Research

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For most short-lived eukaryotic proteins, attachment to ubiquitin is an obligatory step in the cascade of events that culminates in their degradation. Protein turnover via the highly conserved ubiquitin-proteasome system is central to a great variety of regulatory mechanisms, including many of medical relevance. We wish to understand, at a mechanistic and molecular level, how specific proteins are rapidly degraded within cells even while most proteins are spared. This entails detailed functional analysis of the various elements of the ubiquitin-proteasome system, from the degradation signals targeting proteins for destruction, to the components of the ubiquitin conjugation machinery, to the proteasome itself.


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Doa10 is a yeast ubiquitin ligase (E3) discovered in our lab that is embedded in the endoplasmic reticulum (ER) / nuclear envelope. It has the unusual ability to localize in both the nucleus and ER and gain access to distinct sets of substrates in this way. For example, Doa10 is implicated in ER-associated degradation (ERAD), a process that identifies misfolded proteins in the ER and targets them for efficient disposal by the proteasome. We are working towards a thorough functional and structural understanding of this integral membrane ubiquitin ligase, including its interactions with other proteins, its mechanism of substrate recognition, and its specific roles in ERAD and the degradation of nuclear regulatory proteins.


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One of the least understood aspects of the machinery comprising the ubiquitin-proteasome system is how the proteasome itself is assembled. The eukaryotic 20S proteasome has 14 different types of subunit arranged into four heptameric rings. The 19S regulator that binds to, and regulates the activity of, the 20S proteasome is composed of at least 18 distinct proteins. In addition, a number of accessory proteins are thought to be intimately involved in proteasome assembly, and we are working towards an understanding of their function as putative assembly factors. We are interested in understanding the assembly process of the proteasome because this has important implications for how cells regulate their proteolytic capacity to suit their needs. Moreover, a regulated process such as proteasome assembly can provide an interesting target for pharmacological intervention.


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We are also interested in analyzing the function and dynamics of protein modification by other "ubiquitin-like proteins." SUMO is a ubiquitin-like protein that can be conjugated to substrates via a mechanism entirely analogous to ubiquitination. SUMO has been implicated in the regulation of various cellular processes, including the cell cycle and nucleocytoplasmic trafficking. Little is known about the in vivo targets of sumoylation or the regulation of this process. We are interested in determining the role that sumoylation plays in the normal function of the cell. Our current goals include identifying novel SUMO-protein ligases, characterizing the functions of the SUMO-specific proteases Ulp1 and Ulp2, and elucidating the role of SUMO in the response to DNA damage.