Prions and regulation of prion variants in Saccharomyces cerevisiae

Prions are infectious proteins. In their prion conformation, they catalyze the transformation of non-prion isomers into prions. Another characteristic common to currently identified prions is that they form amyloid aggregates. They occur in several mammals and multiple species of yeast, acting in a variety of biological processes, and their numbers continue to grow. Prions can adopt different infectious conformations known as “strains” in mammals or “variants” in yeast, each with distinct, epigenetically inheritable phenotypes. Mechanisms by which prion variants are determined remain unclear. In this thesis, I describe the characterization of a potential novel prion in Saccharomyces cerevisiae, Riq1p. I show that Riq1p can form SDS/heat-resistant aggregates and is sufficient to maintain a reversibly curable, nonsense-suppression phenotype when it is used to replace Sup35p’s prion determining domain. These findings are significant because, unlike the majority of previously identified prion proteins, Riq1p is richer in glutamine (Q) than asparagine (N) and its prion-like behavior cannot be isolated to its Q/N rich domain. In this way, Riq1p provides an important exception to amino-acid composition trends proposed to govern prion behavior and suggests that such theories require revision. I also report the results of a targeted screen for genes affecting early stages of de novo conversion of the prion protein Sup35p into its [PSI+] conformation. From this screen, I identify several chaperone genes that affect the variant of the S. cerevisiae prion Rnq1p/[PIN+]. I show that the deletion of specific chaperone genes alters [PIN+] variant phenotypes, including [PSI+] induction efficiency, Rnq1p aggregate morphology/size, and variant dominance. Genetic analysis demonstrated that the deletion-induced phenotypic changes are stably inherited in a non-Mendelian manner even after restoration of the deleted gene, confirming that they are due to a bona fide change in the [PIN+] variant. Taken together, these results not only show that it is possible to alter an established prion variant in vivo, but also that molecular chaperones may play an important part in this mechanism for regulating prion variants.