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Molecular and Cellular Biology, September 2001, p. 5767-5777, Vol. 21, No. 17
0270-7306/01/$04.00+0   DOI: 10.1128/MCB.21.17.5767-5777.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Control of DNA Rereplication via Cdc2 Phosphorylation Sites in the Origin Recognition Complex

Amit Vas, Winnie Mok, and Janet Leatherwood*

Department of Molecular Genetics and Microbiology, State University of New York, Stony Brook, New York 11794-5222

Received 28 March 2001/Accepted 7 May 2001

Cdc2 kinase is a master regulator of cell cycle progression in the fission yeast Schizosaccharomyces pombe. Our data indicate that Cdc2 phosphorylates replication factor Orp2, a subunit of the origin recognition complex (ORC). Cdc2 phosphorylation of Orp2 appears to be one of multiple mechanisms by which Cdc2 prevents DNA rereplication in a single cell cycle. Cdc2 phosphorylation of Orp2 is not required for Cdc2 to activate DNA replication initiation. Phosphorylation of Orp2 appears first in S phase and becomes maximal in G2 and M when Cdc2 kinase activity is required to prevent reinitiation of DNA replication. A mutant lacking Cdc2 phosphorylation sites in Orp2 (orp2-T4A) allowed greater rereplication of DNA than congenic orp2 wild-type strains when the limiting replication initiation factor Cdc18 was deregulated. Thus, Cdc2 phosphorylation of Orp2 may be redundant with regulation of Cdc18 for preventing reinitiation of DNA synthesis. Since Cdc2 phosphorylation sites are present in Orp2 (also known as Orc2) from yeasts to metazoans, we propose that cell cycle-regulated phosphorylation of the ORC provides a safety net to prevent DNA rereplication and resulting genetic instability.

* Corresponding author. Mailing address: Department of Molecular Genetics and Microbiology, State University of New York, Stony Brook, NY 11794-5222. Phone: (631) 632-9644. Fax: (631) 632-9797. E-mail: janet.leatherwood{at}sunysb.edu.

Molecular and Cellular Biology, September 2001, p. 5767-5777, Vol. 21, No. 17
0270-7306/01/$04.00+0   DOI: 10.1128/MCB.21.17.5767-5777.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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