Functional relationship between Claspin and Rad17 |
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Authors: | Yoshimura Akari Akita Motomu Hosono Yoshifumi Abe Takuya Kobayashi Masahiko Yamamoto Ken-ichi Tada Shusuke Seki Masayuki Enomoto Takemi |
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Affiliation: | aMolecular Cell Biology Laboratory, Research Institute of Pharmaceutical Sciences, Faculty of Pharmacy, Musashino University, 1-1-20 Shinmachi, Nishitokyo-shi, Tokyo 202-8585, Japan;bMolecular Cell Biology Laboratory, Graduate School of Pharmaceutical Sciences, Tohoku University, Aoba 6-3, Aramaki, Aoba-ku, Sendai, Miyagi 980-8578, Japan;cDepartment of Molecular Pathology, Center for the Development of Molecular Target, Drugs, Cancer Research Institute, Kanazawa University, Kakuma-cho Nu 7, Kanazawa, Ishikawa 920-1192, Japan;dFaculty of Pharmaceutical Sciences, Teikyo Heisei University, 4-1 Uruido Minami, Ichihara, Chiba 290-0193, Japan |
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Abstract: | 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 2 days, 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. |
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Keywords: | Claspin Rad17 DT40 DNA replication |
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