An advanced cell cycle tag toolbox reveals principles underlying temporal control of structure-selective nucleases

Julia Bittmann, Rokas Grigaitis, Lorenzo Galanti, Silas Amarell, Florian Wilfling, Joao Matos, Boris Pfander
2020 eLife  
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
more » ... ldtype 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.
doi:10.7554/elife.52459 fatcat:35m2hdtvbzfqhesioy745wnkjq