Chk1 is required to maintain claspin stability.
Claspin is a Chk1-interacting protein that participates in the DNA replication checkpoint. Expression of Claspin fluctuates in a cell cycle-dependent manner, but the mechanisms involved in the regulation of Claspin protein levels have not been explored. In this study, we show that Claspin expression is downregulated by the proteasome-mediated degradation pathway and that Chk1 is required to maintain Claspin stability. Downregulation of Chk1 expression by siRNA or inhibition of Chk1 activity by UCN01 decreases Claspin levels in cells. Conversely, overexpression of Chk1 increases Claspin levels. These data indicate a role of Chk1 in regulating Claspin stability in the cell. Since Claspin has also been shown to participate in Chk1 activation following DNA damage, we further explored the exact role of Claspin during Chk1 activation following replication stress. We observed that while Rad17 is required for early Chk1 activation after hydroxyurea treatment, Claspin is only required to sustain Chk1 activation. Based on these findings, we propose that Claspin functions at late stages of Chk1 activation following DNA damage. Once Chk1 is activated, it stabilizes Claspin, which in turn helps to maintain Chk1 activation during replication stress. In summary, these data indicate that the interaction between Claspin and Chk1 is complex. These proteins regulate each other and thus ensure the proper cell cycle progression and replication checkpoint control.
Chini
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CC
Department of Oncology, Mayo Clinic and Foundation, Rochester, MN 55905, USA.
Wood
J
J
Chen
J
J
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D002453:Cell Cycle / Q000502:physiology; D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1; D006367:HeLa Cells; D006801:Humans; D046988:Proteasome Endopeptidase Complex / Q000502:physiology; D011494:Protein Kinases / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology
D048868:Adaptor Proteins, Signal Transducing; C488158:CLSPN protein, human; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D046988:Proteasome Endopeptidase Complex