The prion hypothesis provides an explanation for a collection of previously inexplicable phenomena, ranging from the appearance, progression and spread of mammalian neurodegenerative disease to the non-Mendelian inheritance of unique traits in fungi. According to this idea, prion-associated phenotypes arise when a protein, known as a prion, adopts an alternative physical state and persist when that form self-replicates. This self- replication is mediated by the assembly of alternatively folded prions into aggregates, which template the con- version of other forms of the protein to a like state. The ability of prions to harness their conformational flexibil- ity is a central event in establishing distinct phenotypes, but this process becomes a multistep endeavor within the context of a living cell. Protein quality control pathways, prion biogenesis, and cell biology modify prion folding in vivo to create transmissible changes in physiology. A molecular understanding of how these forces intersect is a gap in current knowledge, limiting our ability to correlate prion folding mechanisms in vitro and their physiological consequences in vivo. As transitions between prion-associated phenotypes necessarily in- volve changes in protein state, these forces must converge on events that regulate prion dynamics in vivo. The long-term goal of this research is to elucidate the cellular mechanisms that influence prion folding to pro- duce transmissible changes in physiology. The overall objective of this application is to determine the molecu- lar mechanisms through which the prion folding pathway, cellular quality control, and other aspects of cell biol- ogy combine to create prion-associated phenotypes. Our approach is to determine the pathways through which variations in Sup35 sequence, conformation, and expression levels alter prion propagation in vivo. The central hypothesis is that the physical characteristics and abundance of Sup35 temper the efficiency with which mo- lecular chaperones recognize and/or process the prion form, thereby allowing distinct phenotypes to arise and persist but also to occasionally interconvert. Guided by strong preliminary data using the experimentally trac- table Sup35/[PSI+] prion of S. cerevisiae, this hypothesis will be tested through three specific aims: 1) Deter- mine the molecular mechanism by which sequence variants of Sup35 alter [PSI+] propagation, 2) Determine the molecular mechanism by which excess Hsp104 leads to [PSI+] loss, and 3) Determine the molecular mechanism by which Sup35 aggregation creates the [PSI+] phenotype. These proposed studies are innovative because they use a unique combination of experimental and mathematical analyses to test a new and dynamic model for prion-associated phenotypes. The proposed research is significant because it addresses the cellular regulation of prion folding, a poorly understood but significant factor that allows the prion hypothesis to create protein-based epigenetic elements in yeast. This knowledge, gained in the experimentally tractable yeast sys- tem, has the potential to provide new hypotheses that can be tested in more complex systems where a prion mechanism has been implicated.

Public Health Relevance

The proposed research is relevant to public heath because the misfolding of prion proteins is associated with a wide array of familial, sporadic and transmissible neurodegenerative disease in man. Our current understanding of how protein misfolding correlates with disease characteristics is limited;thus, our proposed studies are relevant to NIH's mission because the knowledge gained here has the potential to provide a new framework for understanding disease dynamics in man.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM069802-07
Application #
8326047
Study Section
Special Emphasis Panel (ZRG1-CB-J (03))
Program Officer
Maas, Stefan
Project Start
2006-02-01
Project End
2012-08-10
Budget Start
2012-06-01
Budget End
2012-08-10
Support Year
7
Fiscal Year
2012
Total Cost
$73,145
Indirect Cost
$27,612
Name
Brown University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
001785542
City
Providence
State
RI
Country
United States
Zip Code
02912
Holmes, William M; Mannakee, Brian K; Gutenkunst, Ryan N et al. (2014) Loss of amino-terminal acetylation suppresses a prion phenotype by modulating global protein folding. Nat Commun 5:4383
Pezza, John A; Villali, Janice; Sindi, Suzanne S et al. (2014) Amyloid-associated activity contributes to the severity and toxicity of a prion phenotype. Nat Commun 5:4384
Holmes, William M; Klaips, Courtney L; Serio, Tricia R (2014) Defining the limits: Protein aggregation and toxicity in vivo. Crit Rev Biochem Mol Biol 49:294-303
DiSalvo, Susanne; Derdowski, Aaron; Pezza, John A et al. (2011) Dominant prion mutants induce curing through pathways that promote chaperone-mediated disaggregation. Nat Struct Mol Biol 18:486-92
DiSalvo, Susanne; Serio, Tricia R (2011) Insights into prion biology: integrating a protein misfolding pathway with its cellular environment. Prion 5:76-83
Tuite, Mick F; Serio, Tricia R (2010) The prion hypothesis: from biological anomaly to basic regulatory mechanism. Nat Rev Mol Cell Biol 11:823-33
Derdowski, Aaron; Sindi, Suzanne S; Klaips, Courtney L et al. (2010) A size threshold limits prion transmission and establishes phenotypic diversity. Science 330:680-3
Sindi, Suzanne S; Serio, Tricia R (2009) Prion dynamics and the quest for the genetic determinant in protein-only inheritance. Curr Opin Microbiol 12:623-30
Pezza, John A; Langseth, Sara X; Raupp Yamamoto, Rochele et al. (2009) The NatA acetyltransferase couples Sup35 prion complexes to the [PSI+] phenotype. Mol Biol Cell 20:1068-80
Satpute-Krishnan, Prasanna; Langseth, Sara X; Serio, Tricia R (2007) Hsp104-dependent remodeling of prion complexes mediates protein-only inheritance. PLoS Biol 5:e24

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