Cdk2-dependent phosphorylation of p27 facilitates its Myc-induced release from cyclin E/cdk2 complexes

Area of Science:

• Cell cycle regulation
• Oncogenesis

Background:

• Myc activation induces cyclin E/cdk2 activity and p27 degradation.
• p27 dissociation from cyclin E/cdk2 precedes its degradation.
• p27 interacts with both cyclin E/cdk2 and cyclin D/cdk4 complexes.

Purpose of the Study:

• To investigate the mechanism of p27 release from cyclin E/cdk2.
• To determine the role of p27 phosphorylation in its dissociation from cyclin E/cdk2.
• To elucidate the role of T187 phosphorylation in Myc-induced p27 degradation.

Main Methods:

• In vitro kinase assays using cyclin E/cdk2 and p27.
• Site-directed mutagenesis of p27 at T187.
• In vivo studies in Myc-transformed cells to assess p27 phosphorylation and complex formation.

Main Results:

• Cyclin E/cdk2 phosphorylates p27 at threonine 187 (T187) in vitro.
• Mutation of T187 to valine stabilizes cyclin E/cdk2 complexes, indicating active cdk2.
• In vivo, p27 bound to cyclins E and A, but not D-type cyclins, is phosphorylated at T187.
• Myc-induced p27 release from cdk2 requires cdk2 kinase activity and is delayed by the T187V mutation.
• Phosphorylated p27 transiently accumulates in a non-cdk2 bound form after Myc induction.

Conclusions:

• p27 is released from cyclin E/cdk2 upon phosphorylation at T187.
• This phosphorylation-dependent release is crucial for p27 degradation in Myc-transformed cells.
• Phosphorylated p27 transiently forms a complex independent of cdk2 before degradation.