Involvement of RAD9-dependent damage checkpoint control in arrest of cell cycle, induction of cell death, and chromosome instability caused by defects in origin recognition complex in Saccharomyces cerevisiae.
Perturbation of origin firing in chromosome replication is a possible cause of spontaneous chromosome instability in multireplicon organisms. Here, we show that chromosomal abnormalities, including aneuploidy and chromosome rearrangement, were significantly increased in yeast diploid cells with defects in the origin recognition complex. The cell cycle of orc1-4/orc1-4 temperature-sensitive mutant was arrested at the G2/M boundary, after several rounds of cell division at the restrictive temperature. However, prolonged incubation of the mutant cells at 37 degrees C led to abrogation of G2 arrest, and simultaneously the cells started to lose viability. A sharp increase in chromosome instability followed the abrogation of G2 arrest. In orc1-4/orc1-4 rad9delta/rad9delta diploid cells grown at 37 degrees C, G2 arrest and induction of cell death were suppressed, while chromosome instability was synergistically augmented. These findings indicated that DNA lesions caused by a defect in Orc1p function trigger the RAD9-dependent checkpoint control, which ensures genomic integrity either by stopping the cell cycle progress until lesion repair, or by inducing cell death when the lesion is not properly repaired. At semirestrictive temperatures, orc2-1/orc2-1 diploid cells demonstrated G2 arrest and loss of cell viability, both of which require RAD9-dependent checkpoint control. However, chromosome instability was not induced in orc2-1/orc2-1 cells, even in the absence of the checkpoint control. These data suggest that once cells lose the damage checkpoint control, perturbation of origin firing can be tolerated by the cells. Furthermore, although a reduction in origin-firing capacity does not necessarily initiate chromosome instability, the Orc1p possesses a unique function, the loss of which induces instability in the chromosome.
Watanabe
Keiichi
K
Department of Molecular Biology, Graduate School of Biological Sciences, Nara Institute of Science and Technology (NAIST), Ikoma, Nara 630-0101, Japan.
Morishita
Jun
J
Umezu
Keiko
K
Shirahige
Katsuhiko
K
Maki
Hisaji
H
D002453:Cell Cycle / Q000235:genetics; D018797:Cell Cycle Proteins / Q000502:physiology*; D016923:Cell Death / Q000235:genetics / Q000502:physiology; D002869:Chromosome Aberrations; D015825:Chromosomes, Fungal; D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D004171:Diploidy; D018816:Genes, cdc; D006358:Hot Temperature; D019656:Loss of Heterozygosity; D008957:Models, Genetic; D009154:Mutation; D051738:Origin Recognition Complex; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics* / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins; D013696:Temperature; D013997:Time Factors
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C497214:ORC1 protein, S cerevisiae; C497215:ORC2 protein, S cerevisiae; D051738:Origin Recognition Complex; D029701:Saccharomyces cerevisiae Proteins; C069930:rad9 protein