1. Cell Biology
Download icon

A FRET-based study reveals site-specific regulation of spindle position checkpoint proteins at yeast centrosomes

  1. Yuliya Gryaznova
  2. Ayse Koca Caydasi
  3. Gabriele Malengo
  4. Victor Sourjik
  5. Gislene Pereira  Is a corresponding author
  1. German Cancer Research Centre, Germany
  2. Centre for Molecular Biology, Germany
Research Article
  • Cited 21
  • Views 1,909
  • Annotations
Cite this article as: eLife 2016;5:e14029 doi: 10.7554/eLife.14029

Abstract

The spindle position checkpoint (SPOC) is a spindle pole body (SPB, equivalent of mammalian centrosome) associated surveillance mechanism that halts mitotic exit upon spindle mis-orientation. Here, we monitored the interaction between SPB proteins and the SPOC component Bfa1 by FRET microscopy. We show that Bfa1 binds to the scaffold-protein Nud1 and the γ-tubulin receptor Spc72. Spindle misalignment specifically disrupts Bfa1-Spc72 interaction by a mechanism that requires the 14-3-3-family protein Bmh1 and the MARK/PAR-kinase Kin4. Dissociation of Bfa1 from Spc72 prevents the inhibitory phosphorylation of Bfa1 by the polo-like kinase Cdc5. We propose Spc72 as a regulatory hub that coordinates the activity of Kin4 and Cdc5 towards Bfa1. In addition, analysis of spc72∆ cells shows that a mitotic-exit-promoting dominant signal, which is triggered upon elongation of the spindle into the bud, overrides the SPOC. Our data reinforce the importance of daughter-cell-associated factors and centrosome-based regulations in mitotic exit and SPOC control.

Article and author information

Author details

  1. Yuliya Gryaznova

    DKFZ-ZMBH Alliance, German Cancer Research Centre, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.

  2. Ayse Koca Caydasi

    DKFZ-ZMBH Alliance, German Cancer Research Centre, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.

  3. Gabriele Malengo

    DKFZ-ZMBH Alliance, Centre for Molecular Biology, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.

  4. Victor Sourjik

    DKFZ-ZMBH Alliance, Centre for Molecular Biology, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.

  5. Gislene Pereira

    DKFZ-ZMBH Alliance, German Cancer Research Centre, Heidelberg, Germany
    For correspondence
    gislene.pereira@cos.uni-heidelberg.de
    Competing interests
    The authors declare that no competing interests exist.

Reviewing Editor

  1. Yukiko M Yamashita, University of Michigan, United States

Publication history

  1. Received: December 23, 2015
  2. Accepted: May 8, 2016
  3. Accepted Manuscript published: May 9, 2016 (version 1)
  4. Accepted Manuscript updated: May 20, 2016 (version 2)
  5. Version of Record published: May 24, 2016 (version 3)
  6. Version of Record updated: June 3, 2016 (version 4)

Copyright

© 2016, Gryaznova 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.

Metrics

  • 1,909
    Page views
  • 435
    Downloads
  • 21
    Citations

Article citation count generated by polling the highest count across the following sources: Scopus, Crossref, PubMed Central.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)

Categories and tags

Research organism

    1. Related to

    Spatial signals link exit from mitosis to spindle position

    Jill Elaine Falk et al.
  2. Further reading

Further reading

    1. Cell Biology

    Spatial signals link exit from mitosis to spindle position

    Jill Elaine Falk et al.
    Research Article Updated

    In budding yeast, if the spindle becomes mispositioned, cells prevent exit from mitosis by inhibiting the mitotic exit network (MEN). The MEN is a signaling cascade that localizes to spindle pole bodies (SPBs) and activates the phosphatase Cdc14. There are two competing models that explain MEN regulation by spindle position. In the 'zone model', exit from mitosis occurs when a MEN-bearing SPB enters the bud. The 'cMT-bud neck model' posits that cytoplasmic microtubule (cMT)-bud neck interactions prevent MEN activity. Here we find that 1) eliminating cMT– bud neck interactions does not trigger exit from mitosis and 2) loss of these interactions does not precede Cdc14 activation. Furthermore, using binucleate cells, we show that exit from mitosis occurs when one SPB enters the bud despite the presence of a mispositioned spindle. We conclude that exit from mitosis is triggered by a correctly positioned spindle rather than inhibited by improper spindle position.

    1. Cell Biology
    2. Chromosomes and Gene Expression

    Repressing Ago2 mRNA translation by Trim71 maintains pluripotency through inhibiting let-7 microRNAs

    Qiuying Liu et al.
    Research Article Updated

    The regulation of stem cell fate is poorly understood. Genetic studies in Caenorhabditis elegans lead to the hypothesis that a conserved cytoplasmic double-negative feedback loop consisting of the RNA-binding protein Trim71 and the let-7 microRNA controls the pluripotency and differentiation of stem cells. Although let-7-microRNA-mediated inhibition of Trim71 promotes differentiation, whether and how Trim71 regulates pluripotency and inhibits the let-7 microRNA are still unknown. Here, we show that Trim71 represses Ago2 mRNA translation in mouse embryonic stem cells. Blocking this repression leads to a specific post-transcriptional increase of mature let-7 microRNAs, resulting in let-7-dependent stemness defects and accelerated differentiation in the stem cells. These results not only support the Trim71-let-7-microRNA bi-stable switch model in controlling stem cell fate, but also reveal that repressing the conserved pro-differentiation let-7 microRNAs at the mature microRNA level by Ago2 availability is critical to maintaining pluripotency.

    1. Cell Biology
    2. Developmental Biology

    A Non-stop identity complex (NIC) supervises enterocyte identity and protects from pre-mature aging

    Neta Erez et al.
    Research Article

    A hallmark of aging is loss of differentiated cell identity. Aged Drosophila midgut differentiated enterocytes (ECs) lose their identity, impairing tissue homeostasis. To discover identity regulators, we performed an RNAi screen targeting ubiquitin-related genes in ECs. Seventeen genes were identified, including the deubiquitinase Non-stop (CG4166). Lineage tracing established that acute loss of Non-stop in young ECs phenocopies aged ECs at cellular and tissue levels. Proteomic analysis unveiled that Non-stop maintains identity as part of a Non-stop identity complex (NIC) containing E(y)2, Sgf11, Cp190, (Mod) mdg4, and Nup98. Non-stop ensured chromatin accessibility, maintaining the EC-gene signature, and protected NIC subunit stability. Upon aging, the levels of Non-stop and NIC subunits declined, distorting the unique organization of the EC nucleus<strong>.</strong> Maintaining youthful levels of Non-stop in wildtype aged ECs safeguards NIC subunits, nuclear organization, and suppressed aging phenotypes. Thus, Non-stop and NIC, supervise EC identity and protects from premature aging.