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Published on: October 11, 2022
Cdc2 tyrosine phosphorylation is not required for the S-phase DNA damage checkpoint in fission yeast
Naveen Kommajosyula1, Nicholas Rhind
1Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01609, USA.
Abstract:
The S-phase DNA damage checkpoint slows replication when damage occurs during S phase. Cdc25, which activates Cdc2 by dephosphorylating tyrosine-15, has been shown to be a downstream target of the checkpoint in metazoans, but its role is not clear in fission yeast. The dephosphorylation of Cdc2 has been assumed not to play a role in S-phase regulation because cells replicate in the absence of Cdc25, demonstrating that tyrosine-15 phosphorylated dc2 is sufficient for S phase. However, it has been reported recently that Cdc25 is involved in the slowing of S phase in response to damage in fission yeast, suggesting a modulatory role for Cdc2 dephosphorylation in S phase. We have investigated the role of Cdc25 and the tyrosine phosphorylation of Cdc2 in the S-phase damage checkpoint, and our results show that Cdc2 phosphorylation is not a target of the checkpoint. The checkpoint was not compromised in a Cdc25 overexpressing strain, a strain carrying nonphosphorylatable form of Cdc2, or in a strain lacking Cdc25. Our results are consistent with a strictly Cdc2-Y15 phosphorylation-independent mechanism of the fission yeast S-phase DNA damage checkpoint.
Insights
The fission yeast S-phase DNA damage checkpoint does not rely on Cdc25 or Cdc2 tyrosine-15 phosphorylation. This study reveals a phosphorylation-independent mechanism for regulating DNA replication during S phase.
Area of Science:
- Cell Cycle Regulation
- DNA Damage Response
- Molecular Biology
Background:
- The S-phase DNA damage checkpoint halts replication upon DNA damage during S phase.
- Cdc25 activates Cdc2 by dephosphorylating tyrosine-15 and is a known checkpoint target in metazoans.
- The role of Cdc2 dephosphorylation in fission yeast S-phase regulation was previously unclear.
Purpose of the Study:
- To investigate the role of Cdc25 and Cdc2 tyrosine phosphorylation in the fission yeast S-phase DNA damage checkpoint.
- To determine if Cdc2 phosphorylation is a target of the S-phase damage checkpoint in fission yeast.
Main Methods:
- Genetic analysis of fission yeast strains.
- Overexpression of Cdc25.
- Use of a strain with a non-phosphorylatable form of Cdc2.
- Analysis of a strain lacking Cdc25.
Main Results:
- The S-phase DNA damage checkpoint functions independently of Cdc25.
- Cdc2 phosphorylation at tyrosine-15 is not a target of the S-phase damage checkpoint.
- Checkpoint function remained intact in Cdc25-overexpressing, non-phosphorylatable Cdc2, and Cdc25-lacking strains.
Conclusions:
- Fission yeast S-phase DNA damage checkpoint operates via a mechanism strictly independent of Cdc2-Y15 phosphorylation.
- Cdc25 and Cdc2 dephosphorylation do not play a critical role in this checkpoint pathway.
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