Destruction of Claspin by SCFbetaTrCP restrains Chk1 activation and facilitates recovery from genotoxic stress.
We show that Claspin, an adaptor protein required for Chk1 activation, becomes degraded at the onset of mitosis. Claspin degradation was triggered by its interaction with, and ubiquitylation by, the SCFbetaTrCP ubiquitin ligase. This interaction was phosphorylation dependent and required the activity of the Plk1 kinase and the integrity of a betaTrCP recognition motif (phosphodegron) in the N terminus of Claspin. Uncoupling of Claspin from betaTrCP by mutating the conserved serines in Claspin's phosphodegron or by knocking down betaTrCP stabilized Claspin in mitosis, impaired Chk1 dephosphorylation, and delayed G2/M transition during recovery from cell cycle arrest imposed by DNA damage or replication stress. Moreover, the inability to degrade Claspin allowed partial reactivation of Chk1 in cells exposed to DNA damage after passing the G2/M transition. Our data suggest that degradation of Claspin facilitates timely reversal of the checkpoint response and delineates the period permissive for Chk1 activation during cell cycle progression.
Mailand
Niels
N
Institute of Cancer Biology and Centre for Genotoxic Stress Research, Danish Cancer Society, Strandboulevarden 49, DK-2100, Copenhagen, Denmark.
Bekker-Jensen
Simon
S
Bartek
Jiri
J
Lukas
Jiri
J
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000502:physiology*; D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D002453:Cell Cycle / Q000187:drug effects; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D000071877:Checkpoint Kinase 1; D019925:Cyclin A / Q000235:genetics / Q000378:metabolism; D004249:DNA Damage / Q000502:physiology*; D004317:Doxorubicin / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D044843:SKP Cullin F-Box Protein Ligases / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; C488158:CLSPN protein, human; D018797:Cell Cycle Proteins; D019925:Cyclin A; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D025801:Ubiquitin; D004317:Doxorubicin; D044843:SKP Cullin F-Box Protein Ligases; D011494:Protein Kinases; D011505:Protein-Tyrosine Kinases; C499349:WEE1 protein, human; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1