Martin Singleton
Structural studies of eukaryotic chromosome segregation
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Kinetochores are the proteinaceous complexes that form the connection between chromosomes and spindle microtubules during mitosis and meiosis. In addition to anchoring the centromeric DNA to the highly dynamic microtubules, parts of the kinetochore are responsible for generating the so-called spindle assembly checkpoint (SAC). This ensures that all chromatids are correctly attached to the spindle and the anaphase to metaphase transition does not occur prematurely. Kinetochores are extremely large and complex structures, but can be broken down into distinct sub-complexes, which are amenable to more detailed study.
We are examining some of these sub-complexes from both budding yeast and human kinetochores in order to better understand the assembly and overall architecture of the kinetochore using X-ray crystallography. But determining the three-dimensional structures of the complexes and their constituent proteins, we can gain insight into their function and potential interactions within the kinetochore.
One particular area of interest is the manner in which the inner layers of the kinetochore recognise and bind to the correct site on the chromosome. The exact nature of the centromere varies considerably between species, but a common feature is the presence of modified nucleosomes, which are required to direct kinetochore formation. How this occurs, what specifies the location of the modified nucleosomes, and how the components of the kinetochore interact with them is not yet known.
We aim to answer these questions by both biochemical and biophysical techniques. By determining the composition and stoichiometry of the protein complexes involved, and working out their three-dimensional structures and interactions with chromatin, we can begin to understand this elaborate protein-DNA interface and answer some of these questions.
References
Westermann S, Drubin D, Barnes G. Structures and functions of yeast kinetochore complexes. Annu Rev Biochem 2007; 76: 563-591.
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