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

The formation of the mitotic spindle is a critical step in the equal separation of DNA from the mother cell to the two daughter cells. Cytoskeleton Associated Protein-2 (CKAP2) is a microtubule polymerase that associates with microtubules and centrosomes during interphase and mitosis. Loss of CKAP2 results in chromosomal instability and onset of aneuploidy. To identify proteins interacting with CKAP2, we immunoprecipitated endogenous CKAP2 together with interacting proteins from mitotic RPE1 cells and analysed the samples by mass spectrometry. This experiment identified the mitotic protein kinase Aurora Kinase A, together with its cofactor TPX2, as interacting proteins of CKAP2. Notably, CKAP2 co-localizes and interacts with Aurora Kinase A and TPX2 throughout mitosis, but not with the related protein Aurora Kinase B. Consistently, Aurora Kinase A phosphorylates CKAP2 both in mitotic cells and in vitro . Phosphorylated CKAP2 has reduced affinity for microtubules in vitro . Importantly, expression of Aurora A phospho-mimic mutants of CKAP2 in RPE1 cells results in phenotypes of a small mitotic spindle and reduced microtubule stability. Thus, Aurora Kinase A phosphorylates CKAP2 to regulate mitotic spindle growth and stability.