CCAN Assembly Configures Composite Binding Interfaces to Promote Cross-Linking of Ndc80 Complexes at the Kinetochore

Altunkaya, Gülsah Pekgöz; Malvezzi, Francesca; Demianova, Zuzana; Zimniak, Tomasz; Litos, Gabriele; Weissmann, Florian; Mechtler, Karl; Herzog, Franz und Westermann, Stefan (2016): CCAN Assembly Configures Composite Binding Interfaces to Promote Cross-Linking of Ndc80 Complexes at the Kinetochore. In: Current Biology, Bd. 26, Nr. 17: S. 2370-2378

Abstract

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 his tone -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 Cnnl 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.

• Dokumententyp: Zeitschriftenartikel
• Fakultät: Chemie und Pharmazie > Department Biochemie
• Themengebiete: 500 Naturwissenschaften und Mathematik > 540 Chemie
• ISSN: 0960-9822
• Sprache: Englisch