CEP192 localises mitotic Aurora-A activity by priming its interaction with TPX2

Aurora-A is an essential cell-cycle kinase with critical roles in mitotic entry and spindle dynamics. These functions require binding partners such as CEP192 and TPX2, which modulate both kinase activity and localisation of Aurora-A. Here we investigate the structure and role of the centrosomal Aurora-A:CEP192 complex in the wider molecular network. We find that CEP192 wraps around Aurora-A, occupies the binding sites for mitotic spindle-associated partners, and thus competes with them. Comparison of two different Aurora-A conformations reveals how CEP192 modifies kinase activity through the site used for TPX2-mediated activation. Deleting the Aurora-A-binding interface in CEP192 prevents centrosomal accumulation of Aurora-A, curtails its activation-loop phosphorylation, and reduces spindle-bound TPX2:Aurora-A complexes, resulting in error-prone mitosis. Thus, by supplying the pool of phosphorylated Aurora-A necessary for TPX2 binding, CEP192:Aurora-A complexes regulate spindle function. We propose an evolutionarily conserved spatial hierarchy, which protects genome integrity through fine-tuning and correctly localising Aurora-A activity. The mechanisms underlying the spatiotemporal control of Aurora-A kinase activity during mitosis are not yet determined. This study elucidates both the structure of the human Aurora-A:CEP192 complex and its critical role at the apex of Aurora-A activation in mitosis.CEP192 wraps around the N-lobe of the Aurora-A kinase domain occupying sites that are typically used by the competing microtubule-associated partners TACC3 and TPX2.CEP192 binds the F-pocket/C-helix region of the N-lobe in a way that inhibits Aurora-A, whereas TPX2 binding to the same region activates the kinase.The centrosomal CEP192-Aurora-A complex is the primary driver of Aurora-A auto-phosphorylation during mitosis.Deleting the Aurora-A binding interface of CEP192 causes mitotic spindle defects reminiscent of Aurora-A inhibition.CEP192 facilitates Aurora-A:TPX2 complex formation and activity on the mitotic spindle. The structure of the human Aurora-A:CEP192 complex reveals its importance in facilitating Aurora-A autophosphorylation and activity on the mitotic spindle.