Mechanistic Analysis of the Ubiquitin-Proteasome System

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Project Summary / Abstract The ubiquitin-proteasome system (UPS) regulates the activity, localization and stability of thousands of proteins in the cell. By catalyzing the covalent attachment or removal of ubiquitin to target proteins, the enzymes of the UPS regulate virtually every cellular process, including cell signaling, cell survival and cell division. The widespread influence of the UPS on biology also has important consequences for human health, as mutations in components of the UPS cause diseases such as cancer. Encouragingly, the vast number of enzymes and protein-protein interactions in the UPS are also providing new drug targets to treat disease. There is growing interest in developing drugs that either inhibit the UPS or harness its ability to eliminate harmful proteins. For these reasons, mechanistic studies of the UPS have the potential to reveal new insights into biological regulation and may also help us understand mechanisms of disease pathogenesis and reveal opportunities for therapy. The focus of this proposal is to discover new substrates of the UPS and to characterize the specificity and regulation of enzymes that add and remove ubiquitin to these proteins. Although many components of the UPS are well-studied, we still have little insight into the molecular targets for hundreds of UPS enzymes encoded in the genome. This study will help fill this critical gap in our knowledge by identifying new enzyme-substrate relationships and providing new insights into how the enzymes of the system are regulated. It will apply modern quantitative proteomic methods to identify proteins that are modified by ubiquitin, and how these modifications change during cell division. This study will systematically examine the ability of deubiquitylating enzymes (DUBs) to remove ubiquitin from substrates, identifying new functional roles for these enzymes. The study will also examine the substrate specificity and regulation of specific ubiquitin conjugating enzymes and ubiquitin ligases, the proteins that conjugate ubiquitin to other proteins. Together, the experiments outlined in this study will identify new enzyme-substrate relationships in the UPS and provide new insights into how the UPS controls cell division. By defining these important regulatory pathways, the findings will provide a framework for understanding how these pathways are altered in cancer and may provide new targets for the development of drugs to treat disease.