Cryo-EM structure of the complete inner kinetochore of the budding yeast point centromere.

Dendooven, Tom; Zhang, Ziguo; Yang, Jing; McLaughlin, Stephen H; Schwab, Johannes; Scheres, Sjors H W; Yatskevich, Stanislau; Barford, David · Sci Adv · 2023

basic_science · Level V

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Abstract

The point centromere of budding yeast specifies assembly of the large kinetochore complex to mediate chromatid segregation. Kinetochores comprise the centromere-associated inner kinetochore (CCAN) complex and the microtubule-binding outer kinetochore KNL1-MIS12-NDC80 (KMN) network. The budding yeast inner kinetochore also contains the DNA binding centromere-binding factor 1 (CBF1) and CBF3 complexes. We determined the cryo-electron microscopy structure of the yeast inner kinetochore assembled onto the centromere-specific centromere protein A nucleosomes (CENP-A<sup>Nuc</sup>). This revealed a central CENP-A<sup>Nuc</sup> with extensively unwrapped DNA ends. These free DNA duplexes bind two CCAN protomers, one of which entraps DNA topologically, positioned on the centromere DNA element I (CDEI) motif by CBF1. The two CCAN protomers are linked through CBF3 forming an arch-like configuration. With a structural mechanism for how CENP-A<sup>Nuc</sup> can also be linked to KMN involving only CENP-QU, we present a model for inner kinetochore assembly onto a point centromere and how it organizes the outer kinetochore for chromosome attachment to the mitotic spindle.

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