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Highly Synchronous Mitotic Progression in Schizosaccharomyces pombe Upon Relief of Transient Cdc2-asM17 Inhibition

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Cell Cycle Oscillators

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2329))

Abstract

Synchronized progression of a cell population through the cell division cycle supports the biochemical and functional dissection of cell cycle controls and execution. The concerted behaviour of the population reflects the attributes of each cell within that population. The reversible imposition of a block to cell cycle progression at the G2–M boundary through transient inactivation of the Cdk1-Cyclin B activating phosphatase, Cdc25, with the temperature sensitive cdc25-22 mutant, has been widely used to study fission yeast mitosis and DNA replication. However, the biology of the compromised Cdc25-22 phosphatase generates significant division abnormalities upon release from mitotic arrest. We show how reversible inhibition of Cdc2-asM17, with the ATP analog 3-BrB-PP1, generates higher levels of synchrony with timing and morphology much more reminiscent of a normal division. We also describe a version of the H1 kinase assay of Cdk1-Cyclin B activity that is widely used to monitor mitotic progression which does not require radiolabeled ATP.

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Acknowledgments

Daniel Mulvihill (University of Kent, UK) and Janni Petersen (Flinders University, Australia) for critical reading of the manuscript and CRUK (A27336 and A24458) and Wellcome (200847/Z/16/Z) for funding.

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Authors and Affiliations

  1. Cell Division Group, CRUK Manchester Institute, The University of Manchester, Alderley Park, UK

    Pawan Singh, Lenka Halova & Iain Michael Hagan

Authors
  1. Pawan Singh
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  2. Lenka Halova
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  3. Iain Michael Hagan
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Correspondence to Iain Michael Hagan .

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Editors and Affiliations

  1. School of Science and Technology, Nottingham Trent University, Nottingham, UK

    Amanda S. Coutts

  2. Scientific Writer, Newcastle upon Tyne, UK

    Louise Weston

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Singh, P., Halova, L., Hagan, I.M. (2021). Highly Synchronous Mitotic Progression in Schizosaccharomyces pombe Upon Relief of Transient Cdc2-asM17 Inhibition. In: Coutts, A.S., Weston, L. (eds) Cell Cycle Oscillators . Methods in Molecular Biology, vol 2329. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1538-6_10

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  • DOI: https://doi.org/10.1007/978-1-0716-1538-6_10

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1537-9

  • Online ISBN: 978-1-0716-1538-6

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