The fission yeast gene cdc18+ is required for entry into S phase and for coupling mitosis to the successful completion of S phase. Cdc18 is a highly unstable protein that is expressed only once per cell cycle at the G1/S boundary. Overexpression of Cdc18 causes a mitotic delay and reinitiation of DNA replication, suggesting that the inactivation of Cdc18 plays a role in preventing rereplication within a given cell cycle. In this paper, we present evidence that Cdc18 is associated with active cyclin- dependent kinase in vivo. We have expressed Cdc18 as a glutathione S- transferase fusion in fission yeast and demonstrated that the fusion protein is functional in vivo. We find that the Cdc18 fusion protein copurifies with a kinase activity capable of phosphorylating historic H1 and Cdc18. The activity was identified by a variety of methods as the cyclin-dependent kinase containing the product of the cdc2+ gene. The amino terminus of Cdc18 is required for association with cyclin-dependent kinase, but the association does not require the consensus cyclin-dependent kinase phosphorylation sites in this region. Additionally, both G1/S and mitotic forms of cyclin- dependent kinase phosphorylate and interact with Cdc18. These interactions between Cdc18 and cyclin-dependent kinases suggest mechanisms by which cyclin-dependent kinases could activate the initiation of DNA replication and could prevent rereplication.
|Original language||English (US)|
|Number of pages||6|
|Journal||Proceedings of the National Academy of Sciences of the United States of America|
|State||Published - Jun 10 1997|
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