The G2 DNA damage checkpoint targets both Wee1 and Cdc25

Jeanette M. Raleigh, Matthew J. O'Connell

Research output: Contribution to journalArticlepeer-review

134 Scopus citations

Abstract

The onset of mitosis is controlled by the cyclin dependent kinase Cdc2p. Cdc2p activity is controlled through the balance of phosphorylation and dephosphorylation of tyrosine-15 (Y15) by the Wee1p kinase and Cdc25p phosphatase. In the fission yeast Schizosaccharomyces pombe, detection of DNA damage in G2 activates a checkpoint that prevents entry into mitosis through the maintenance of Y15 phosphorylation of Cdc2p, thus ensuring DNA repair precedes chromosome segregation. The protein kinase Chk1p is the endpoint of this checkpoint pathway. We have previously reported that overexpression of Chk1p causes a wee1+-dependent G2 arrest, and this or irradiation leads to hyperphosphorylation of Wee1p. Moreover, Chk1p directly phosphorylates Wee1p in vitro. These data suggested that Wee1p is a key target of Chk1p action in checkpoint control. However, cells lacking wee1+ are checkpoint proficient and sustained Chk1p overexpression arrests cell cycle progression independently of Wee1p. Therefore, up-regulation of Wee1p alone cannot enforce a checkpoint arrest. Chk1p can also phosphorylate Cdc25p in vitro. These phosphorylation events are thought to promote the interaction with 14-3-3 proteins the cytoplasmic retention of the 14-3-3/Cdc25p complexes. However we show here that the G2 DNA damage checkpoint is intact in cells that regulate mitotic entry independently of Cdc25p. Further, these cells are still sensitive to Chk1p-mediated arrest, and so downregulation of Cdc25p is also insufficient to regulate checkpoint arrest. Conversely, inactivation of both wee1+ and cdc25+ abolishes checkpoint control. We also show that activation of the G2 DNA damage checkpoint induces a transient increase in Wee1p levels. We conclude that the G2 DNA damage checkpoint simultaneously signals via both up-regulation of Wee1p and down-regulation of Cdc25p, thus providing a double-lock mechanism to ensure cell cycle arrest and genomic stability.

Original languageEnglish
Pages (from-to)1727-1736
Number of pages10
JournalJournal of Cell Science
Volume113
Issue number10
StatePublished - 2000
Externally publishedYes

Keywords

  • Cdc25
  • Checkpoint
  • Chk1p
  • Fission yeast
  • Wee1

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