Dynamic association of a tumor amplified kinase, Aurora-A, with the centrosome and mitotic spindle.
Aurora-A kinase, also known as STK15/BTAK kinase, is a member of a serine/threonine kinase superfamily that includes the prototypic yeast Ipl1 and Drosophila aurora kinases as well as other mammalian and non-mammalian aurora kinases involved in the regulation of centrosomes and chromosome segregation. The Aurora-A gene is amplified and overexpressed in a wide variety of human tumors. Aurora-A is centrosome-associated during interphase, and binds the poles and half-spindle during mitosis; its over-expression has been associated with centrosome amplification and multipolar spindles. GFP-Aurora-A was used to mark centrosomes and spindles, and monitor their movements in living cells. Centrosome pairs labeled with GFP-Aurora-A are motile throughout interphase undergoing oscillations and tumbling motions requiring intact microtubules and ATP. Fluorescence recovery after photobleaching (FRAP) was used to examine the relative molecular mobility of GFP-Aurora-A, and GFP-labeled alpha-tubulin, gamma-tubulin, and NuMA. GFP-Aurora-A rapidly exchanges in and out of the centrosome and mitotic spindle (t(1/2) approximately 3 sec); in contrast, both tubulins are relatively immobile indicative of a structural role. GFP-NuMA mobility was intermediate in both interphase nuclei and at the mitotic spindle (t(1/2) approximately 23-30 sec). Deletion mapping identifies a central domain of Aurora-A as essential for its centrosomal localization that is augmented by both the amino and the carboxyl terminal ends of the protein. Interestingly, amino or carboxy terminal deletion mutants that maintained centrosomal targeting exhibited significantly slower molecular exchange. Collectively, these studies contrast the relative cellular dynamics of Aurora-A with other cytoskeletal proteins that share its micro-domains, and identify essential regions required for targeting and dynamics.
Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas 77030, USA. stenoien@bcm.tmc.edu
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D034961:Antigens, Nuclear; D064096:Aurora Kinase A; D064127:Aurora Kinases; D018797:Cell Cycle Proteins; D002455:Cell Division / Q000235:genetics; D002465:Cell Movement / Q000235:genetics; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000378:metabolism; D018385:Centrosome / Q000201:enzymology*; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins; D009154:Mutation / Q000235:genetics; D009369:Neoplasms / Q000201:enzymology* / Q000235:genetics; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins / Q000378:metabolism; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D017434:Protein Structure, Tertiary / Q000235:genetics; D021381:Protein Transport / Q000235:genetics / Q000502:physiology; D011993:Recombinant Fusion Proteins; D008941:Spindle Apparatus / Q000201:enzymology*; D014404:Tubulin / Q000378:metabolism; D029867:Xenopus Proteins
D034961:Antigens, Nuclear; D018797:Cell Cycle Proteins; D008164:Luminescent Proteins; C050045:NUMA1 protein, human; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D014404:Tubulin; D029867:Xenopus Proteins; D049452:Green Fluorescent Proteins; D011494:Protein Kinases; C112134:AURKA protein, Xenopus; C576917:AURKA protein, human; D064096:Aurora Kinase A; D064127:Aurora Kinases; D017346:Protein Serine-Threonine Kinases