Functional relationship between Claspin and Rad17

► This is the first study to characterize Claspin gene knockout cells using DT40 cells. ► Claspin plays a critical role in DNA replication in the absence of exogenous stress. ► This is the first study to reveal the functional relationship between Claspin and Rad17. ► RAD17− cells observed a greater...

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Bibliographic Details
Published in:Biochemical and biophysical research communications 2011-10, Vol.414 (2), p.298-303
Main Authors: Yoshimura, Akari, Akita, Motomu, Hosono, Yoshifumi, Abe, Takuya, Kobayashi, Masahiko, Yamamoto, Ken-ichi, Tada, Shusuke, Seki, Masayuki, Enomoto, Takemi
Format: Article
Language:English
Subjects:
Adaptor Proteins, Signal Transducing - genetics
Adaptor Proteins, Signal Transducing - metabolism
Animals
Cell Cycle Proteins - genetics
Cell Cycle Proteins - metabolism
Cell Line
Chickens
Claspin
DNA replication
DNA Replication - genetics
DT40
Gene Knockout Techniques
Humans
Rad17
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Summary:► This is the first study to characterize Claspin gene knockout cells using DT40 cells. ► Claspin plays a critical role in DNA replication in the absence of exogenous stress. ► This is the first study to reveal the functional relationship between Claspin and Rad17. ► RAD17− cells observed a greater defect in MMS-induced checkpoint activation than Claspin−/− cells. ► Knocking out RAD17 gene showed almost no additive effects in Claspin−/− cells. Claspin was originally identified as a Check1 (Chk1)-interacting protein. Claspin and Rad17 are reportedly involved in the DNA damage-induced phosphorylation of Chk1, a hallmark of checkpoint activation. To understand the cellular functions of Claspin and the functional relationship between Claspin and Rad17, we generated Claspin−/− and Claspin−/−/RAD17− cells using chicken DT40 cells, which contain an exogenously introduced Claspin that can be suppressed by the addition of doxycycline (Dox). In the presence of Dox, Claspin−/− cells ceased growth within 2days, leading to cell death. In addition, a remarkable reduction in the rate of DNA elongation was observed in Claspin-depleted cells, suggesting that Claspin plays a critical role in DNA replication in the absence of exogenous stress. When cells were exposed to methyl methanesulfonate (MMS), a DNA damaging agent, RAD17− cells showed a greater defect in checkpoint activation than Claspin−/− cells as monitored by progression of cell cycle and phosphorylation of Chk1. Knocking out RAD17 gene showed almost no additive effects on cell death and DNA elongation rates in Claspin-depleted cells.
ISSN:0006-291X
1090-2104
DOI:10.1016/j.bbrc.2011.09.037