Phosphorylation- and polo-box-dependent binding of Plk1 to Bub1 is required for the kinetochore localization of Plk1

Polo-like kinase 1 (PIk1) is required for the generation of the tension-sensing 3F3/2 kinetochore epitope and facilitates kinetochore localization of Mad2 and other spindle checkpoint proteins. Here, we investigate the mechanism by which PIk1 itself is recruited to kinetochores. We show that PIk1 binds to budding uninhibited by benzimidazole 1 (Bub1) in mitotic human cells. The PIk1-Bub1 interaction requires the polo-box domain (PBD) of PIk1 and is enhanced by cyclin-dependent kinase 1 (Cdk1)-mediated phosphorylation of Bub1 at T609. The PBD-dependent binding of PIk1 to Bub1 facilitates phosphorylation of Bub1 by PIk1 in vitro. Depletion of Bub1 in HeLa cells by RNA interference (RNAi) diminishes the kinetochore localization of PIk1. Ectopic expression of the wild-type Bub1, but not the Bub1-T609A mutant, in Bub1-RNAi cells restores the kinetochore localization of PIk1. Our results suggest that phosphorylation of Bub1 at T609 by Cdk1 creates a docking site for the PBD of PIk1 and facilitates the kinetochore recruitment of PIk1.