1. UNIVERSITY OF SUSSEX
Catrina Anne Miles
Doctor of Philosophy (Genome Damage and Stability)
An investigation into factors affecting condensin association with mitotic centromeres
SUMMARY
The SMC protein family (Structural Maintenance of Chromosomes) consists a group of highly
conserved protein complexes, central to chromosome dynamics and key cell cycle events.
Condensin is a member of the SMC protein family, best known for its role in chromosome
condensation and segregation in mitosis. The condensin complex has been shown to
become enriched at specific chromosome loci in a cell-cycle specific manner, however the
details of how it becomes associated with chromatin remain unclear. A particular area of
interest regarding condensin association and activity is at the centromeres and
pericentromeres, where condensin has been consistently shown to be enriched specifically
duringmitosis.
This work is comprised of four results chapters investigating factors affecting condensin
association with mitotic centromeres in budding yeast, using chromatin
immunoprecipitation (ChIP). We started by establishing a robust ChIP assay suitable for
probing condensin enrichment at the centromeric regions, and conducting genetic control
experiments to ensure the functionality of the experimental technique. In the next chapter
we explored the importance of the kinetochore with regards to condensin enrichment, and
found that perturbing the budding yeast kinetochore results in a loss of centromeric
condensin association during mitosis. We then used condensin phosphorylation-site and
mitotic kinase mutants to examine the role of condensin subunit phosphorylation in its
association with chromatin. Our result showed that Ipl1 (aurora kinase) and condensin
phosphorylation is important for its enrichment at the centromere, but rather surprisingly
that Cdc5 (polo-like kinase) a known activator of condensin does not appear to be. The final
chapter investigates the function of condensin’s intrinsic ATPase activity, and we found that
ATP-binding activity but not ATP-hydrolysis is important for condensin association at mitotic
centromeres.