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bioRxiv · 10.64898/2026.05.16.725582

Mitotic kinase regulation of DNA replication forks

Shrestha, S.·Paukovich, N.·Greenfield, S. R.·MacFadden, A.·Smith, S. N.·Bevers, S.·McClure, A. W.
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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 Pol phosphorylation by M-CDK results in less lagging strand initiation. We further show that a phospho-resistant mutant of Pol bypasses M-CDK inhibition of Pol 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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Shrestha, S., Paukovich, N., Greenfield, S. R., MacFadden, A., Smith, S. N., Bevers, S., McClure, A. W.. 2026-05-19. Mitotic kinase regulation of DNA replication forks. https://doi.org/10.64898/2026.05.16.725582

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