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The DNA damage/replication checkpoints act by sensing the presence of damaged DNA or stalled replication forks and initiate signaling pathways that arrest cell cycle progression. Here we report the cloning and characterization of Xenopus orthologues of the RFCand PCNA-related checkpoint proteins. XRad17 shares regions of homology with the five subunits of Replication factor C. XRad9, XRad1, and XHus1 (components of the 9-1-1 complex) all show homology to the DNA polymerase processivity factor PCNA. We demonstrate that these proteins associate with chromatin and are phosphorylated when replication is inhibited by aphidicolin. Phosphorylation of X9-1-1 is caffeine sensitive, but the chromatin association of XRad17 and the X9-1-1 complex after replication block is unaffected by caffeine. This suggests that the X9-1-1 complex can associate with chromatin independently of XAtm/XAtr activity. We further demonstrate that XRad17 is essential for the chromatin binding and checkpoint-dependent phosphorylation of X9-1-1 and for the activation of XChk1 when the replication checkpoint is induced by aphidicolin. XRad17 is not, however, required for the activation of XCds1 in response to dsDNA ends.

Original publication

DOI

10.1091/mbc.e03-03-0138

Type

Journal article

Journal

Mol Biol Cell

Publication Date

09/2003

Volume

14

Pages

3898 - 3910

Keywords

Amino Acid Sequence, Animals, Aphidicolin, Cell Cycle, Cell Cycle Proteins, Cell Nucleus, Chromatin, Cloning, Molecular, DNA Replication, DNA-Binding Proteins, Female, Male, Molecular Sequence Data, Oocytes, Phosphorylation, Proliferating Cell Nuclear Antigen, Sequence Homology, Spermatozoa, Xenopus Proteins, Xenopus laevis