Phospho‐regulated Bim1/EB1 interactions trigger Dam1c ring assembly at the budding yeast outer kinetochore

• Published in: The EMBO journal Vol. 40; no. 18; pp. e108004 - n/a
• Main Authors: Dudziak, Alexander, Engelhard, Lena, Bourque, Cole, Klink, Björn Udo, Rombaut, Pascaline, Kornakov, Nikolay, Jänen, Karolin, Herzog, Franz, Gatsogiannis, Christos, Westermann, Stefan
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 15.09.2021; Blackwell Publishing Ltd; John Wiley and Sons Inc
• Subjects: Assembling; Assembly; Attachment; Bik1; Bim1; bi‐orientation; Chromosome Segregation; Chromosomes; Complex formation; Conserved sequence; Crosslinking; Dam1/DASH complex; EMBO06; EMBO31; Error correction; Homology; Kinases; Kinetochores; Kinetochores - metabolism; Localization; Metaphase; Microtubule Proteins - metabolism; Microtubule-Associated Proteins - metabolism; Microtubules; Mitosis; Mitosis - physiology; Multiprotein Complexes - metabolism; Oligomers; Phosphorylation; Protein Binding; Proteins; Rings (mathematics); Saccharomycetales - physiology; Spindle Apparatus - metabolism; Yeast; Yeasts; bi‐orientation; Bik1; Bim1; chromosome segregation; Dam1/DASH complex; bi-orientation
• ISSN: 0261-4189; 1460-2075
• DOI: 10.15252/embj.2021108004

Kinetochores form the link between chromosomes and microtubules of the mitotic spindle. The heterodecameric Dam1 complex (Dam1c) is a major component of the Saccharomyces cerevisiae outer kinetochore, assembling into 3 MDa‐sized microtubule‐embracing rings, but how ring assembly is specifically initiated in vivo remains to be understood. Here, we describe a molecular pathway that provides local control of ring assembly during the establishment of sister kinetochore bi‐orientation. We show that Dam1c and the general microtubule plus end‐associated protein (+TIP) Bim1/EB1 form a stable complex depending on a conserved motif in the Duo1 subunit of Dam1c. EM analyses reveal that Bim1 crosslinks protrusion domains of adjacent Dam1c heterodecamers and promotes the formation of oligomers with defined curvature. Disruption of the Dam1c‐Bim1 interaction impairs kinetochore localization of Dam1c in metaphase and delays mitosis. Phosphorylation promotes Dam1c‐Bim1 binding by relieving an intramolecular inhibition of the Dam1 C‐terminus. In addition, Bim1 recruits Bik1/CLIP‐170 to Dam1c and induces formation of full rings even in the absence of microtubules. Our data help to explain how new kinetochore end‐on attachments are formed during the process of attachment error correction. SYNOPSIS Kinetochore‐microtubule attachment for budding yeast chromosome segregation requires ring formation of the Dam1 complex (Dam1c). A combination of genetic, biochemical and structural approaches reveals how association of microtubule plus‐end associated proteins triggers formation of Dam1c rings. Plus end‐binding protein Bim1 binds and crosslinks the protrusion domains of Dam1c. Interaction with Bim1 is required for efficient Dam1c kinetochore loading in metaphase and for timely mitotic progression. Mps1 kinase activity promotes Dam1c‐Bim1 complex formation. Bim1 cooperates with CLIP‐170 homolog Bik1 to induce Dam1c ring assembly in vitro independent of microtubules. The Dam1c‐Bim1‐Bik1 complex might represent an early configuration during end‐on attachment formation. Graphical Abstract A combination of genetic, biochemical and structural approaches reveals how phosphorylation‐regulated association of microtubule plus‐end associated proteins facilitates formation of complexes for end‐on spindle attachment.