Disruption of mechanisms that prevent rereplication triggers a DNA damage response.
Eukaryotes replicate DNA once and only once per cell cycle due to multiple, partially overlapping mechanisms efficiently preventing reinitiation. The consequences of reinitiation are unknown. Here we show that the induction of rereplication by mutations in components of the prereplicative complex (origin recognition complex [ORC], Cdc6, and minichromosome maintenance proteins) causes a cell cycle arrest with activated Rad53, a large-budded morphology, and an undivided nucleus. Combining a mutation disrupting the Clb5-Orc6 interaction (ORC6-rxl) and a mutation stabilizing Cdc6 (CDC6(Delta)NT) causes a cell cycle delay with a similar phenotype, although this background is only partially compromised for rereplication control and does not exhibit overreplication detectable by fluorescence-activated cell sorting. We conducted a systematic screen that identified genetic requirements for the viability of these cells. ORC6-rxl CDC6(Delta)NT cells depend heavily on genes required for the DNA damage response and for double-strand-break repair by homologous recombination. Our results implicate an Mre11-Mec1-dependent pathway in limiting the extent of rereplication.
Archambault
Vincent
V
The Rockefeller University, 1230 York Ave., Box 237, New York, NY 10021, USA.
Ikui
Amy E
AE
Drapkin
Benjamin J
BJ
Cross
Frederick R
FR
D048868:Adaptor Proteins, Signal Transducing; D002453:Cell Cycle / Q000235:genetics* / Q000502:physiology; D018797:Cell Cycle Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D064447:Checkpoint Kinase 2; D004249:DNA Damage / Q000502:physiology*; D004261:DNA Replication / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000235:genetics / Q000378:metabolism; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000378:metabolism; D017930:Genes, Reporter; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D004718:Saccharomycetales / Q000235:genetics / Q000502:physiology
D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C440013:Ddc1 protein, S cerevisiae; D047908:Intracellular Signaling Peptides and Proteins; C440014:LCD1 protein, S cerevisiae; C441579:MRC1 protein, S cerevisiae; D009687:Nuclear Proteins; D010750:Phosphoproteins; C501395:RAD17 protein, S cerevisiae; C086930:RAD50 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C119636:TOF1 protein, S cerevisiae; C086929:XRS2 protein, S cerevisiae; D064447:Checkpoint Kinase 2; C088937:MEC1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; C067022:RAD53 protein, S cerevisiae; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; C094191:MRE11 protein, S cerevisiae