Activation of Aurora-A kinase by protein phosphatase inhibitor-2, a bifunctional signaling protein.
Aurora-A kinase is necessary for centrosome maturation, for assembly and maintenance of a bipolar spindle, and for proper chromosome segregation during cell division. Aurora-A is an oncogene that is overexpressed in multiple human cancers. Regulation of kinase activity apparently depends on phosphorylation of Thr-288 in the T-loop. In addition, interactions with targeting protein for Xenopus kinesin-like protein 2 (TPX2) allosterically activate Aurora-A. The Thr-288 phosphorylation is reversed by type-1 protein phosphatase (PP1). Mutations in the yeast Aurora, Ipl1, are suppressed by overexpression of Glc8, the yeast homolog of phosphatase inhibitor-2 (I-2). In this study, we show that human I-2 directly and specifically stimulated recombinant human Aurora-A activity in vitro. The I-2 increase in kinase activity was not simply due to inhibition of PP1 because it was not mimicked by other phosphatase inhibitors. Furthermore, activation of Aurora-A was unaffected by deletion of the I-2 N-terminal PP1 binding motif but was eliminated by deletion of the I-2 C-terminal domain. Aurora-A and I-2 were recovered together from mitotic HeLa cells. Kinase activation by I-2 and TPX2 was not additive and occurred without a corresponding increase in T-loop phosphorylation. These results suggest that both I-2 and TPX2 function as allosteric activators of Aurora-A. This implies that I-2 is a bifunctional signaling protein with separate domains to inhibit PP1 and directly stimulate Aurora-A kinase.
Satinover
David L
DL
Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, VA 22908, USA.
Leach
Craig A
CA
Stukenberg
P Todd
PT
Brautigan
David L
DL
D000818:Animals; D064127:Aurora Kinases; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D066298:In Vitro Techniques; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D054623:Receptor-Like Protein Tyrosine Phosphatases, Class 2; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism; D000097847:Protein Phosphatase Inhibitory Proteins
D064127:Aurora Kinases; D018797:Cell Cycle Proteins; D004791:Enzyme Inhibitors; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D010749:Phosphoprotein Phosphatases; D010750:Phosphoproteins; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases; D011506:Proteins; D054623:Receptor-Like Protein Tyrosine Phosphatases, Class 2; D011994:Recombinant Proteins; D029867:Xenopus Proteins; C486551:TPX2 protein, Xenopus; C122167:TPX2 protein, human; C024833:protein phosphatase inhibitor-2; D000097847:Protein Phosphatase Inhibitory Proteins; C112134:AURKA protein, Xenopus; C516868:PTPRU protein, human