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Mitotic kinase regulation of DNA replication forks
Sudikchya Shrestha1, Natasia Paukovich1,2, Samuel R Greenfield1,3,2
1Department of Biochemistry and Molecular Genetics, University of Colorado Anschutz, Aurora, CO, USA.
Abstract:
While DNA replication forks initiate in S phase, they do not necessarily complete and terminate prior to cell entry into mitosis. How mitotic proteins regulate leftover replication forks is not well-understood. Using reconstituted DNA replication forks with purified proteins, we show that the budding yeast mitotic kinases Clb2-CDK (M-CDK) and Cdc5 (Plk1 homolog) phosphorylate and regulate several replication elongation proteins. Mrc1 phosphorylation by both kinases results in slower replication, and Polα phosphorylation by M-CDK results in less lagging strand initiation. We further show that a phospho-resistant mutant of Polα bypasses M-CDK inhibition of Polα activity in reconstituted replication reactions. Yeast cells expressing the phospho-resistant mutant exhibit faster cell cycle progression revealing a potential negative feedback mechanism between DNA replication forks and mitotic progression.
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