Cdc4, a Protein Required for the Onset of S Phase, Serves an Essential Function during G₂/M Transition in Saccharomyces cerevisiae

Abstract

Saccharomyces cerevisiae proteins Cdc4 and Cdc20 contain WD40 repeats and participate in proteolytic processes. However, they are thought to act at two different stages of the cell cycle: Cdc4 is involved in the proteolysis of the Cdk inhibitor, Sic1, necessary for G₁/S transition, while Cdc20 mediates anaphase-promoting complex-dependent degradation of anaphase inhibitor Pds1, a process necessary for the onset of chromosome segregation. We have isolated three mutant alleles of CDC4(cdc4-10, cdc4-11, and cdc4-16) which suppress the nuclear division defect of cdc20-1cells. However, the previously characterized mutation cdc4-1 and a new allele, cdc4-12, do not alleviate the defect of cdc20-1 cells. This genetic interaction suggests an additional role for Cdc4 in G₂/M. Reexamination of the cdc4-1 mutant revealed that, in addition to being defective in the onset of S phase, it is also defective in G₂/M transition when released from hydroxyurea-induced S-phase arrest. A second function for CDC4 in late S or G₂ phase was further confirmed by the observation that cells lacking the CDC4gene are arrested both at G₁/S and at G₂/M. We subsequently isolated additional temperature-sensitive mutations in the CDC4 gene (such as cdc4-12) that render the mutant defective in both G₁/S and G₂/M transitions at the restrictive temperature. While the G₁/S block in both cdc4-12 and cdc4Δ mutants is abolished by the deletion of the SIC1 gene (causing the mutants to be arrested predominantly in G₂/M), the preanaphase arrest in the cdc4-12 mutant is relieved by the deletion of PDS1. Collectively, these observations suggest that, in addition to its involvement in the initiation of S phase, Cdc4 may also be required for the onset of anaphase.

ACKNOWLEDGMENTS

We thank Alice Tay and her lab members for sequencing the DNA clones, Sara Zaman for the sic1Δ strain and the SIC1-myc3 plasmid, and the technical staff for their help. We are grateful to Breck Byers for the pds1Δ strain and John Kilmartin for antitubulin antibody.

This work was supported by the National Science and Technology Board, Singapore.