Abstract

Many signaling pathways employ covalent modification of target proteins by ubiquitin to control the timing of particular events. In order to more easily identify protein complexes and signaling networks, we have developed a bioinformatic and experimental platform that facilitates the parallel analysis of interaction networks and ubiquitin modifications. We have employed this platform to eluciduate the interaction landscape of several classes of proteins in the ubiquitin system, including a large fraction of human deubiquitinating enzymes, E3 ubiquitin ligases, and the autophagy system. Recently, we have developed technology that allows the identification of sites of ubiquitination of proteins on a large scale. We employ this approach to examine how proteasome inhibition re-sculpts the ubiquitinated proteome, and the role of protein synthesis in this process. We also have demonstrated the utility of this approach in the identification of substrates for ubiquitin ligases. These tools and technologies provide a facile system for understanding the complexities of the ubiquitin system and its role in cell function.