Pharmacology
background shadow background pic Gardner

Richard Gardner, Ph.D.,
Associate Professor

Lab Website
Publications
Course Websites
  phcol402

Email: gardnerr@uw.edu
Box 357280
HSC E421

Office: 206.685.0506
Lab: 206.616.1976
Fax: 206.685.3822
 

 

 

 

 

 

 

Overview of our research

We're generally interested in how the cell regulates protein function through post-translational modifications. Such modifications are critical to the cell because, once a protein is made, the cell must be able to control the level of activity to balance it with need. Of the various known protein modifications, we're very interested in exploring how the small protein ubiquitin is used as a modifer of other proteins to alter their stability, enzymatic activity, interactions, localization, or other functions.

FigureUbiquitin was discovered as a covalent modifier of histone H2A nearly 30 years ago, and has since emerged as one of the cell's most broadly utilized protein modifications. From a systems perspective, ubiquitin can be thought of as a universal cellular rheostat, deployed in an array of different configurations that are used in distinct ways to regulate a myriad of activities. And though the full extent of ubiquitin's action in the cell is unknown, it's easy to imagine that every cellular process is controlled in some way by ubiquitination. The large number of genes encoding known or predicted ubiquitination-associated proteins in eukaryotic genomes - more than 900 in humans alone - hints at ubiquitin's vast regulatory potential. Functions for a considerable number of these ubiquitination-associated proteins have been identified in a variety of organisms. Most, however, remain functionally uncharacterized. Even for those with known functions, only a handful of substrates have been identified for each, but it's likely that they target many more substrates.

So, to expand our knowledge and approach a complete understanding of ubiquitin's scope within the cell, we're interested in discovering new ubiquitination pathways, especially those that function in the nucleus to regulate chromatin-associated processes or act in novel ways. To accomplish this, we're using classical genetic and biochemical techniques, as well as a variety of new high throughput methods, in yeast. Our ultimate goal is to discover new ubiquitination pathways in yeast, and then determine if analogous pathways similarly function in metazoans.

Specific ongoing research projects

Nuclear protein quality control

The age-dependent accumulation of structurally aberrant, aggregation-prone proteins has been implicated in the pathogenesis of over 30 heritable disorders including Alzheimer's, Parkinson's, and Huntington's. Although many of these disorders are attributed to aberrant protein accumulation and aggregation in the cytoplasm and extracellular space, a striking majority (>20) are linked to accumulation and aggregation specifically in the nucleus. Why the nucleus is particularly susceptible to the toxic effects of aggregation-prone proteins is not clear. We know the cell generally protects itself from accumulating aberrant proteins through numerous Protein Quality Control (PQC) mechanisms that either repair structurally aberrant proteins by the action of chaperones or destroy the aberrant proteins through cellular degradation systems. However, studies of PQC have primarily focused on the cytoplasm and ER and we have a solid understanding of how PQC chaperone and degradation systems in these compartments work. Studies of PQC in the nucleus have been few to this point, so it's poorly understood how the cell safeguards against the nuclear accumulation of aberrant proteins. To close this gap in our cellular PQC knowledge and learn more about this fundamental aspect of nuclear biology that impacts human disease, we've focused our attention on nuclear PQC to address these key unresolved questions: (1) What PQC systems act in the nucleus to protect it from toxic aberrant proteins? (2) What do nuclear PQC systems recognize as aberrant, and are they incapable of targeting specific aberrant proteins? (3) Are lesions in nuclear PQC systems required for disease pathogenesis? (4) What nuclear processes are disrupted by nuclear accumulation of aberrant proteins?

Protein deubiquitination in the nucleus

Ubiquitination is an important modification for regulating a variety of chromatin-associated processes including gene activation, DNA replication, and DNA repair. Although groups in the chromatin field has identified some chromatin targets of ubiquitination (such as histone H2B, RNA and DNA polymerase, origin of replication complex proteins, etc), we don't yet understand how this modification is dynamically regulated to control its myriad chromatin processes. In particular, we don't understand how deubiquitination is employed to tightly regulate chromatin function or other nuclear processes. We've recently identified and characterized Ubp10 as yeast deubiquitinating enzyme that removes ubiquitin from histone H2B to regulate silent chromatin. We now also have evidence that Ubp10 deubiquitinates H2B in active chromatin for an as yet unknown purpose, may regulate transcription factor stability perhaps as a way to control transcription, and may also function in ribosome biogenesis in an as yet unidentified step. Characterizing these new Ubp10 functions will open up our view of ubiquitination and deubiquitination in the nucleus, and bring us closer to understanding the dynamic nature of ubiquitin both in regulating chromatin function and in controlling other critical nuclear processes.

Develop novel proteomics methods to identify substrates for ubiquitin-protein ligases and deubiquitinating enzymes

The defining challenge of the ubiquitin field is the identification of substrates for ubiquitin-protein ligases and deubiquitinating enzymes. Until now, substrates have been identified on a case-by-case basis, which is often tedious and prone to failure. To improve our ability to understand ubiquitin-protein ligase and deubiquitinating enzyme function, we're developing ubiquitin proteomics technologies that will help us identify the entire cohort of substrates for any ubiquitin-protein ligase or deubiquitinating enzymes. Because there are hundreds of putative ubiquitin-protein ligases and dozens of deubiquitinating enzymes encoded in the human genome, these technologies will dramatically enhance our knowledge not only of basic cellular biology, but will provide inroads into diseases that result, in part, from dysregulation of ubiquitination, such as cancer and neurodegenerative diseases.



Recent Publications 

Cellular maintenance of nuclear protein homeostasis. - ABSTRACT

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