Human claspin is required for replication checkpoint control.
Claspin is a newly identified protein that regulates Chk1 activation in Xenopus. In the present study we investigated the role of human Claspin in the DNA damage/replication checkpoint in mammalian cells. We observed that human Claspin is a cell cycle regulated protein that peaks at S/G2 phase. Claspin localizes in the nuclei, but it only associates with Chk1 following replication stress or other types of DNA damage. In addition, Claspin is phosphorylated in response to replication stress, and this phosphorylation appears to be required for its association with Chk1. Moreover, Claspin interacts with the checkpoint proteins ATR and Rad9. Given that both the ATR and Rad9-Rad1-Hus1 complexes are involved in Chk1 activation, it is possible that Claspin works as an adaptor molecule bringing these molecules together. Using small interfering RNA technology, we have shown that down-regulation of Claspin expression inhibits Chk1 activation in response to replication stress. More importantly, down-regulation of Claspin augments the premature chromatin condensation induced by hydroxyurea, inhibits the UV-induced reduction of DNA synthesis, and decreases cell survival. Taken together, these data imply a potentially critical role for Claspin in replication checkpoint control in mammalian cells.
Chini
Claudia Christiano Silva
CC
Department of Oncology, Mayo Clinic and Foundation, Rochester, Minnesota 55905, USA.
Chen
Junjie
J
D048868:Adaptor Proteins, Signal Transducing; D064007:Ataxia Telangiectasia Mutated Proteins; D002352:Carrier Proteins / Q000737:chemistry*; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival; D002843:Chromatin / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis; D004249:DNA Damage; D004268:DNA-Binding Proteins; D015536:Down-Regulation; D004720:Endonucleases / Q000378:metabolism; D016195:G2 Phase; D006367:HeLa Cells; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology; D020014:K562 Cells; D008856:Microscopy, Fluorescence; D008938:Mitosis; D010766:Phosphorylation; D011233:Precipitin Tests; D017346:Protein Serine-Threonine Kinases; D034741:RNA, Small Interfering / Q000378:metabolism; D016196:S Phase; D029702:Schizosaccharomyces pombe Proteins; D013997:Time Factors; D014162:Transfection; D014407:Tumor Cells, Cultured; D014466:Ultraviolet Rays; D029867:Xenopus Proteins
D048868:Adaptor Proteins, Signal Transducing; C416368:CLSPN protein, Xenopus; C488158:CLSPN protein, human; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; D034741:RNA, Small Interfering; D029702:Schizosaccharomyces pombe Proteins; D029867:Xenopus Proteins; C106332:hus1 protein, S pombe; C069930:rad9 protein; D004247:DNA; C576898:Atr protein, Xenopus; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D004720:Endonucleases; D006918:Hydroxyurea