Ubiquitylation of Cyclin E Requires the Sequential Function of SCF Complexes Containing Distinct hCdc4 Isoforms

SummaryCyclin E, an activator of cyclin-dependent kinase 2 (Cdk2), is targeted for proteasomal degradation by phosphorylation-dependent multiubiquitylation via the ubiquitin ligase SCFhCdc4. 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 multiubiquitylation of cyclin E requires the sequential function of two distinct splice variant isoforms of the F box protein hCdc4 known as α and γ. SCFhCdc4α 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 SCFhCdc4α-Pin1 complex, cyclin E becomes an efficient ubiquitylation substrate of SCFhCdc4γ. Furthermore, in the context of Cdc4α and cyclin E, mutational data suggest that Pin1 isomerizes a noncanonical proline-proline bond, with the possibility that Cdc4α may serve as a cofactor for altering the specificity of Pin1.