In budding yeast, a single cenH3 (Cse4) nucleosome occupies the ∼120-bp functional centromere, however conflicting structural models for the particle have been proposed. To resolve this controversy, we have applied H4S47C-anchored cleavage mapping, which reveals the precise position of histone H4 in every nucleosome in the genome. We find that cleavage patterns at centromeres are unique within the genome and are incompatible with symmetrical structures, including octameric nucleosomes and (Cse4/H4)2 tetrasomes. Centromere cleavage patterns are compatible with a precisely positioned core structure, one in which each of the 16 yeast centromeres is occupied by oppositely oriented Cse4/H4/H2A/H2B hemisomes in two rotational phases within the population. Centromere-specific hemisomes are also inferred from distances observed between closely-spaced H4 cleavages, as predicted from structural modeling. Our results indicate that the orientation and rotational position of the stable hemisome at each yeast centromere is not specified by the functional centromere sequence.
DNA is tightly packaged in cells for a variety of reasons—to allow it to fit inside the nucleus, to protect it from damage, and to help control the production of proteins from genes. The basic unit of packaged DNA is called a nucleosome, which consists of DNA wrapped around a structure formed by two pairs of four different proteins.
These proteins, which are called histones, have a role that extends beyond providing structural support for DNA. When cells divide, for example, pairs of ‘sister chromosomes’ are pulled apart to ensure that the two daughter cells both have the same chromosomes as the original cell. The sister chromosomes are pulled apart from a single position called a centromere, and the nucleosomes at this position contain a histone that is different from the histones found everywhere else in the cell. However, until recently it was not clear if the nucleosomes that contained these special cenH3 histones had the same structure as other nucleosomes.
Now Henikoff et al. have used a method called H4S47C-anchored cleavage mapping to study every nucleosome in the genome of the yeast S. cerevisiae. This mapping technique uses DNA sequencing to measure the precise distances between fixed points on the DNA in the nucleosome. Knowing these distances tells researchers a great deal about the number and position of the histones within each nucleosome in the genome.
Using this approach, Henikoff et al. found that nucleosomes at centromeres are different from other nucleosomes in histone number and arrangement. In particular, the nucleosome at each yeast centromere contains only one each of the four different histones in an asymmetrical orientation, in contrast to all other yeast nucleosomes, which contain two sets of four histones in a symmetrical arrangement. Furthermore, each nucleosome at a centromere can adopt one of two orientations: these orientations are mirror images of each other, and they occur with equal probability. It should also be possible to use the mapping technique developed by Henikoff et al. to study the larger and more complex centromeres found in other organisms, including humans.