A requirement for replication in activation of the ATR-dependent DNA damage checkpoint.
Using the Xenopus egg extract system, we investigated the involvement of DNA replication in activation of the DNA damage checkpoint. We show here that DNA damage slows replication in a checkpoint-independent manner and is accompanied by replication-dependent recruitment of ATR and Rad1 to chromatin. We also find that the replication proteins RPA and Polalpha accumulate on chromatin following DNA damage. Finally, damage-induced Chk1 phosphorylation and checkpoint arrest are abrogated when replication is inhibited. These data indicate that replication is required for activation of the DNA damage checkpoint and suggest a unifying model for ATR activation by diverse lesions during S phase.
Lupardus
Patrick J
PJ
Department of Molecular Pharmacology, Stanford University, Stanford, California 94305-5174, USA.
Byun
Tony
T
Yee
Muh-Ching
MC
Hekmat-Nejad
Mohammad
M
Cimprich
Karlene A
KA
D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000378:metabolism*; D000071877:Checkpoint Kinase 1; D004249:DNA Damage; D004261:DNA Replication / Q000528:radiation effects; D004268:DNA-Binding Proteins; D004720:Endonucleases / Q000235:genetics / Q000378:metabolism; D005260:Female; D066298:In Vitro Techniques; D008741:Methyl Methanesulfonate / Q000633:toxicity; D008954:Models, Biological; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014466:Ultraviolet Rays / Q000009:adverse effects; D014981:Xenopus; D029867:Xenopus Proteins
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D029867:Xenopus Proteins; D008741:Methyl Methanesulfonate; D011494:Protein Kinases; C576898:Atr protein, Xenopus; D064007:Ataxia Telangiectasia Mutated Proteins; D000071877:Checkpoint Kinase 1; C000605875:Chek1 protein, Xenopus; D017346:Protein Serine-Threonine Kinases; D004720:Endonucleases