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Manipulation and Analysis of Cell Cycle-Dependent Processes in Budding Yeast

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Interphase APC/C-Cdc20 inhibition by cyclin A2-Cdk2 ensures efficient mitotic entry.

Jamin B Hein¹, Jakob Nilsson¹

¹The Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Blegdamsvej 3B, 2200 Copenhagen, Denmark.

Nature Communications

|March 11, 2016

Summary

The Anaphase Promoting Complex/Cyclosome (APC/C) is regulated by cyclin A2-Cdk2 during interphase, preventing premature activation and ensuring proper cell division. This discovery reveals a novel positive feedback loop crucial for cell cycle progression.

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Area of Science:

Cell Biology
Molecular Biology
Biochemistry

Background:

Cell cycle progression relies on precise regulation of the Anaphase Promoting Complex/Cyclosome (APC/C).
The APC/C is activated by co-activators Cdh1 or Cdc20.
The regulation of APC/C-Cdc20 during interphase remains largely unknown.

Purpose of the Study:

To investigate the regulation of APC/C-Cdc20 activity during interphase.
To elucidate the role of cyclin A2-Cdk2 in controlling APC/C-Cdc20 during interphase.
To uncover novel feedback mechanisms governing cell cycle progression.

Main Methods:

In vitro binding and phosphorylation assays to study protein interactions.
Analysis of APC/C-Cdc20 activity in response to Cdc20 phosphorylation.
Cell cycle analysis using non-degradable cyclin variants to assess mitotic entry.

Main Results:

Cyclin A2-Cdk2 directly binds and phosphorylates Cdc20 during interphase, inhibiting APC/C-Cdc20 activity.
Inhibition of Cdc20 phosphorylation leads to premature APC/C-Cdc20 activation and destabilization of key cell cycle substrates like cyclin A2 and B1.
This premature activation results in a prolonged G2 phase and delayed entry into mitosis.
Expression of non-degradable cyclin A2, but not cyclin B1, rescues the mitotic entry defect.

Conclusions:

A novel positive feedback loop involving cyclin A2-Cdk2 inhibition of interphase APC/C-Cdc20 has been identified.
This mechanism allows for the accumulation of cyclin A2, promoting timely entry into mitosis.
The findings provide new insights into the intricate regulation of cell cycle control and the APC/C complex.