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Phosphorylation of human Fen1 by cyclin-dependent kinase modulates its role in replication fork regulation

Oncogene volume 22, pages 4301–4313 (2003)Cite this article

Abstract

Cyclin-dependent kinase (Cdk) Cdk1–Cyclin A can phosphorylate Flap endonuclease 1 (Fen1), a key-enzyme of the DNA replication machinery, in late S phase. Cdk1–cyclin A forms a complex in vitro and in vivo with Fen1. Furthermore, Fen1 phosphorylation is detected in vivo and depends upon Cdks activity. As a functional consequence of phosphorylation by Cdk1–Cyclin A in vitro, endo- and exonuclease activities of Fen1 are reduced whereas its DNA binding is not affected. Moreover, phosphorylation of Fen1 by Cdk1–Cyclin A abrogates its proliferating cell nuclear antigen (PCNA) binding thus preventing stimulation of Fen1 by PCNA. Concomitantly, human cells expressing the S187A mutant defective for Cdk1–Cyclin A phosphorylation accumulate in S phase consistent with a failure in cell cycle regulation through DNA replication. Our results suggest a novel regulatory role of Cdks onto the end of S phase by targeting directly a key enzyme involved in DNA replication.

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Acknowledgements

We thank HP Nasheuer for providing purified Cdks complexes, JP Magyar for the Cdk1 clone, R Freire for GST-PCNA plasmid, L Meijer for the gift of Olomoucine, T Hennet for the use of FACScan™ flow cytometer, S Hasan and ZO Jónsson for stimulating discussions. This work has been supported by grants from the Swiss National Science Foundation (Grant 31-61361.00) to GH and SK, and by the Kanton of Zürich to GH and UH.

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  1. Ghislaine Henneke and Stéphane Koundrioukoff: Both authors contributed equally to this work

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  1. Institute of Veterinary Biochemistry and Molecular Biology, University of Zürich, Winterthurerstrasse 190, Zürich, CH-8057, Switzerland

    Ghislaine Henneke, Stéphane Koundrioukoff & Ulrich Hübscher

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  1. Ghislaine Henneke
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  2. Stéphane Koundrioukoff
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Correspondence to Ulrich Hübscher.

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Henneke, G., Koundrioukoff, S. & Hübscher, U. Phosphorylation of human Fen1 by cyclin-dependent kinase modulates its role in replication fork regulation. Oncogene 22, 4301–4313 (2003). https://doi.org/10.1038/sj.onc.1206606

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