An Aurora Kinase A/TPX2 complex phosphorylates CKAP2 to control mitotic spindle growth

Faithful chromosome segregation requires the precise assembly of the mitotic spindle. Cytoskeleton-associated protein 2 (CKAP2) is a microtubule-associated protein with potent microtubule-polymerizing activity that localizes to spindle microtubules during mitosis. Loss or overexpression of CKAP2 causes chromosomal instability and aneuploidy, yet its regulation remains poorly understood. To identify CKAP2 interactors, we immunoprecipitated endogenous CKAP2 from mitotic RPE1 cells and analysed co-purifying proteins by mass spectrometry. This revealed a specific interaction between CKAP2 and the mitotic kinase Aurora A and its activator TPX2, but not with Aurora B, as previously suggested. We further show that CKAP2 co-localizes with Aurora A-TPX2 complexes throughout mitosis, and Aurora A directly phosphorylates CKAP2 in cells and in vitro. Phosphorylation of CKAP2 By Aurora kinase A decreases its microtubule affinity in cells and in vitro. These findings identify CKAP2 as a direct interactor of TPX2 and a substrate of Aurora kinase A and uncover a regulatory pathway controlling spindle growth and stability during mitosis.