Polo-like kinase 4 homodimerization is not required for catalytic activation, autodestruction, or centriole assembly

Abstract

Polo-like kinase 4 (Plk4) is the master-regulator of centriole assembly and cell cycle-dependent regulation of its activity maintains proper centrosome number. During most of the cell cycle, Plk4 levels are nearly undetectable due to its ability to autophosphorylate and trigger its own ubiquitin-mediated degradation. However, during mitotic exit, Plk4 forms a single aggregate on the centriole surface to stimulate centriole duplication. Whereas most Polo-like kinase family members are monomeric, Plk4 is unique because it forms homodimers. Notably, Plk4 trans-autophosphorylates a degron near its kinase domain, a critical step in autodestruction. While it is thought that the purpose of homodimerization is to promote trans-autophosphorylation, this has not been tested. Here, we generate separation-of-function Plk4 mutants that fail to dimerize, and we show that homodimerization is required to create a binding site for the Plk4 activator, Asterless. Surprisingly, Plk4 dimer mutants are catalytically active in cells, promote centriole assembly, and can trans-autophosphorylate by a process based its on concentration-dependent aggregation. Our findings implicate a concentration-dependent pathway of Plk4 activation that does not require Asterless binding or homodimerization. Lastly, we propose a model of Plk4 cell cycle regulation that utilizes both activation pathways - Asterless-dependent and aggregation-driven - to restrict centriole assembly to mother centrioles.