CCAN assembly configures composite binding interfaces to promote cross-linking of Ndc80 complexes at the kinetochore

Affiliation
Research Institute of Molecular Pathology (IMP), Dr. Bohr-Gasse 7, 1030 Vienna, Austria.
Pekgöz Altunkaya, Gülsah;
Affiliation
Research Institute of Molecular Pathology (IMP), Dr. Bohr-Gasse 7, 1030 Vienna, Austria.
Malvezzi, Francesca;
Affiliation
Research Institute of Molecular Pathology (IMP), Dr. Bohr-Gasse 7, 1030 Vienna, Austria.
Demianova, Zuzana;
Affiliation
Gene Center and Department of Biochemistry, Ludwig-Maximilians Universität München, Fedor-Lynen Strasse 25, 81377 Munich, Germany.
Zimniak, Tomasz;
Affiliation
Research Institute of Molecular Pathology (IMP), Dr. Bohr-Gasse 7, 1030 Vienna, Austria.
Litos, Gabriele;
ORCID
0000-0002-5174-6561
Affiliation
Research Institute of Molecular Pathology (IMP), Dr. Bohr-Gasse 7, 1030 Vienna, Austria.
Weissmann, Florian;
ORCID
0000-0002-3392-9946
Affiliation
Research Institute of Molecular Pathology (IMP), Dr. Bohr-Gasse 7, 1030 Vienna, Austria.
Mechtler, Karl;
Affiliation
Gene Center and Department of Biochemistry, Ludwig-Maximilians Universität München, Fedor-Lynen Strasse 25, 81377 Munich, Germany.
Herzog, Franz;
GND
123695961
LSF
57732
Affiliation
Research Institute of Molecular Pathology (IMP), Dr. Bohr-Gasse 7, 1030 Vienna, Austria; Department of Molecular Genetics, Faculty of Biology, Center for Medical Biotechnology, University of Duisburg-Essen, Universitätsstrasse 5, 45117 Essen, Germany. Electronic address: stefan.westermann@uni-due.de.
Westermann, Stefan

Partitioning of the genome requires kinetochores, large protein complexes that mediate dynamic attachment of chromosomes to the spindle. Kinetochores contain two supramolecular protein assemblies. The ten-protein KMN network harbors key microtubule-binding sites in the Ndc80 complex and mediates assembly of checkpoint complexes via the KNL-1/Spc105 protein [1, 2]. As KMN does not contact DNA directly, it relies on different centromere-binding proteins for recruitment and cell-cycle-dependent assembly. These proteins are collectively referred to as the CCAN (constitutive centromere-associated network) [2-4]. The molecular mechanisms by which CCAN subunits associate, however, have remained incompletely defined. In particular, it is unclear how CCAN subunits facilitate the assembly of a microtubule-binding interface that contains multiple Ndc80 molecules bound to different receptors [5]. Here, we dissect molecular mechanisms that underlie targeting of the CCAN subunit Cnn1/CENP-T to the sequence-determined point centromeres of budding yeast. Systematic quantitative mass spectrometry experiments reveal association dependencies within the yeast CCAN network. We show that evolutionarily conserved residues in the histone-fold domain of Cnn1 are required for the formation of a stable five-subunit CCAN subassembly with the Ctf3 complex. Cnn1 localizes in a Ctf3-dependent manner to the core of the yeast point centromere, overlapping with the yeast CENP-A protein Cse4. By arranging the N-terminal domains of the CCAN subunits Mcm16, Mcm22, and Cnn1 into close proximity, the Ctf3c-Cnn1-Wip1 complex configures a composite interaction site for two molecules of the Ndc80 complex. Our experiments show how cooperative assembly mechanisms organize the microtubule-binding interface of the kinetochore.

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Citation style:
Pekgöz Altunkaya, G., Malvezzi, F., Demianova, Z., Zimniak, T., Litos, G., Weissmann, F., Mechtler, K., Herzog, F., Westermann, S., 2016. CCAN assembly configures composite binding interfaces to promote cross-linking of Ndc80 complexes at the kinetochore. https://doi.org/10.1016/j.cub.2016.07.005
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