Centromeres in the thermotolerant yeast K. marxianus mediate attachment to a single microtubule

TL;DR

This study shows that the yeast K. marxianus uses a single centromeric nucleosome to attach chromosomes to one microtubule during cell division.

Contribution

The study reveals that K. marxianus centromeres mediate attachment to a single microtubule despite having a longer centromere than S. cerevisiae.

Findings

K. marxianus kinetochores localize similarly to S. cerevisiae throughout the cell cycle.

Electron tomography shows each chromosome in K. marxianus is bound by a single microtubule.

Nucleosome mapping suggests only one centromeric nucleosome is present in K. marxianus.

Abstract

Eukaryotic chromosome segregation requires spindle microtubules to attach to chromosomes through kinetochores. The chromosomal locus that mediates kinetochore assembly is the centromere and is epigenetically specified in most organisms by a centromeric histone H3 variant called CENP-A. An exception to this is budding yeast which have short, sequenced-defined point centromeres. In S. cerevisiae, a single CENP-A nucleosome is formed at the centromere and is sufficient for kinetochore assembly. The thermophilic budding yeast Kluyveromyces marxianus also has a point centromere but its length is nearly double the S. cerevisiae centromere and the number of centromeric nucleosomes and kinetochore attachment sites is unknown. Purification of native kinetochores from K. marxianus yielded a mixed population, with one subpopulation that appeared to consist of doublets, making it unclear whether K. marxianus shares the same attachment architecture as S. cerevisiae. Here, we demonstrate that though the doublet kinetochores have a functional impact on kinetochore strength, kinetochore localization throughout the cell cycle appears conserved between these two yeasts. In addition, whole spindle electron tomography demonstrates that a single microtubule binds to each chromosome. Single-molecule nucleosome mapping analysis suggests the presence of a single centromeric nucleosome. Taken together, we propose that the K. marxianus point centromere assembles a single centromeric nucleosome that mediates attachment to one microtubule.

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