Cks promotes recruitment of cyclin B1 as an APC/C substrate. (A) Cells were cotransfected with cyclin B1–cerulean and either Cdk1*–EYFP or Cdk1*–1N–EYFP. A cell undergoing mitotic exit without cytokinesis is shown. DIC, differential interference contrast. Bar, 5 µM. (B) Reduced APC/C^Cdc20 binding of cyclin B1–Cdk1–1N complexes in cells, whereas total APC/C phosphorylation is normal. Checkpoint-arrested mitotic cells selected for expression of indicated constructs were collected. GFP IPs (pulling down Cdk1–EYFP) on equalized extracts were blotted for coprecipitation of the indicated proteins. Cyclin B1–Cherry is not recognized by the anti-GFP antibodies. WCE, whole cell extract. (C) Quantitative comparison of destruction times of cyclin B1 bound to Cdk1 (white) or Cdk1–1N (light gray). The destruction times of the cells expressing Cdk1–1N, which failed cytokinesis, are presented in the dark gray box. The plots contain data from three independent experiments (see Fig. S2 B for destruction curve). Markers are given in kilodaltons.