A novel mechanism for activation of the protein kinase Aurora A.
Segregation of chromosomes during mitosis requires interplay between several classes of protein on the spindle, including protein kinases, protein phosphatases, and microtubule binding motor proteins [1-4]. Aurora A is an oncogenic cell cycle-regulated protein kinase that is subject to phosphorylation-dependent activation [5-11]. Aurora A localization to the mitotic spindle depends on the motor binding protein TPX2 (Targeting Protein for Xenopus kinesin-like protein 2), but the protein(s) involved in Aurora A activation are unknown [11-13]. Here, we purify an activator of Aurora A from Xenopus eggs and identify it as TPX2. Remarkably, Aurora A that has been fully deactivated by Protein Phosphatase 2A (PP2A) becomes phosphorylated and reactivated by recombinant TPX2 in an ATP-dependent manner. Increased phosphorylation and activation of Aurora A requires its own kinase activity, suggesting that TPX2 stimulates autophosphorylation and autoactivation of the enzyme. Consistently, wild-type Aurora A, but not a kinase inactive mutant, becomes autophosphorylated on the regulatory T loop residue (Thr 295) after TPX2 treatment. Active Aurora A from bacteria is further activated at least 7-fold by recombinant TPX2, and TPX2 also impairs the ability of protein phosphatases to inactivate Aurora A in vitro. This concerted mechanism of stimulation of activation and inhibition of deactivation implies that TPX2 is the likely regulator of Aurora A activity at the mitotic spindle and may explain why loss of TPX2 in model systems perturbs spindle assembly [14-16]. Our finding that a known binding protein, and not a conventional protein kinase, is the relevant activator for Aurora A suggests a biochemical model in which the dynamic localization of TPX2 on mitotic structures directly modulates the activity of Aurora A for spindle assembly.
Eyers
Patrick A
PA
Howard Hughes Medical Institute and Department of Pharmacology, University of Colorado School of Medicine, Denver, CO 80262, USA.
Erikson
Eleanor
E
Chen
Lin G
LG
Maller
James L
JL
D000818:Animals; D064127:Aurora Kinases; D001345:Autoradiography; D018797:Cell Cycle Proteins; D020090:Chromosome Segregation / Q000502:physiology; D004793:Enzyme Reactivators / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008938:Mitosis / Q000502:physiology*; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D008941:Spindle Apparatus / Q000502:physiology*; D014981:Xenopus; D029867:Xenopus Proteins
D018797:Cell Cycle Proteins; D004793:Enzyme Reactivators; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; C486551:TPX2 protein, Xenopus; D029867:Xenopus Proteins; D011494:Protein Kinases; C112134:AURKA protein, Xenopus; D064127:Aurora Kinases; D017346:Protein Serine-Threonine Kinases