Differential susceptibility of yeast S and M phase CDK complexes to inhibitory tyrosine phosphorylation.
BACKGROUND
Several checkpoint pathways employ Wee1-mediated inhibitory tyrosine phosphorylation of cyclin-dependent kinases (CDKs) to restrain cell-cycle progression. Whereas in vertebrates this strategy can delay both DNA replication and mitosis, in yeast cells only mitosis is delayed. This is particularly surprising because yeasts, unlike vertebrates, employ a single family of cyclins (B type) and the same CDK to promote both S phase and mitosis. The G2-specific arrest could be explained in two fundamentally different ways: tyrosine phosphorylation of cyclin/CDK complexes could leave sufficient residual activity to promote S phase, or S phase-promoting cyclin/CDK complexes could somehow be protected from checkpoint-induced tyrosine phosphorylation.


RESULTS
We demonstrate that in Saccharomyces cerevisiae, several cyclin/CDK complexes are protected from inhibitory tyrosine phosphorylation, allowing Clb5,6p to promote DNA replication and Clb3,4p to promote spindle assembly, even under checkpoint-inducing conditions that block nuclear division. In vivo, S phase-promoting Clb5p/Cdc28p complexes were phosphorylated more slowly and dephosphorylated more effectively than were mitosis-promoting Clb2p/Cdc28p complexes. Moreover, we show that the CDK inhibitor (CKI) Sic1p protects bound Clb5p/Cdc28p complexes from tyrosine phosphorylation, allowing the accumulation of unphosphorylated complexes that are unleashed when Sic1p is degraded to promote S phase. The vertebrate CKI p27(Kip1) similarly protects Cyclin A/Cdk2 complexes from Wee1, suggesting that the antagonism between CKIs and Wee1 is evolutionarily conserved.


CONCLUSIONS
In yeast cells, the combination of CKI binding and preferential phosphorylation/dephosphorylation of different B cyclin/CDK complexes renders S phase progression immune from checkpoints acting via CDK tyrosine phosphorylation.
Keaton
Mignon A
MA
Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC 27710, USA.
Bardes
Elaine S G
ES
Marquitz
Aron R
AR
Freel
Christopher D
CD
Zyla
Trevin R
TR
Rudolph
Johannes
J
Lew
Daniel J
DJ
D000199:Actins / Q000037:antagonists & inhibitors / Q000378:metabolism; D019086:Bridged Bicyclo Compounds, Heterocyclic; D017489:CDC28 Protein Kinase, S cerevisiae / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism*; D019926:Cyclin B / Q000378:metabolism; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; D004261:DNA Replication / Q000502:physiology; D008938:Mitosis / Q000502:physiology*; D046912:Multiprotein Complexes / Q000378:metabolism; D010766:Phosphorylation; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D016196:S Phase / Q000502:physiology*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D008941:Spindle Apparatus / Q000378:metabolism; D053778:Thiazolidines; D014443:Tyrosine / Q000378:metabolism; D020666:ras-GRF1
D000199:Actins; D019086:Bridged Bicyclo Compounds, Heterocyclic; C542351:CDC25 protein, S cerevisiae; C076558:CLB5 protein, S cerevisiae; D018797:Cell Cycle Proteins; D019926:Cyclin B; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; D046912:Multiprotein Complexes; C086926:SIC1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D053778:Thiazolidines; D020666:ras-GRF1; D014443:Tyrosine; C099685:SWE1 protein, S cerevisiae; D011505:Protein-Tyrosine Kinases; D017489:CDC28 Protein Kinase, S cerevisiae; D017027:Protein Tyrosine Phosphatases; C037067:latrunculin A