Abstract
Chromosome orientation and alignment within the mitotic spindle requires the Aurora B protein kinase and the mitotic centromere-associated kinesin (MCAK). Here, we report the regulation of MCAK by Aurora B. Aurora B inhibited MCAK's microtubule depolymerizing activity in vitro, and phospho-mimic (S/E) mutants of MCAK inhibited depolymerization in vivo. Expression of either MCAK (S/E) or MCAK (S/A) mutants increased the frequency of syntelic microtubule-kinetochore attachments and mono-oriented chromosomes. MCAK phosphorylation also regulates MCAK localization: the MCAK (S/E) mutant frequently localized to the inner centromere while the (S/A) mutant concentrated at kinetochores. We also detected two different binding sites for MCAK using FRAP analysis of the different MCAK mutants. Moreover, disruption of Aurora B function by expression of a kinase-dead mutant or RNAi prevented centromeric targeting of MCAK. These results link Aurora B activity to MCAK function, with Aurora B regulating MCAK's activity and its localization at the centromere and kinetochore.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Amino Acid Sequence
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Animals
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Antineoplastic Agents, Phytogenic / pharmacology
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Aurora Kinase B
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Aurora Kinases
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Autoradiography
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CHO Cells
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Cell Cycle / drug effects
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Cell Cycle / physiology*
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Centromere / metabolism*
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Chromatography, High Pressure Liquid
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Chromosomes / metabolism
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Cricetinae
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Electrophoresis, Gel, Two-Dimensional
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Fluorescence Recovery After Photobleaching / methods
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Fluorescent Antibody Technique / methods
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Green Fluorescent Proteins
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HeLa Cells
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Humans
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In Vitro Techniques
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Kinesins / metabolism*
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Kinetochores / metabolism
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Luminescent Proteins / metabolism
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Mitosis / drug effects
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Mitosis / physiology*
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Models, Biological
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Mutation
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Nocodazole / pharmacology
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Paclitaxel / pharmacology
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Phosphorylation
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Protein Serine-Threonine Kinases / metabolism*
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RNA, Small Interfering / pharmacology
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Rats
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Transfection
Substances
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Antineoplastic Agents, Phytogenic
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KIF2C protein, human
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Luminescent Proteins
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RNA, Small Interfering
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mitotic centromere-associated kinesin, hamster
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Green Fluorescent Proteins
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AURKB protein, human
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Aurkb protein, rat
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Aurora Kinase B
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Aurora Kinases
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Protein Serine-Threonine Kinases
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Kinesins
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Paclitaxel
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Nocodazole