Centromere Structure and Function

Abstract

The centromere is the genetic locus that specifies the site of kinetochore assembly, where the chromosome will attach to the kinetochore microtubule. The pericentromere is the physical region responsible for the geometry of bi-oriented sister kinetochores in metaphase. In budding yeast the 125 bp point centromere is sufficient to specify kinetochore assembly. The flanking region is enriched (3X) in cohesin and condensin relative to the remaining chromosome arms. The enrichment spans about 30-50 kb around each centromere. We refer to the flanking chromatin as the pericentromere in yeast. In mammals, a 5-10 Mb region dictates where the kinetochore is built. The kinetochore interacts with a very small fraction of DNA on the surface of the centromeric region. The remainder of the centromere lies between the sister kinetochores. This is typically called centromere chromatin. The chromatin sites that directly interface to microtubules cannot be identified due to the repeated sequence within the mammalian centromere. However in both yeast and mammals, the total amount of DNA between the sites of microtubule attachment in metaphase is highly conserved. In yeast the 16 chromosomes are clustered into a 250 nm diameter region, and 800 kb (16 × 50 kb) or ~1 Mb of DNA lies between sister kinetochores. In mammals, 5-10 Mb lies between sister kinetochores. In both organisms the sister kinetochores are separated by about 1 μm. Thus, centromeres of different organisms differ in how they specify kinetochore assembly, but there may be important centromere chromatin functions that are conserved throughout phylogeny. Recently, centromeric chromatin has been reconstituted in vitro using alpha satellite DNA revealing unexpected features of centromeric DNA organization, replication, and response to stress. We will focus on the conserved features of centromere in this review.

Fig. 1

A DNA basket on the surface of centromeres in metaphase. Left The pericentromere region of all 16 chromosomes in metaphase in budding yeast. The 125 bp CEN region (pink nucleosome at right-most edge of the nucleosome fiber, depicted as yellow DNA wrapped around red histones) lie at the apex of the pericentric chromatin loops (11 nm fiber, yellow strands). The centromere DNA containing loops extend perpendicular to the chromosome axis (Lawrimore et al. 2016). Middle panels Top: end-on view of the Cse4 containing nucleosomes, one from each centromere of the 16 sister chromatids are cylindrically arranged and lie on the surface of the chromosome. Bottom: side view of centromere DNA in metaphase. 80 bp of DNA is in direct contact with the Cse4-containing core (pink), flanking DNA follows a trajectory toward the kinetochore (yellow strands away from the pericentromere). DNA devoid of histones reflect the DNAase I hypersensitive regions (Bloom and Carbon 1982; Bloom et al. 1983) exiting and entering the Cse4 containing nucleosome that protrude from the surface of the chromosome to make a basket. Far right The Cse4 containing nucleosomes are proximal to the pericentric chromatin (yellow DNA strands, bottom), while the DNAase I hypersensitive sites protrude from the chromosome surface toward the kinetochore (top). Sister kinetochores lie ⁓800 nm away on the opposite surface of the sister strands

Fig. 2

Configuration of pericentric chromatin loops surrounding the spindle axis in the budding yeast. The blue spheres are spindle pole bodies, the green rods are kinetochore microtubules. The interpolar microtubules can be seen as blue rods extending about ¾ the length of the spindle from each pole. The yellow strands are pericentric chromatin. The primary loop (horizontal) is attached to a kinetochore microtubule. Chromosome arms (not shown) would extend north and south, from approximately the middle of the pericentromere. Condensin is at the base of the each of the radial subloops (purple staples). Cohesin are the red rings. The position of cohesin is the most thermodynamically favorable and matches the position observed experimentally with the size and number of loops modeled herein

Fig. 3

Electron micrograph showing partially digested chromatin isolated from Xenopus laevis egg extract incubated with human alpha satellite DNA. Loops of double stranded DNA filaments running parallel to each other embedded in a protein matrix can be appreciated. Electron dense particles can be noticed at the base of some of the loops. Bar corresponds to 500 base pairs

Fig. 4

Isw1a, a member of the SWI/SNF chromatin remodeling family functions as a loop extruding enzyme in vitro, adapted from (De Cian et al. 2012). Isw1 interacts with the yeast Cbf1 factor where it functions to maintain nucleosome-free regions at promoters. This and other ATPases at centromere could collectively promote pericentromeric looping