Mechanism of prion species barrier at short phylogenetic distances in a yeast model

Georgia Tech Research Corporation, Atlanta GA

Investigators

Abstract

Amyloids are highly ordered self-seeded fibrous protein aggregates possibly representing an ancient protein fold and serving as a molecular tool for propagation of prions, which are infectious proteins. Prion infection is thought to spread via nucleated polymerization, so that polymeric prion "seeds" immobilize a normal monomeric protein of the same amino acid sequence and convert it into a prion. Transmission of the prion state from a pre-existing amyloid polymer to the newly mobilized protein molecules is a strictly sequence-specific process. The so-called "species barrier" prevents transmission of the prion state even between closely related proteins. If the barrier is overcome, this results in a cross-species prion transmission. The overall goal of this project is to uncover the molecular mechanisms that control the prion species barrier at low levels of protein sequence divergence. Cross-species prion transmission involves immobilization of a divergent protein into heteropolymers (co-aggregation), and propagation of the newly generated amyloid via its fragmentation, producing new "seeds". The species barrier between highly divergent prion domains is due to their inability to co-aggregate. It is hypothesized that at low levels of sequence divergence, a heterologous prion domain can be incorporated into the heteropolymers, but is unable to acquire the conformational state which would allow for efficient fragmentation and production of new prion "seeds" in the heterologous host. The project is designed to test this hypothesis. Specific Research Objectives are as follows: 1) to study cross-seeding capabilities of the divergent prion domains in vitro; 2) to identify the sequence differences that are responsible for the species barrier and defective propagation of the heterologous prions; 3) to investigate the role of cellular environment in the propagation of heterologous prions and the species barrier. Prion proteins of the closely related yeast species will be studied. Level of variation among these proteins corresponds to the range of divergence observed in the species barrier and cross-species prion transmission phenomena in the mammalian systems. A combination of both in vitro and in vivo approaches will be employed, followed by a direct comparison of the results. Prions can be found in mammals, yeast, and other fungi. They control inheritance of non-Mendelian traits in yeast and other fungi, and provide a mechanism for the "structural inheritance," possibly playing an important role in evolution. Results of this research will contribute to development of the strategies for prevention of cross-species prion transmission among closely related animals, and may have far-reaching implications for understanding the mechanisms controlling the specificity of protein-protein interactions involved in structural inheritance. The research project relies heavily on the participation of graduate and undergraduate students, and is being carried out by a diverse team of males and females. The project will be integrated courses for undergraduares and graduate students, who will contribute to it through their doctoral projects. The PI's laboratory participates in the multi-institutional Center for Fundamental and Applied Molecular Evolution (FAME), which promotes interdisciplinary research and education on various topics related to chemical and biological evolution. New genetically marked strains of the Saccharomyces species other than S. cerevisiae, constructed in the course of this project, will help to advance further studies of these organisms.