Investigating the Mechanism of Sup35 Prion Curing by Excess Hsp104

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DESCRIPTION (provided by applicant): In most cases, proteins adopt a single three-dimensional fold that specifies their functions in vivo. There are several protein quality control pathways that help to ensure that proteins adopt their proper fold and that improperly folded proteins are degraded. The breakdown of these protein quality control pathways, particularly as a result of aging, can lead to a number of devastating human diseases. The most severe of these disorders, such as Alzheimer's, Parkinson's and Huntington's diseases are caused by the self- perpetuating misfolding and aggregation of otherwise normal cellular proteins. The appearance and persistence of these protein aggregates can lead to the disease state by inhibiting the function of the aggregated protein, or by producing toxic aggregates. In either case, control of the aggregation pathway is a key determinant in the progression of pathology. While these disease-associated aggregates are stable in vitro, the same complexes appear to be dynamic in vivo, suggesting that existing cellular mechanisms could be exploited through therapeutic intervention to target these pathogenic complexes for clearance. Perhaps the best-studied example of misfolded protein dynamics involves the yeast prion protein Sup35. Sup35 forms self-perpetuating, heritable aggregates that are continually remodeled by the molecular chaperone Hsp104. A similar remodeling activity is considered essential for the development and spread of self-perpetuating protein aggregates in higher eukaryotes. When Hsp104 levels are elevated in yeast, Sup35 prion aggregates, but not those comprised of other prion proteins, are lost over time. While in vitro studies suggest this effect results from resolubilization of aggregated protein, the available in vivo studies do not support this hypothesis. To elucidate the molecular basis of this process, I will determine both the characteristics of protein aggregates that confer susceptibility to excess Hsp104 and the mechanism by which excess Hsp104 leads to a loss of these complexes. Together, these studies will elucidate the dynamic equilibrium between soluble and aggregated forms of a misfolded protein in vivo, uncover the role of protein disaggregation in this process, and highlight key events in this pathway that could be exploited for therapeutic intervention. Many devastating aging-related diseases are caused by the breakdown of protein quality control pathways that lead to the buildup of toxic protein aggregates. The proposed work aims to elucidate the mechanism of action of a chaperone protein that functions in removing aggregated proteins from cells, providing insights into a pathway that could potentially be the target of future therapies.