title: Molecular architecture of the Dam1 complex-microtubule interaction.
abstract: Mitosis is a highly regulated process that allows the equal distribution of the genetic material to the daughter cells. Chromosome segregation requires the formation of a bipolar mitotic spindle and assembly of a multi-protein structure termed the kinetochore to mediate attachments between condensed chromosomes and spindle microtubules. In budding yeast, a single microtubule attaches to each kinetochore, necessitating robustness and processivity of this kinetochore-microtubule attachment. The yeast kinetochore-localized Dam1 complex forms a direct interaction with the spindle microtubule. In vitro, the Dam1 complex assembles as a ring around microtubules and couples microtubule depolymerization with cargo movement. However, the subunit organization within the Dam1 complex, its higher-order oligomerization and how it interacts with microtubules remain under debate. Here, we used chemical cross-linking and mass spectrometry to define the architecture and subunit organization of the Dam1 complex. This work reveals that both the C termini of Duo1 and Dam1 subunits interact with the microtubule and are critical for microtubule binding of the Dam1 complex, placing Duo1 and Dam1 on the inside of the ring structure. Integrating this information with available structural data, we provide a coherent model for how the Dam1 complex self-assembles around microtubules.
authors.affiliation: Wellcome Trust Centre for Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.
authors.forename: Thibault
authors.initials: T
authors.lastname: Legal
authors.affiliation: Wellcome Trust Centre for Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.
authors.forename: Juan
authors.initials: J
authors.lastname: Zou
authors.affiliation: Wellcome Trust Centre for Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.
authors.forename: Alicja
authors.initials: A
authors.lastname: Sochaj
authors.affiliation: Wellcome Trust Centre for Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK Department of Bioanalytics, Institute of Biotechnology, Technische Universität Berlin, Berlin, Germany.
authors.forename: Juri
authors.initials: J
authors.lastname: Rappsilber
authors.affiliation: Wellcome Trust Centre for Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK julie.welburn@ed.ac.uk.
authors.forename: Julie P I
authors.identifier: http://orcid.org/0000-0002-5440-6060
authors.initials: JP
authors.lastname: Welburn
mesh_terms: D005656:Fungal Proteins / Q000032:analysis / Q000378:metabolism*; D018386:Kinetochores / Q000737:chemistry / Q000378:metabolism*; D008870:Microtubules / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D011485:Protein Binding; D060066:Protein Interaction Maps; D021122:Protein Subunits / Q000032:analysis / Q000378:metabolism; D004718:Saccharomycetales / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D021122:Protein Subunits
keywords: Dam1 complex; kinetochore; microtubule; mitosis; self-assembly; three-dimensional MS cross-linking