Figure 7.
Mutations in the hydrophobic patch of cyclin A alter both its binding to Cdk2 and its localization. (A) Localization. HeLa cells were transfected with myc epitope-tagged cyclin A (M210A, L214A, W217A) alone (top) or with CDK2 (bottom). Cells were fixed and processed for immunofluorescence using the anti-myc epitope antibody. (B) Cdk binding. Myc epitope-tagged cyclin A and cyclin A hydrophobic patch mutants were transfected into HeLa cells. Eighteen hours after transfection, cell lysates were incubated with p9Cks1-Sepharose beads. The amounts of myc-cyclin A and myc-cyclin A HPM bound to p9Cks1 was compared by immunoblotting using an anti-myc epitope antibody. Top, the amounts of myc-cyclin A and myc-cyclin A hydrophobic patch mutant expressed in each cell lysate.