title: The NIMA-family kinase Nek6 phosphorylates the kinesin Eg5 at a novel site necessary for mitotic spindle formation.
abstract: Nek6 and Nercc1 (also known as Nek9) belong to the NIMA family of protein kinases. Nercc1 is activated in mitosis, whereupon it binds, phosphorylates and activates Nek6. Interference with Nek6 or Nercc1 in mammalian cells causes prometaphase-metaphase arrest, and depletion of Nercc1 from Xenopus egg extracts prevents normal spindle assembly. Herein we show that Nek6 is constitutively associated with Eg5 (also known as Kinesin-5 and Kif11), a kinesin that is necessary for spindle bipolarity. Nek6 phosphorylated Eg5 at several sites in vitro and one of these sites, Ser1033, is phosphorylated in vivo during mitosis. Whereas CDK1 phosphorylates nearly all Eg5 at Thr926 during mitosis, Nek6 phosphorylates approximately 3% of Eg5, primarily at the spindle poles. Eg5 depletion caused mitotic arrest, resulting in cells with a monopolar spindle. This arrest could be rescued by wild-type Eg5 but not by Eg5[Thr926Ala]. Despite substantial overexpression, Eg5[Ser1033Ala] rescued 50% of cells compared with wild-type Eg5, whereas an Eg5[Ser1033Asp] mutant was nearly as effective as wild type. Thus, during mitosis Nek6 phosphorylates a subset of Eg5 polypeptides at a conserved site, the phosphorylation of which is crucial for the mitotic function of Eg5.
authors.affiliation: Department of Molecular Biology and Medical Services, Massachusetts General Hospital and Department of Medicine, Harvard Medical School, Boston, MA 02114, USA.
authors.forename: Joseph
authors.initials: J
authors.lastname: Rapley
authors.forename: Marta
authors.initials: M
authors.lastname: Nicolàs
authors.forename: Aaron
authors.initials: A
authors.lastname: Groen
authors.forename: Laura
authors.initials: L
authors.lastname: Regué
authors.forename: M Teresa
authors.initials: MT
authors.lastname: Bertran
authors.forename: Carme
authors.initials: C
authors.lastname: Caelles
authors.forename: Joseph
authors.initials: J
authors.lastname: Avruch
authors.forename: Joan
authors.initials: J
authors.lastname: Roig
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D006367:HeLa Cells; D006801:Humans; D016547:Kinesins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D008941:Spindle Apparatus / Q000378:metabolism*; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: C088335:KIF11 protein, Xenopus; D011994:Recombinant Proteins; D029867:Xenopus Proteins; D017346:Protein Serine-Threonine Kinases; D016547:Kinesins