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A Mass Spectrometry-Based Approach to Identify Phosphoprotein Phosphatases and their Interactors
Published on: April 29, 2022
Protein phosphatase 2A and separase form a complex regulated by separase autocleavage
Andrew J Holland1, Franziska Böttger, Olaf Stemmann
1Faculty of Life Sciences, Michael Smith Building, Oxford Road, University of Manchester, Manchester M13 9PT, United Kingdom.
The Journal of Biological Chemistry
|July 3, 2007
Summary
Separase self-cleavage is essential for cell division, as it disrupts the binding of protein phosphatase 2A (PP2A). This interaction regulates centromeric cohesion, ensuring proper sister chromatid separation during anaphase.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Anaphase onset relies on separase activation, a protease that cleaves cohesins to separate sister chromatids.
- Separase undergoes self-cleavage, but its biological role remains unclear.
- Separase activity is inhibited by securin and Cdk1/cyclin B1 before anaphase.
Purpose of the Study:
- To elucidate the biological significance of separase self-cleavage.
- To identify novel separase interacting partners and their roles in regulating separase activity.
Main Methods:
- Co-immunoprecipitation assays to identify separase-binding proteins.
- Site-directed mutagenesis to create non-cleavable separase variants.
- Analysis of centromeric cohesion in cells expressing separase mutants.
Main Results:
- Protein phosphatase 2A (PP2A) was identified as a novel separase-interacting partner, binding via its B' (B56) subunit.
- PP2A association requires a domain near separase autocleavage sites; cleavage disrupts this interaction.
- Expression of non-cleavable separase, but not a PP2A-binding deficient mutant, caused premature loss of centromeric cohesion.
Conclusions:
- Separase self-cleavage regulates its interaction with PP2A.
- This regulation is crucial for controlling centromeric cohesion and ensuring accurate chromosome segregation during anaphase.
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