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Proc Natl Acad Sci U S A. 2019 Jul 2;116(27):13374-13383. doi: 10.1073/pnas.1903418116. Epub 2019 Jun 17.

An ATR and CHK1 kinase signaling mechanism that limits origin firing during unperturbed DNA replication.

Author information

1
Department of Radiation Oncology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213.
2
Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY 10016.
3
Department of Cell Biology, Center for Biologic Imaging, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261.
4
Department of Biostatistics, University of Pittsburgh, Pittsburgh, PA 15260.
5
Department of Biomedical Informatics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15206.
6
Department of Radiation Oncology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213; bakkenistcj@upmc.edu.
7
Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261.

Abstract

DNA damage-induced signaling by ATR and CHK1 inhibits DNA replication, stabilizes stalled and collapsed replication forks, and mediates the repair of multiple classes of DNA lesions. We and others have shown that ATR kinase inhibitors, three of which are currently undergoing clinical trials, induce excessive origin firing during unperturbed DNA replication, indicating that ATR kinase activity limits replication initiation in the absence of damage. However, the origins impacted and the underlying mechanism(s) have not been described. Here, we show that unperturbed DNA replication is associated with a low level of ATR and CHK1 kinase signaling and that inhibition of this signaling induces dormant origin firing at sites of ongoing replication throughout the S phase. We show that ATR and CHK1 kinase inhibitors induce RIF1 Ser2205 phosphorylation in a CDK1-dependent manner, which disrupts an interaction between RIF1 and PP1 phosphatase. Thus, ATR and CHK1 signaling suppresses CDK1 kinase activity throughout the S phase and stabilizes an interaction between RIF1 and PP1 in replicating cells. PP1 dephosphorylates key CDC7 and CDK2 kinase substrates to inhibit the assembly and activation of the replicative helicase. This mechanism limits origin firing during unperturbed DNA replication in human cells.

KEYWORDS:

ATR; CDK1; RIF1; dormant origins; replication

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