Ubiquitylation of cyclin E requires the sequential function of SCF complexes containing distinct hCdc4 isoforms

Cyclin E, an activator of cyclin-dependent kinase 2 (Cdk2), is targeted for proteasomal degradation by phosphorylation-dependent multiubiquitylation via the hCdC4 ubiquitin ligase SCF. SCF ubiquitin ligases are composed of a core of conserved subunits and one variable subunit (an F box protein) involved in substrate recognition. We show here that multi ubiquitylation of cyclin E requires the sequential function of two distinct splice variant isoforms of the F box protein hCdc4 known as alpha and gamma. SCFhCdc4 alpha binds a complex containing cyclin E, Cdk2, and the prolyl cis/trans isomerase Pin1 and promotes the activity of Pin1 without directly ubiquitylating cyclin E. However, due to the action of this SCFhCdc4 alpha-Pin1 complex, cyclin E becomes an efficient ubiquitylation substrate of SCFhCdc4 gamma. Furthermore, in the context of Cdc4 alpha and cyclin E, mutational data suggest that Pin1 isomerizes a noncanonical proline-proline bond, with the possibility that Cdc4 alpha may serve as a cofactor for altering the specificity of Pin1.