|Ph.D Student||Braten Ori|
|Subject||Non-Canonical Modes of Protein Ubiquitination|
|Department||Department of Medicine||Supervisor||? 18? Aaron Ciechanover|
|Full Thesis text|
Polymers of ubiquitin are formed on various protein substrates in eukaryotic cells. In the 'canonical' mode of ubiquitination, substrates are degraded by the 26S proteasome following their conjugation by multiple Ub moieties that are linked to one another via an isopeptide bond between the C-terminal Gly residue of the distal moiety and the ε-NH2 group of Lys48 of the proximal one. However, several modes of non-canonical ubiquitination exist that mediate a wide variety of biological outcomes. Here, we study non-canonical ubiquitination by focusing on monoubiquitin-dependent proteasomal degradation (Part A), and the generation of free ubiquitin chains (Part B).
While protein monoubiquitination regulates various processes, including membrane receptor internalization and chromatin remodeling, it is generally deemed non-destructive. However, emerging reports indicate that several specific substrates can be degraded following their monoubiquitination. In Part A, we apply a systematic approach for the identification of monoubiquitination-dependent proteasomal substrates. We combine techniques for ubiquitin replacement, ubiquitin conjugates enrichment and data analysis algorithms, and suggest that monoubiquitination-dependent degradation is more widespread than assumed previously, and may play key roles in various cellular processes.
Although ubiquitin chains are typically bound to a specific protein substrate, several studies have demonstrated the existence of free ubiquitin chains. Several physiological functions have been attributed to these chains, among them playing a role in signal transduction and serving as storage of ubiquitin for utilization under stress. In Part B, we establish a system for the detection of free ubiquitin chains and monitoring their level under changing conditions. Using this system, we show that the ubiquitin ligase UFD4 (ubiquitin fusion degradation 4) is involved in free chain generation. We also show that generation of these chains is stimulated in response to a variety of stresses, particularly those caused by DNA damage. However, it appears that the stress-induced synthesis of free chains is catalyzed by a different ligase, HUL5 (HECT ubiquitin ligase 5).