Since all eukaryotes and few prokaryotes keep their genomes in the form of linear DNA molecules, these organisms depend on an intact telomere structure both to maintain chromosome length due to the semi-conservative nature of DNA replication, as well as to differentiate the chromosome ends from the DSB repair machinery to prevent illicit repair events.
Though it is clear that the 32 telomeres of a haploid S. cerevisiae cell cluster in 2-8 foci at the nuclear periphery, a number of questions remained unanswered. It is unclear “who is where” among telomeric foci; that is whether telomeres always reside in a particular cluster, if some telomeres move freely outside of the clusters. My goal was to shed light on these questions using multicolor fluorescence microscopy to observe to differentially labeled telomeres in living cells. I aimed firstly to investigate the likelihood of a telomere residing within or outside of telomeric clusters, and secondly to see whether the composition of clusters is stable or stochastic. In other words, is telomere position in any given cluster reproducible amongst a population of cells? Alternatively, are telomere-telomere interactions random?
Furthermore, a major question that remained unclear is where the components of telomerase themselves are located within the nucleus, and the relationship between telomere position and telomere length regulation. At the beginning of my thesis it was known that there were at least two different pathways that recruit telomeres to the NE, one relying on the telomere bound
Sir complex and the peripheral protein Esc1, and the other involving the yKu heterodimer and a yet unknown anchoring partner yKu has been shown to interact with TLC1 (Stellwagen et al., 2003) and my goal was to investigate whether the interaction is involved in the positioning of telomeres at the NE.
Also in this context, I explored the relationship between telomere location and major aspects of telomere biology, namely capping and length regulation.
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