Abstract
The centromere is essential to the propagation of eukaryotic cells as it allows sister chromatids to segregate during mitosis. It is thus important to understand how this region responds to DNA damage and which proteins are involved. Double-strand breaks are one type of DNA damage that can occur in a cell. In Saccharomyces cerevisiae, most double-strand breaks are repaired through homologous recombination, which involves the generation of single-stranded DNA. This process likely disrupts centromere function when DNA damage occurs near centromeres. To understand how cells restore centromere function after DNA damage, I induced a double-strand break and monitored product formation. I then performed chromatin immunoprecipitation using an antibody against the kinetochore protein Mif2 to monitor centromere restoration. De novo centromere establishment is known to require CHL4, which encodes an outer kinetochore protein. To test whether CHL4 is required for centromere re-establishment after DNA damage, I deleted CHL4 and compared the phenotypes with wild type. This allowed me to understand the effect of CHL4 and to create a system that can monitor the effect of other proteins on centromere repair after a double-strand break. I showed that the deletion of CHL4 did not affect cell viability and demonstrated that CHL4 has little impact on centromere repair. However, this study did not rule out the possibility that CHL4 affects centromere repair independently of Mif2. Although this study only monitored the impact of CHL4, this system will be useful for testing other proteins to understand the mechanism of centromere re-establishment after DNA damage.