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Evaluation of the Spindle Assembly Checkpoint Integrity in Mouse Oocytes
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The HECT E3 ligase Smurf2 is required for Mad2-dependent spindle assembly checkpoint
Evan C Osmundson1, Dipankar Ray, Finola E Moore
1Department of Molecular Pharmacology and Biological Chemistry, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA.
The Journal of Cell Biology
|October 15, 2008
Summary
Smurf2, a novel mitotic regulator, is essential for the spindle assembly checkpoint by controlling Mad2 stability and localization. Its absence causes premature cell division and chromosome misalignment.
Area of Science:
- Cell Biology
- Molecular Biology
- Genetics
Background:
- The metaphase-anaphase transition is regulated by the anaphase-promoting complex/cyclosome (APC/C) activated by Cdc20.
- The spindle assembly checkpoint ensures proper chromosome attachment before anaphase onset by delaying APC/C-Cdc20 activation.
Purpose of the Study:
- To investigate the role of the HECT ubiquitin ligase Smurf2 in the spindle assembly checkpoint.
- To elucidate the mechanism by which Smurf2 regulates mitotic progression.
Main Methods:
- Smurf2 depletion using siRNA.
- Expression of catalytically inactive Smurf2.
- Nocodazole treatment to induce spindle checkpoint arrest.
- Analysis of cyclin B, securin, and Mad2 levels and localization.
- Chromosome alignment and cytokinesis assays.
Main Results:
- Smurf2 depletion or inactivation caused chromosome misalignment, lagging chromosomes, premature anaphase, and failed cytokinesis.
- Smurf2 inactivation prevented prometaphase arrest and accumulation of cyclin B and securin in nocodazole-treated cells.
- Smurf2 depletion led to enhanced Mad2 degradation and mislocalization, indicating Smurf2 regulates Mad2 stability and localization.
Conclusions:
- Smurf2 is a novel and essential regulator of the spindle assembly checkpoint.
- Smurf2 controls mitotic progression by influencing the stability and localization of the checkpoint effector Mad2.
- Smurf2 plays a critical role in ensuring accurate chromosome segregation and cell division.
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