Defining the Function of Proteasomal Deubiquitinases

The 26S proteasome is a massive, intricately regulated ATP-dependent protease responsible for the degradation of most cellular proteins. The goal of this research program is to determine how degradation by the 26S proteasome is regulated by deubiquitinases. Since misregulation of protein degradation is a hallmark of many human cancers and neurological disorders, the results of these studies can be used to design new drugs that target key regulators of the 26S proteasome. A major focus of this research program will be on the deubiquitinase UCHL5/UCH37, which is recruited to the proteasome by the ubiquitin-binding receptor ADRM1/RPN13. Deficiencies in UCH37 leads to embryonic lethality in mice and overexpression of UCH37 along with RPN13 is associated with poor prognoses in several cancers. A combination of x-ray crystallography, NMR, in vitro biochemistry, protein engineering, and cell-based approaches will be used to understand how UCH37 selects its ubiquitin chain targets, why the cleavage of those targets is necessary for promoting proteasomal degradation, and which proteins in a cellular proteome require UCH37 activity for efficient degradation. We will also develop a set of single-chain antibodies (referred to as nanobodies) capable of interfering with distinct activities of UCH37. These new tools will be instrumental in defining the function of UCH37 in different biochemical pathways such as the cell cycle and the cellular response to oxidative stress and DNA damage. Considering the tumor suppressor BAP1 shares a similar sequence with the catalytic domain of UCH37, our tools and approaches can also be used to better understand its role outside of histone deubiquitination. The molecular insights that will result from the proposed studies will provide a foundation for the development of new therapeutic agents that target cancers requiring excessive proteasome activity for survival.