Structure and Function of a Eukaryotic Centromere

Project Summary
Goals: The centromere serves as the binding site for the kinetochore and is essential for the
faithful segregation of chromosomes throughout cell division. The point centromere in yeast is
encoded by a ~115 bp specific DNA sequence, whereas regional centromeres span 6-10 kbp in
fission yeast to 5-10 Mbp in human. Despite the apparent diversity in centromere organization,
the distance between sister kinetochores in metaphase ranges from 800 nm to 1,000 nm in
yeast, worms, flies, flower moths, plants, horses and human. Understanding the physical
structure of centromere chromatin (pericentromere in yeast, defined as the chromatin between
sister kinetochores) will provide fundamental insights how centromere DNA is organized into a
stiff spring that resists microtubule pulling forces during mitosis.
Approach: Our laboratory develops computational tools to interrogate the structure and
dynamics of hundreds of kilobase pairs of pericentromeric DNA. Together with experimentally
obtained images of fluorescent probes of pericentromeric structure (e.g. pericentromere DNA,
cohesin, condensin) we make quantitative comparisons between simulations and experimental
results through transformation of in silico models into microscope images (model convolution).
We will test the proposal that the mechanism for building tension between sister kinetochores is
a chromatin bottlebrush organized by the loop-extruding proteins condensin and cohesin. The
bottlebrush provides a biophysical mechanism that transforms pericentromeric chromatin into a
spring due to the steric repulsion between radial loops. The bottlebrush as an organizing
principle for chromosome organization has emerged from multiple approaches in the field. We
will leverage the powerful features of chromosome engineering in yeast to explore the
consequences of reducing the number of centromeres, and exploit synthetic bottlebrushes and
statistical physics of polymer models to reveal basic principles linking bottlebrush structure to
the functional readout of force/tension.
Innovation: We will combine our experience in chromosome engineering and advanced
bioimaging in yeast with the expertise of collaborators in statistical physics and applied math
(Forest UNC-CH) and synthetic bio-inspired materials (Freeman UNC-CH). Testing our
hypotheses will elucidate important information about the organization and function of
centromeres, potentially providing a paradigm shifting foundation for the remarkable
conservation of distance between sister kinetochores throughout phylogeny.

Grant Number: 5R01GM032238-38
NIH Institute/Center: NIH
Principal Investigator: Kerry Bloom