title: Aurora-A phosphorylates Augmin complex component Hice1 protein at an N-terminal serine/threonine cluster to modulate its microtubule binding activity during spindle assembly.
abstract: Proper assembly of mitotic spindles requires Hice1, a spindle-associated protein. Hice1 possesses direct microtubule binding activity at its N-terminal region and contributes to intraspindle microtubule nucleation as a subunit of the Augmin complex. However, whether microtubule binding activity of Hice1 is modulated by mitotic regulators remains unexplored. Here, we found that Aurora-A kinase, a major mitotic kinase, specifically binds to and phosphorylates Hice1. We identified four serine/threonine clusters on Hice1 that can be phosphorylated by Aurora-A in vitro. Of the four clusters, the Ser/Thr-17-21 cluster was the most critical for bipolar spindle assembly, whereas other phospho-deficient point mutants had a minimal effect on spindle assembly. Immunostaining with a phospho-Ser-19/20 phospho-specific antibody revealed that phosphorylated Hice1 primarily localizes to spindle poles during prophase to metaphase but gradually diminishes after anaphase. Consistently, the phospho-mimic 17-21E mutant reduced microtubule binding activity in vitro and diminished localization to spindles in vivo. Furthermore, expression of the 17-21E mutant led to decreased association of Fam29a, an Augmin component, with spindles. On the other hand, expression of the phospho-deficient 17-21A mutant permitted intraspindle nucleation but delayed the separation of early mitotic spindle poles and the timely mitotic progression. Taken together, these results suggest that Aurora-A modulates the microtubule binding activity of Hice1 in a spatiotemporal manner for proper bipolar spindle assembly.
authors.affiliation: Department of Biological Chemistry, University of California, Irvine, California 92697-4037, USA.
authors.forename: Connie Y
authors.initials: CY
authors.lastname: Tsai
authors.forename: Bryan
authors.initials: B
authors.lastname: Ngo
authors.forename: Anjali
authors.initials: A
authors.lastname: Tapadia
authors.forename: Pang-Hung
authors.initials: PH
authors.lastname: Hsu
authors.forename: Guikai
authors.initials: G
authors.lastname: Wu
authors.forename: Wen-Hwa
authors.initials: WH
authors.lastname: Lee
mesh_terms: D064127:Aurora Kinases; D045744:Cell Line, Tumor; D006801:Humans; D008677:Metaphase / Q000502:physiology*; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010766:Phosphorylation / Q000502:physiology; D011418:Prophase / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D008941:Spindle Apparatus / Q000235:genetics / Q000378:metabolism*
chemical_list: C000628759:HAUS8 protein, human; D008869:Microtubule-Associated Proteins; D064127:Aurora Kinases; D017346:Protein Serine-Threonine Kinases