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Evaluation of the Spindle Assembly Checkpoint Integrity in Mouse Oocytes
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The Bub1-TPR Domain Interacts Directly with Mad3 to Generate Robust Spindle Checkpoint Arrest.

Ioanna Leontiou1, Nitobe London2, Karen M May1

  • 1Institute of Cell Biology, University of Edinburgh, King's Buildings, Max Born Crescent, Edinburgh EH9 3BF, UK.

Current Biology : CB
|July 2, 2019
PubMed
Summary

Researchers bypassed the normal spindle checkpoint pathway in yeast by inducing Mps1 kinase and Bub1 protein interactions. This artificial activation of the spindle checkpoint, dependent on Mad1, Mad2, and Mad3, reveals key domains in Bub1 essential for mitotic arrest.

Keywords:
Bub1Mad3Mps1TPR domainmitosisspindle checkpoint

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Area of Science:

  • Cell Biology
  • Molecular Biology
  • Genetics

Background:

  • The spindle checkpoint ensures accurate chromosome segregation by delaying anaphase onset when kinetochore-microtubule attachments are incorrect.
  • This checkpoint is activated by Mps1 kinase phosphorylating KNL1 on MELT motifs, recruiting Bub1-Bub3 complexes, which then recruit Mad1-Mad2 to form a signaling platform.
  • The platform facilitates the assembly of the Mitotic Checkpoint Complex (MCC), which inhibits the APC/C, delaying anaphase.

Purpose of the Study:

  • To investigate whether the spindle checkpoint can be activated independently of kinetochore-mediated signaling.
  • To identify the specific domains of Bub1 required for spindle checkpoint activation and MCC formation.
  • To dissect the roles of Bub1's tetratricopeptide repeat (TPR) domain in protein interactions during mitosis.

Main Methods:

  • Induction of heterodimers between Mps1 kinase and Bub1 in budding and fission yeast.
  • Analysis of metaphase arrest dependency on Mad1, Mad2, and Mad3.
  • Domain dissection of Bub1 to identify regions essential for checkpoint activation and protein binding.

Main Results:

  • Artificial induction of Mps1-Bub1 heterodimers bypassed the need for kinetochore-localized KNL1 and triggered a metaphase arrest.
  • This arrest was dependent on Mad1, Mad2, and Mad3, indicating MCC formation.
  • Bub1's tetratricopeptide repeat (TPR) domain was found to be both necessary and sufficient for binding Mad3 and recruiting it to the signaling complex.

Conclusions:

  • The spindle checkpoint can be artificially activated by directly linking Mps1 kinase activity to Bub1, independent of upstream kinetochore signals.
  • Bub1's TPR domain plays a crucial role in recruiting Mad3, facilitating MCC assembly and mitotic arrest.
  • This study provides insights into the minimal requirements for spindle checkpoint activation and the function of Bub1 domains in chromosome segregation fidelity.