The ubiquitin-like proteins (Ubls) are small polypeptides that function as post-translational modifiers. Like ubiquitin, most Ubls are covalently attached to a lysine residue on target proteins. The small ubiquitin-related modifiers (SUMO) play important roles in a number of critical biological processes, such as proliferation and regulation of the cell cycle, yet their specific cellular functions have remained poorly understood. Like ubiquitin, SUMO proteins can also form oligomeric “chains”, but the functions of these structures were even less well understood. To this end, I created the first spectral library for the identification of Ub/Ubl proteins and Ub/Ubl chain linkages in mass spectrometry experiments. This tool has dramatically improved our ability to use MS to analyze the contents of biological samples for Ub and Ubls, and to identify specific types of Ub and Ubl chains in model organisms. I also used MS to conduct the first comprehensive SUMO system protein-protein interactome in any organism. In total, 452 high confidence protein-protein interactions were detected for S. cerevisiae SUMO system proteins, encompassing a total of 321 interacting partners. Yeast SUMO system components were found to interact with proteins involved in a number of different biological processes, and my mapping effort increased the number of known SUMO system interacting partners >50-fold. This study revealed that a number of transcriptional co-repressors and chromatin remodelling proteins interact physically with specific SUMO system components, with a clear division of labour between SUMO system enzymes. Finally, I conducted the first global analysis of SUMO chain function, using a combination of genetic, high-content microscopy, and high-density transcriptomics screens. Consistent with my interactomics work, this study demonstrated that inhibition of SUMO chain synthesis leads to severe chromatin condensation defects, which in-turn leads to chromosome missegregation, unscheduled transcription of stress-and nutrient-regulated genes, and aberrant intragenic transcription. Together, my work thus revealed a major role for the SUMO system in the maintenance of higher order chromatin structure and transcriptional repression.
| Identifer | oai:union.ndltd.org:TORONTO/oai:tspace.library.utoronto.ca:1807/36004 |
| Date | 13 August 2013 |
| Creators | Srikumar, Tharan |
| Contributors | Raught, Brian |
| Source Sets | University of Toronto |
| Language | en_ca |
| Detected Language | English |
| Type | Thesis |
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