Progress 09/01/05 to 08/31/07
Outputs
OUTPUTS: The major output during the funding period has been research efforts to identify factors that influence prion propagation, prion transmission, and protein aggregation using yeast prions as a model system. Genetic screens were conducted to identify factors required for the faithful propagation and transmission of the yeast prion [PSI+]. Forty-two mutants with potential defects in prion propagation or transmission were identified. Characterization and cloning of several of the strongest mutants are well underway. It is anticipated the results of this study will be disseminated to the scientific community through publications in scholarly journals.
PARTICIPANTS: Research was conducted by the principle investigator (L.A. Strawn) in the laboratory of Dr. Heather True-Krob at Washington University School of Medicine as part of her postdoctoral training.
TARGET AUDIENCES: The target audience is the scientific community, which will gain information on factors influencing prion propagation and transmission.
PROJECT MODIFICATIONS: The change in approach was outlined in the progress report for the first year of the funding period.
Impacts
Protein aggregation is the basis for several disorders, particularly neurodegenerative disorders, in both humans and animals. In the case of prion diseases, the disease-causing agent is transmissible between humans and animals through the food supply. Normally, a barrier to transmission of prions between species exists. However, in some instances there is evidence that the species barrier can be overcome. Studies on protein aggregation and factors that influence prion transmission and propagation in yeast may provide insight into the heterotypic aggregation that may occur during interspecies prion transmission. Factors identified using the yeast model system may provide the basis for future studies using animal model systems.
Publications
- Strawn, L.A., and H.L. True. 2006. Deletion of RNQ1 Gene Reveals Novel Functional Relationship between Divergently Transcribed Bik1p/CLIP-170 and Sfi1p in Spindle Pole Body Separation. Curr. Genet. 50:347-366.
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Progress 09/01/05 to 08/31/06
Outputs
The goal of this project is to use yeast prions as a model system to screen for factors that influence prion propagation, prion transmission, and heterotypic protein aggregation. We proposed to use a cytosolic protein-protein interaction system with the Sup35p prion-forming domains of two yeast species (both induced into the [PSI+] prion conformation) as our screenable system. We set up two different protein-protein interaction systems, but neither system worked correctly due to technical problems with the systems. Since we were unable to set up a suitable screening method to permit studying the interaction of prion proteins from two species within the same cell, we adopted another approach to understand more about heterotypic protein aggregation. Prion proteins from one species are not always capable of inducing conversion of the same protein from another species into a prion state (a species barrier to transmission exists). Conversely, within one species two
different prions can sometimes influence conversion frequencies of each other. It is intriguing how a strong species barrier may be the result of minor changes in primary sequence, yet one prion protein can serve to induce a prion conformation in a completely unrelated protein. By studying the heterotypic aggregation of two different prions from one species we will gain information that can be applied to the heterotypic aggregation that may occur during interspecies prion transmission. The Saccharomyces cerevisiae [RNQ+] prion influences the formation of [PSI+] potentially by acting as a seed onto which Sup35p can aggregate to form [PSI+] (Derkatch et al. 2000 EMBO J 19:1942-52). However, this system is currently limited in utility because the function of Rnq1p is unknown. We set up genetic screens to identify factors that are related to the function of Rnq1p. While screening for genetic interactions with RNQ1, we serendipitously uncovered a novel interactor with the divergently
transcribed BIK1 gene. This was uncovered because our deletion of RNQ1 lead to a great decrease in the expression of Bik1p. The characterization of this interaction is currently in press for publication, and we are completing a second screen for mutants directly related to RNQ1. We hope to use information gained about Rnq1p function to devise novel strategies to examine heterotypic aggregation between Rnq1p in [RNQ+] and Sup35p in [PSI+].
Impacts
Protein aggregation is the basis for several disorders, particularly neurodegenerative disorders, in both humans and animals. In the case of prion diseases, the disease-causing agent is transmissible between animals and humans through the food supply. Normally, a barrier to transmission of prions between species exists. However, in some instances there is evidence that the species barrier can be overcome. Studies on heterotypic protein aggregation and factors that influence protein aggregation may provide insight into the heterotypic aggregation that may occur during interspecies prion transmission.
Publications
- Strawn, L.A., and H.L. True. 2006. Deletion of RNQ1 Gene Reveals Novel Functional Relationship between Divergently Transcribed Bik1p/CLIP-170 and Sfi1p in Spindle Pole Body Separation. Curr. Genet., In press.
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