Segregation of Unreplicated Chromosomes in Saccharomyces cerevisiae Reveals a Novel G₁/M-Phase Checkpoint

Abstract

Saccharomyces cerevisiae dbf4 and cdc7 cell cycle mutants block initiation of DNA synthesis (i.e., are iDS mutants) at 37&C and arrest the cell cycle with a 1C DNA content. Surprisingly, certain dbf4 and cdc7 strains divide their chromatin at 37&C. We found that the activation of the Cdc28 mitotic protein kinase and the Dbf2 kinase occurred with the correct relative timing with respect to each other and the observed division of the unreplicated chromatin. Furthermore, the division of unreplicated chromatin depended on a functional spindle. Therefore, the observed nuclear division resembled a normal mitosis, suggesting that S. cerevisiae commits to M phase in late G₁ independently of S phase. Genetic analysis of dbf4 and cdc7 strains showed that the ability to restrain mitosis during a late G₁ block depended on the genetic background of the strain concerned, since the dbf4 and cdc7 alleles examined showed the expected mitotic restraint in other backgrounds. This restraint was genetically dominant to lack of restraint, indicating that an active arrest mechanism, or checkpoint, was involved. However, none of the previously described mitotic checkpoint pathways were defective in the iDS strains that carry out mitosis without replicated DNA, therefore indicating that the checkpoint pathway that arrests mitosis in iDS mutants is novel. Thus, spontaneous strain differences have revealed that S. cerevisiae commits itself to mitosis in late G₁ independently of entry into S phase and that a novel checkpoint mechanism can restrain mitosis if cells are blocked in late G₁. We refer to this as the G₁/M-phase checkpoint since it acts in G₁ to restrain mitosis.