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

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(Nuc)). This revealed a central CENP-A(Nuc) 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(Nuc) 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.

Automated summary

The point centromere of budding yeast controls the assembly of large kinetochore complex for chromatid segregation. The kinetochore complex consists of the inner kinetochore (CCAN) and the outer kinetochore KNL1-MIS12-NDC80 (KMN) network. The cryo-electron microscopy structure reveals the assembly of the yeast inner kinetochore onto the centromere-specific CENP-A nucleosomes with unwrapped DNA ends. The CCAN protomers bind the free DNA duplexes and are connected through CBF3. A model is presented for the assembly of the inner kinetochore onto a point centromere and its organization of the outer kinetochore for chromosome attachment to the mitotic spindle.