|Ph.D Thesis||Department of Biology|
|Supervisor:||Prof. Glickman Michael|
|Full Thesis text|
Many regulatory proteins are removed from the cell in a timely and specific manner by a large multi subunit enzyme called the 26S proteasome. For proteins to be recognized by the proteasome, they are usually first covalently attached to a polyUbiquitin chain via a cascade of three ubiquitinating enzymes: ubiquitin activating, conjugating, and specific recruitment by a ubiquitin ligase. This process is reversible; the ubiquitin tag can be removed by proteases known as DUBs (deubiquitinating enzymes). Ubiquitinated substrates are shuttled to the 26S proteasome for processing into short peptides. The 26S proteasome is a multisubunit protease composed of a cylindrical catalytic core (20S-CP) and a regulatory particle (19S-RP) that together perform the essential degradation of polyUbiquitin labeled substrates.
Mitochondria form an ever-changing branched snake-like network, driven by two opposing events: fusion of membrane tips and fission into smaller organelles. Rapid cycles of collisions and divisions maintain the dynamic mitochondrial network which is crucial for energy production, mitochondrial DNA maintenance, mitochondrial inheritance, and cell viability. These processes are constantly under tight regulation balancing and have important implications for apoptosis, respiration and mitochondrial inheritance. This dynamic nature of mitochondrial morphology regulates a variety of mitochondrial functions and thus need to be tightly regulated.
In this thesis, I present data that Fzo1, a transmembrane GTPase required for mitochondria fusion, is ubiquitnated at mitochondria by the ubiquitin ligase SCF-Mdm30 and degraded by the proteasome. The determination that a mitochondrial outer membrane protein is ubiquitinated while still embedded and degraded by the proteasome lead us to posit the existence of an analogous mitochondrial associated degradation (MAD) pathway. We have identified and characterized a novel UPS component, Vms1, which is recruited to mitochondria upon mitochondrial stress where it plays a role in the degradation of ubiquitinated proteins.
The 19S-RP is separated into a lid and base sub-complexes. The only enzymatic function described for the lid is deubiquitination attributed to Rpn11. A carboxy terminus mutation termed rpn11-m1 exhibits both proteasome structural defects and an impaired mitochondria cellular network. We found that this C' terminal region of Rpn11 is sufficient to restore the typical proteasome structure and thus repair mitochondria respiration and morphology. Moreover, Rpn11 plays a structural role in stabilizing interactions with its nearest neighbors within the lid and in the lid-base interface.
Based on these observations, it seems that, the far-reaching arms of the UPS took over multiple facets of mitochondrial biology and dynamics.