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MBC in Press, published online ahead of print March 5, 2004
Mol. Biol. Cell 10.1091/mbc.E03-08-0617

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Submitted on August 22, 2003
Revised on December 31, 2003
Accepted on February 12, 2004

The role and regulation of the preRC component Cdc6 in the initiation of premeiotic DNA replication

Yaara Ofir1, Shira Sagee1, Noga Guttmann-Raviv1, Lilach Pnuelli1, and Yona Kassir1*

1 Department of Biology, Technion - Israel Institute of Technology Haifa 32000 Israel

* Corresponding author. E-mail address: ykassir{at}tx.technion.ac.il.

In all eukaryotes, the initiation of DNA replication is regulated by the ordered assembly of DNA/proteins complexes on origins of DNA replication. In this report we examine the role of Cdc6, a component of the preReplication complex, in the initiation of premeiotic DNA replication in budding yeast. We show that in the meiotic cycle, Cdc6 is required for DNA synthesis and sporulation. Moreover, similarly to the regulation in the mitotic cell cycle, Cdc6 is specifically degraded upon entry into the meiotic S-phase. By contrast, chromatin-immunoprecipitation (ChIP) analysis reveals that the origin-bound Cdc6 is stable throughout the meiotic cycle. Preliminary evidence suggests that this protection reflects a change in chromatin structure that occurs in meiosis. Using the cdc28-degron allele, we show that depletion of Cdc28 leads to stabilization of Cdc6 in the mitotic cycle, but not in the meiotic cycle. We show physical association between Cdc6 and the meiosis-specific hCDK2 homolog, Ime2. These results suggest that under meiotic conditions, Ime2, rather than Cdc28, regulates the stability of Cdc6. ChIP analysis reveals that similarly to the mitotic cell cycle, Mcm2 binds origins in G1 and meiotic S-phases, and at the end of the second meiotic division, it is gradually removed from chromatin.




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