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Characterization of Pch2 localization determinants reveals a nucleolar-independent role in the meiotic recombination checkpoint

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Abstract

The meiotic recombination checkpoint blocks meiotic cell cycle progression in response to synapsis and/or recombination defects to prevent aberrant chromosome segregation. The evolutionarily conserved budding yeast Pch2TRIP13 AAA+ ATPase participates in this pathway by supporting phosphorylation of the Hop1HORMAD adaptor at T318. In the wild type, Pch2 localizes to synapsed chromosomes and to the unsynapsed rDNA region (nucleolus), excluding Hop1. In contrast, in synaptonemal complex (SC)–defective zip1Δ mutants, which undergo checkpoint activation, Pch2 is detected only on the nucleolus. Alterations in some epigenetic marks that lead to Pch2 dispersion from the nucleolus suppress zip1Δ-induced checkpoint arrest. These observations have led to the notion that Pch2 nucleolar localization could be important for the meiotic recombination checkpoint. Here we investigate how Pch2 chromosomal distribution impacts checkpoint function. We have generated and characterized several mutations that alter Pch2 localization pattern resulting in aberrant Hop1 distribution and compromised meiotic checkpoint response. Besides the AAA+ signature, we have identified a basic motif in the extended N-terminal domain critical for Pch2’s checkpoint function and localization. We have also examined the functional relevance of the described Orc1-Pch2 interaction. Both proteins colocalize in the rDNA, and Orc1 depletion during meiotic prophase prevents Pch2 targeting to the rDNA allowing unwanted Hop1 accumulation on this region. However, Pch2 association with SC components remains intact in the absence of Orc1. We finally show that checkpoint activation is not affected by the lack of Orc1 demonstrating that, in contrast to previous hypotheses, nucleolar localization of Pch2 is actually dispensable for the meiotic checkpoint.

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Acknowledgements

We are grateful to David Ontoso, Andrés Clemente, and Shirleen Roeder for reagents. We also thank Isabel Acosta and Sara González-Arranz for the technical assistance, Carlos Vázquez for the advice on microscopy analysis, and José Pérez-Martín and Andrés Clemente for the helpful discussions and ideas.

Funding

This work was supported by grants from the Ministry of Economy and Competitiveness (MINECO) of Spain to JAC and PSS (grants BFU2015-64361-P and BFU2015-65417-R, respectively). EH was supported by a predoctoral contract (FPU1502035) from the Ministry of Education of Spain. JAC is supported by a Ramón y Cajal contract (RYC2013-13950). The IBFG is funded in part by an institutional grant from Junta de Castilla y León (CLU-2017-03).

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  1. Instituto de Biología Funcional y Genómica (IBFG), Consejo Superior de Investigaciones Científicas (CSIC) and University of Salamanca, 37007, Salamanca, Spain

    Esther Herruzo, Beatriz Santos & Pedro A. San-Segundo

  2. Departamento de Microbiología y Genética, University of Salamanca, 37007, Salamanca, Spain

    Beatriz Santos

  3. Instituto de Tecnologías Biomédicas, Hospital Universitario de Canarias, 38320, La Laguna, Tenerife, Spain

    Raimundo Freire

  4. Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas (CSIC), 28040, Madrid, Spain

    Jesús A. Carballo

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  1. Esther Herruzo
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  5. Pedro A. San-Segundo
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Correspondence to Pedro A. San-Segundo.

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This article is part of a Special Issue on Recent advances in meiosis from DNA replication to chromosome segregation “edited by Valérie Borde and Francesca Cole, co-edited by Paula Cohen and Scott Keeney.”

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Herruzo, E., Santos, B., Freire, R. et al. Characterization of Pch2 localization determinants reveals a nucleolar-independent role in the meiotic recombination checkpoint. Chromosoma 128, 297–316 (2019). https://doi.org/10.1007/s00412-019-00696-7

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