In eukaryotic cells, the Cdc2-cyclin B complex is a key regulatory kinase that induces mitosis. In vertebrates, Cdc2 is negatively regulated during interphase by inhibitory phosphorylation on its Thr-14 and Tyr-15 residues. The dual-specificity phosphatase Cdc25C activates Cdc2-cyclin B at the G2/M transition by dephosphorylating these residues. The entry into mitosis is suppressed if the genome has not been replicated properly or has suffered damage. These regulatory mechanisms, which are referred to as the DNA replication and DNA damage checkpoints, respectively, function in part by blocking the action of Cdc25C. In vertebrates, the down-regulation of Cdc25C during a DNA replication checkpoint response involves phosphorylation by the protein kinase Chk1. In frog egg extracts, for example, the Xenopus homologue of Chk1 (Xchk1) phosphorylates Cdc25C on Ser-287. The activation on Xchk1 in response to DNA replication blocks is carried out by Xenopus Atr (Xatr), Furthermore, the Xatr-dependent phosphorylation of Xchk1 requires a novel protein named Claspin. A comprehensive study of the regulatory pathway containing Cdc25, Xchk1, Claspin, and Xatr is proposed. The following experimental objectives will be pursued. 1. Control of Cdc25. Additional studies on the mechanism by which Cdc25 is down-regulated by Xck1 will be conducted. 2. Regulation of Xchk1. The mechanisms which control the action of Xchk1 will be analyzed further. 3. Functional dissection of Claspin. Studies will be carried out to elucidate the biochemical function of Claspin. 4. Identification of Novel Components in the Claspin-Echk1 Pathway. Various searchers will be employed to identify additional regulators in this network.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Method to Extend Research in Time (MERIT) Award (R37)
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Cell Development and Function Integrated Review Group (CDF)
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Zatz, Marion M
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California Institute of Technology
Schools of Arts and Sciences
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Kumagai, Akiko; Dunphy, William G (2017) MTBP, the partner of Treslin, contains a novel DNA-binding domain that is essential for proper initiation of DNA replication. Mol Biol Cell 28:2998-3012
Mu, Ruiling; Tat, John; Zamudio, Robert et al. (2017) CKS Proteins Promote Checkpoint Recovery by Stimulating Phosphorylation of Treslin. Mol Cell Biol 37:
Guo, Cai; Kumagai, Akiko; Schlacher, Katharina et al. (2015) Interaction of Chk1 with Treslin negatively regulates the initiation of chromosomal DNA replication. Mol Cell 57:492-505
Ryu, Hyunju; Yoshida, Makoto M; Sridharan, Vinidhra et al. (2015) SUMOylation of the C-terminal domain of DNA topoisomerase II? regulates the centromeric localization of Claspin. Cell Cycle 14:2777-84
Lee, Joon; Dunphy, William G (2013) The Mre11-Rad50-Nbs1 (MRN) complex has a specific role in the activation of Chk1 in response to stalled replication forks. Mol Biol Cell 24:1343-53
Kumar, Sanjay; Yoo, Hae Yong; Kumagai, Akiko et al. (2012) Role for Rif1 in the checkpoint response to damaged DNA in Xenopus egg extracts. Cell Cycle 11:1183-94
Meng, Zheng; Capalbo, Luisa; Glover, David M et al. (2011) Role for casein kinase 1 in the phosphorylation of Claspin on critical residues necessary for the activation of Chk1. Mol Biol Cell 22:2834-47
Kumagai, Akiko; Shevchenko, Anna; Shevchenko, Andrej et al. (2011) Direct regulation of Treslin by cyclin-dependent kinase is essential for the onset of DNA replication. J Cell Biol 193:995-1007
Ramírez-Lugo, Juan S; Yoo, Hae Yong; Yoon, Su Jin et al. (2011) CtIP interacts with TopBP1 and Nbs1 in the response to double-stranded DNA breaks (DSBs) in Xenopus egg extracts. Cell Cycle 10:469-80
Wawrousek, Karen E; Fortini, Barbara K; Polaczek, Piotr et al. (2010) Xenopus DNA2 is a helicase/nuclease that is found in complexes with replication proteins And-1/Ctf4 and Mcm10 and DSB response proteins Nbs1 and ATM. Cell Cycle 9:1156-66

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