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Zhu, Mang

University of British Columbia

Protein aggregation is often a hallmark of protein homeostasis disruption in the cell. Accumulation of cytotoxic protein species has been associated to many diseases. However, protein aggregation has also been shown to be beneficial and provide biological functions in other cases. With a fine line between a functional and harmful outcome, protein aggregation is highly regulated and under constant monitoring of the protein homeostasis network. In this thesis, we combined cell fractionation with mass spectrometry to identify proteins aggregates and to delineate features associated with them. To identify protein aggregates in unstressed cells, we established a quantitative mass spectrometry approach that combines SILAC labeling and centrifugation. We found that these proteins are longer and lower abundant proteins with fewer hydrophobic residues and more low complexity and disordered regions. We also found features that suggest these proteins have more potential interaction partners and are involved in function macro assemblies. We next examined which yeast proteins aggregate under stress condition by using heat shock. We found that proteins that aggregate under heat-shock were enriched with known stress granule (SG) proteins. There proteins are also abundant and enriched in intrinsically disordered regions (IDRs). We found that several IDRs were sufficient for localization to SG. Lastly, we showed that the IDR of the Ubp3 was critical for yeast SG formation. This work shows the importance of IDRs in mediating protein condensation upon stress. Finally, we examined which newly translated proteins aggregate after heat-shock. We found these proteins were abundant, shorter, highly ordered, and more hydrophobic. Notably these proteins contained more β-sheets. As well, there was an enrichment for proteins with chaperone-binding motifs. Interestingly, these proteins were also more often components of stable protein complexes. We found a specific group of thermal labile proteins that takes longer to reach thermostable native state. Collectively, this thesis uncovered protein features associated with aggregating proteins using a common experimental framework and sheds light on how protein aggregates are formed and regulated. Our work provides a platform for future studies to better comprehend how protein homeostasis is regulated and to potentially prevent unwanted protein aggregation for therapeutic applications.

Text

2021-06-09

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/78575

Genome Science and Technology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/