identify Cdc25, but not Wee1, as a target of the DNA damage checkpoint.

Adenosine Triphosphatases↗
PubMed1997

Cdc2 tyrosine phosphorylation is required for the DNA damage checkpoint in fission yeast.

A common cellular response to DNA damage is cell cycle arrest. This checkpoint control has been the subject of intensive genetic investigation, but the biochemical mechanism that prevents mitosis following DNA damage is unknown. In Schizosaccharomyces pombe, as well as vertebrates, the timing of mitosis under normal circumstances is determined by the balance of kinases and phosphatases that regulate inhibitory phosphorylation of Cdc2. In S. pombe, the phosphorylation occurs on tyrosine-15. This method of mitotic control is also used in S. pombe to couple mitosis with completion of DNA replication, but the role of Cdc2 tyrosine phosphorylation in the Chk1 kinase-mediated DNA damage checkpoint has remained uncertain. We show that, in contrast to recent speculation, the G2 DNA damage checkpoint arrest in S. pombe depends on the inhibitory tyrosine phosphorylation of Cdc2 carried out by the Wee1 and Mik1 kinases. Furthermore, the rate of Cdc2 tyrosine dephosphorylation is reduced by irradiation. This result implicates regulation of Cdc2 tyrosine dephosphorylation, mainly carried out by the Cdc25 tyrosine phosphatase, as an important part of the mechanism by which the DNA damage checkpoint induces Cdc2 inhibition and G2 arrest.

CDC2 Protein Kinase↗
PubMedDate not supplied

Checkpoints: it takes more than time to heal some wounds.

The S-phase DNA damage checkpoint seems to provide a twist on the checkpoint theme. Instead of delaying replication and allowing repair as a consequence, it may activate repair and delay replication as a consequence.

Animals↗