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Authors: Wagner, Tina
Title: Telomere folding in S. cerevisiae depends on chromatin modifiers and recombination factors and is lost during replicative senescence
Online publication date: 29-Jul-2020
Language: english
Abstract: The fact that eukaryotic chromosomes are linear requires the implementation of a protection mechanism that prevents their ends from being recognized as double strand breaks. Telomeres, the repetitive DNA elements at chromosome ends, fulfill this protective function by counteracting inappropriate checkpoint activation and repair. The protective capacity of telomeres has been largely attributed to specific telomere‐associated protein complexes. In addition, telomere loop structures have been proposed to contribute to end protection by sequestering the chromosome end and thereby preventing telomere fusions and degradation. Human telomeres form a lariat structure within the telomeric repeat tract, referred to as a t‐loop. Saccharomyces cerevisiae telomeres were proposed to fold back into subtelomeric regions, however, a robust method to study these structures has been lacking. In this thesis, a chromosome conformation capture‐based approach, dubbed Telo‐3C, was optimized to quantify telomere‐subtelomere interactions at a single unmodified yeast telomere. This methodology allowed for the identification of key regulatory factors that establish telomere folding. Studying the regulation of telomere folding in a genetically versatile system such as budding yeast offers the opportunity to draw parallels to the human system and to reveal regulatory mechanisms of the human telomere folding behavior. Using Telo‐3C, I have determined the homologous recombination machinery, comprising the factors Rad51 and Rad52, as well as the checkpoint kinase Rad53 as key folding regulators of budding yeast telomeres. My results show that the histone modifiers Sir2, Sin3 and Set2 are major regulatory factors of telomere folding indicating that the distinct telomeric chromatin environment might be a critical requirement for its establishment. Interestingly, the structural maintenance of chromosomes (SMC) complexes cohesin, condensin and SMC5/6 are not required for efficient folding of S. cerevisiae telomeres. Therefore, the mechanism by which telomere folding is established might be different from canonical internal chromatin loops. The Telo‐3C analysis demonstrated that wild type length telomeres fold back efficiently, however, as telomeres shorten and cells enter replicative senescence, telomere folding is compromised. I show that the unfolding of chromosome ends during senescence is independent from telomere shortening per se, but seems to be linked to the senescence program. Indeed, telomeres also unfold upon treatment with the DNA damaging agent MMS (methyl methanesulfonate), which induces a similar stress response and global gene expression changes as replicative senescence in budding yeast. A quantitative mass spectrometry approach revealed that the telomere folding regulators Sir2, Sin3 and Set2 are less abundant during replicative senescence. Therefore, their downregulation might contribute to the opening of the telomere fold‐back structure during this process.The data presented in this thesis point towards the homologous recombination machinery and certain chromatin modifiers being major requirements for the folding of yeast telomeres. Additionally, this thesis contributes to our understanding of the topological reorganization that telomeres undergo during senescence, a potent tumor suppressor mechanism in human cells.
DDC: 500 Naturwissenschaften
500 Natural sciences and mathematics
570 Biowissenschaften
570 Life sciences
Institution: Johannes Gutenberg-Universität Mainz
Department: FB 10 Biologie
Place: Mainz
URN: urn:nbn:de:hebis:77-openscience-86cbe9b2-9b31-453e-ba60-fcc21fc9ac633
Version: Original work
Publication type: Dissertation
License: CC BY-NC-ND
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Extent: X, 169 Seiten
Appears in collections:JGU-Publikationen

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