Figure 4.

Human Cdc20 and aurora2/Aik associate in HeLa cells. (A) HeLa cells were synchronized at the indicated stages and extracts produced as described in the text. Thirty micrograms of each extract was analyzed by Western blotting for cell cycle regulated proteins cyclin A, cyclin B1, hCdc20, or aurora2/Aik. As indicates asynchronously dividing cells. (B) For immunoprecipitate kinase assays, extracts were incubated with either preimmune serum (−) or with antibodies to hCdc20 (+, Upper) or aurora2/Aik (+, Lower); immune complexes were recovered and assayed for kinase activity toward MBP, as described in the text. Phosphorylated MBP was detected by SDS/PAGE followed by autoradiography or PhosphorImaging. (C) Cdc20 or aurora2/Aik immunoprecipitates were prepared as in B, eluted from washed beads, precipitated, and analyzed by SDS/PAGE followed by immunoblotting. Immunoprecipitates were probed for human Cdc20 (Upper) or aurora2/Aik (Lower). Arrows indicate the positions of human Cdc20 and aurora2/Aik.