title: The pseudo GTPase CENP-M drives human kinetochore assembly.
abstract: Kinetochores, multi-subunit complexes that assemble at the interface with centromeres, bind spindle microtubules to ensure faithful delivery of chromosomes during cell division. The configuration and function of the kinetochore-centromere interface is poorly understood. We report that a protein at this interface, CENP-M, is structurally and evolutionarily related to small GTPases but is incapable of GTP-binding and conformational switching. We show that CENP-M is crucially required for the assembly and stability of a tetramer also comprising CENP-I, CENP-H, and CENP-K, the HIKM complex, which we extensively characterize through a combination of structural, biochemical, and cell biological approaches. A point mutant affecting the CENP-M/CENP-I interaction hampers kinetochore assembly and chromosome alignment and prevents kinetochore recruitment of the CENP-T/W complex, questioning a role of CENP-T/W as founder of an independent axis of kinetochore assembly. Our studies identify a single pathway having CENP-C as founder, and CENP-H/I/K/M and CENP-T/W as CENP-C-dependent followers.DOI: http://dx.doi.org/10.7554/eLife.02978.001.
authors.affiliation: Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Federica
authors.identifier: http://orcid.org/0000-0002-7002-6316
authors.initials: F
authors.lastname: Basilico
authors.affiliation: Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: Stefano
authors.initials: S
authors.lastname: Maffini
authors.affiliation: Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: John R
authors.initials: JR
authors.lastname: Weir
authors.affiliation: Department of Physical Biochemistry, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: Daniel
authors.initials: D
authors.lastname: Prumbaum
authors.affiliation: Computational Biology and Bioinformatics Group, Institute of Biomedicine of Seville, Campus Hospital Universitario Virgen del Rocio, Seville, Spain.
authors.forename: Ana M
authors.initials: AM
authors.lastname: Rojas
authors.affiliation: Department of Biochemistry and Gene Center, Ludwig-Maximilians-Universität, München, Munich, Germany.
authors.forename: Tomasz
authors.initials: T
authors.lastname: Zimniak
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Anna
authors.initials: A
authors.lastname: De Antoni
authors.affiliation: Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: Sadasivam
authors.initials: S
authors.lastname: Jeganathan
authors.affiliation: Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: Beate
authors.initials: B
authors.lastname: Voss
authors.affiliation: Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: Suzan
authors.initials: S
authors.lastname: van Gerwen
authors.affiliation: Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Veronica
authors.initials: V
authors.lastname: Krenn
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Lucia
authors.initials: L
authors.lastname: Massimiliano
authors.affiliation: Structural Biology and Biocomputing Programme, Spanish National Cancer Centre-CNIO, Madrid, Spain.
authors.forename: Alfonso
authors.initials: A
authors.lastname: Valencia
authors.affiliation: Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: Ingrid R
authors.initials: IR
authors.lastname: Vetter
authors.affiliation: Department of Biochemistry and Gene Center, Ludwig-Maximilians-Universität, München, Munich, Germany.
authors.forename: Franz
authors.initials: F
authors.lastname: Herzog
authors.affiliation: Department of Physical Biochemistry, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Raunser
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Sebastiano
authors.initials: S
authors.lastname: Pasqualato
authors.affiliation: Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany Centre for Medical Biotechnology, Faculty of Biology, University of Duisburg-Essen, Essen, Germany andrea.musacchio@mpi-dortmund.mpg.de.
authors.forename: Andrea
authors.initials: A
authors.lastname: Musacchio
mesh_terms: D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D020558:GTP Phosphohydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000737:chemistry / Q000378:metabolism*; D008954:Models, Biological; D008958:Models, Molecular; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017510:Protein Folding; D055550:Protein Stability; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D034741:RNA, Small Interfering / Q000235:genetics; D017386:Sequence Homology, Amino Acid
chemical_list: C495719:CENPM protein, human; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D046912:Multiprotein Complexes; D009687:Nuclear Proteins; D021122:Protein Subunits; D034741:RNA, Small Interfering; D020558:GTP Phosphohydrolases
keywords: centromeres; kinetochores; mitosis