IDENTIFICATION OF SUBSTRATES FOR UBIQUITIN-PROTEIN LIGASES IN YEAST

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Ubiquitin is one of the cell's most broadly utilized protein modifications, and it is now widely appreciated that ubiquitination serves critical regulatory roles in virtually every aspect of cellular physiology. Protein ubiquitination occurs through an enzymatic cascade in which ubiquitin is passed from an ubiquitin activase to an ubiquitin conjugase, and then attached to lysine residue within a substrate through the action of an ubiquitin-protein ligase. In the cascade, the ubiquitin-protein ligase imparts the substrate specificity, either by directly binding the substrate or by recruiting ancillary factors that bind substrates. Through homology-based searches, hundreds of ubiquitin-protein ligases are predicted to exist in humans, underscoring the importance and depth of ubiquitination in cellular regulation. However, the function for most of these has not yet been characterized, and so we know very little about the cohort of substrates for any particular ubiquitin-protein ligase. To address this problem, our long-term goal is to elucidate, in an unbiased way, the scope of targets for any putative or known ubiquitin-protein ligase, especially those implicated in disease. We are initially developing ubiquitin proteomics technologies using yeast as a model system because of its tractability. However, many yeast ubiquitination pathways also have human counterparts, so we will gain valuable biological information as well as providing a foothold in developing a mammalian version of this technology.