Determination of ubiquitin fitness landscapes under different chemical stresses in a classroom setting

David Mavor(University of California, San Francisco), Kyle A. Barlow(University of California, San Francisco), Samuel Thompson(University of California, San Francisco), Benjamin A. Barad(University of California, San Francisco), Alain R. Bonny(University of California, San Francisco), Clinton L. Cario(University of California, San Francisco), Garrett Gaskins(University of California, San Francisco), Zairan Liu(University of California, San Francisco), Laura Deming(Howard Hughes Medical Institute), Seth D. Axen(University of California, San Francisco), Elena L. Cáceres(University of California, San Francisco), Weilin Chen(University of California, San Francisco), Adolfo Cuesta(University of California, San Francisco), Rachel E. Gate(University of California, San Francisco), Evan M. Green(University of California, San Francisco), Kaitlin R. Hulce(University of California, San Francisco), Weiyue Ji(University of California, San Francisco), Lillian R Kenner(University of California, San Francisco), Bruk Mensa(University of California, San Francisco), Leanna S. Morinishi(University of California, San Francisco), Steven M. Moss(University of California, San Francisco), Marco Mravic(University of California, San Francisco), Ryan K. Muir(University of California, San Francisco), Stefan Niekamp(University of California, San Francisco), Chimno I. Nnadi(University of California, San Francisco), Eugene Palovcak(University of California, San Francisco), Erin M. Poss(University of California, San Francisco), Tyler D. Ross(University of California, San Francisco), Eugenia C. Salcedo(University of California, San Francisco), Stephanie K. See(University of California, San Francisco), Meena Subramaniam(University of California, San Francisco), Allison W. Wong(University of California, San Francisco), Jennifer Li(University of California, San Francisco), Kurt S. Thorn(University of California, San Francisco), Shane Ó Conchúir(QB3), Benjamin P. Roscoe(University of Massachusetts Chan Medical School), Eric D. Chow(University of California, San Francisco), Joseph L. DeRisi(Howard Hughes Medical Institute), Tanja Kortemme(QB3), Daniel N. Bolon(University of Massachusetts Chan Medical School), James S. Fraser(QB3)
eLife
April 25, 2016
10.7554/elife.15802
Cited by 97Open Access
Full Text

Abstract

Ubiquitin is essential for eukaryotic life and varies in only 3 amino acid positions between yeast and humans. However, recent deep sequencing studies indicate that ubiquitin is highly tolerant to single mutations. We hypothesized that this tolerance would be reduced by chemically induced physiologic perturbations. To test this hypothesis, a class of first year UCSF graduate students employed deep mutational scanning to determine the fitness landscape of all possible single residue mutations in the presence of five different small molecule perturbations. These perturbations uncover 'shared sensitized positions' localized to areas around the hydrophobic patch and the C-terminus. In addition, we identified perturbation specific effects such as a sensitization of His68 in HU and a tolerance to mutation at Lys63 in DTT. Our data show how chemical stresses can reduce buffering effects in the ubiquitin proteasome system. Finally, this study demonstrates the potential of lab-based interdisciplinary graduate curriculum.

Related Papers

Transformation of yeast by lithium acetate/single-stranded carrier DNA/polyethylene glycol method
R. Daniel Gietz, Robin A. Woods|Methods in enzymology on CD-ROM/Methods in enzymology|2002|3k
Inhibitors of the proteasome block the degradation of most cell proteins and the generation of peptides presented on MHC class I molecules
Kenneth L. Rock, Colette F. Gramm, Lisa Rothstein et al.|Cell|1994|2.5k
Recognition of the polyubiquitin proteolytic signal
Julia S. Thrower, Laura Hoffman, Martin Rechsteiner et al.|The EMBO Journal|2000|1.7k
Quantitative Proteomics Reveals the Function of Unconventional Ubiquitin Chains in Proteasomal Degradation
Ping Xu, Duc M. Duong, Nicholas T. Seyfried et al.|Cell|2009|1.1k
Recognition Dynamics Up to Microseconds Revealed from an RDC-Derived Ubiquitin Ensemble in Solution
Oliver F. Lange, Nils‐Alexander Lakomek, Christophe Farès et al.|Science|2008|1.1k