An advanced cell cycle tag toolbox reveals principles underlying temporal control of structure-selective nucleases
- Max Planck Institute of Biochemistry, Germany
- ETH Zurich, Switzerland
- ETH Zürich, Switzerland
Abstract
Cell cycle tags allow to restrict target protein expression to specific cell cycle phases. Here, we present an advanced toolbox of cell cycle tag constructs in budding yeast with defined and compatible peak expression that allow comparison of protein functionality at different cell cycle phases. We apply this technology to the question of how and when Mus81-Mms4 and Yen1 nucleases act on DNA replication or recombination structures. Restriction of Mus81-Mms4 to M phase but not S phase allows a wildtype response to various forms of replication perturbation and DNA damage in S phase, suggesting it acts as a post-replicative resolvase. Moreover, we use cell cycle tags to reinstall cell cycle control to a deregulated version of Yen1, showing that its premature activation interferes with the response to perturbed replication. Curbing resolvase activity and establishing a hierarchy of resolution mechanisms are therefore the principal reasons underlying resolvase cell cycle regulation.
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All data generated or analysed during this study are included in the manuscript and supporting files.
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Author details
Funding
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung
- Joao Matos
Eidgenössische Technische Hochschule Zürich
- Joao Matos
Max-Planck-Gesellschaft
- Boris Pfander
Deutsche Forschungsgemeinschaft
- Boris Pfander
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2020, Bittmann et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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An advanced cell cycle tag toolbox reveals principles underlying temporal control of structure-selective nucleases
eLife 9:e52459.
https://doi.org/10.7554/eLife.52459
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