FIGURE 5.

Cyclin A/cdk2 associates with and phosphorylates APC. A, representation of APC. The arrowheads represent the RXL cyclin A binding sites, and the asterisks represent the cdk phosphorylation sites. The Ser-1100 and Ser-1360 sites are indicated as are the GST-APC truncations used in the in vitro phosphorylation studies. The position of the mutation cluster region (MCR) is also indicated. B, cyclin A immunoprecipitates from HeLa cells harvested at times indicated post-thymidine release, immunoblotted for APC. The levels of cyclin A and α-tubulin (loading control) were determined by immunoblotting of the whole cell lysate. C, immunoprecipitated cyclin A complexes were assayed for kinase activity. The phosphorylated band was identified as APC by immunoblotting. The level of phosphorylation relative to control antibody (-Ab) is shown in the bar graph. The percentage of cells in mitosis was determined by MPM2 staining and represented in the line graph.