The effects of amino acid composition on yeast prion formation and prion domain interactions

Abstract

Yeast prions provide a powerful model system for examining prion formation and propagation in vivo. Yeast prion formation is driven primarily by amino acid composition, not by primary amino acid sequence. However, although yeast prion domains are consistently glutamine/asparagine-rich, they otherwise vary significantly in their compositions. Therefore, elucidating the exact compositional requirements for yeast prion formation has proven challenging. We have developed an in vivo method that allows for estimation of the prion propensity of each amino acid within the context of a yeast prion domain.1 Using these values, we are able to predict the prion-propensity of various glutamine/asparagine-rich yeast domains. These results provide insight into the basis for yeast prion formation, and may aid in the discovery of additional novel prion domains. Additionally, we examined whether amino acid composition could drive interactions between heterologous glutamine/asparagine-rich proteins.2 Although inefficient interactions between yeast prion domains have previously been observed, we found that one prion protein, Ure2, is able to interact with compositionally similar domains with unprecedented efficiency. This observation, combined with the growing number of yeast prions, suggests that a broad network of interactions between heterologous glutamine/asparagine-rich proteins may affect yeast prion formation.

Figures and Tables

Figure 1 Mutagenesis method. Oligonucleotides were designed with a degenerate region, flanked by regions complementary to SUP35-27. These degenerate oligonucleotides were used to PCR amplify SUP35-27, generating a library of sequences in which eight codons were randomly mutated. Using plasmid shuffling, the library was used to replace wild-type Sup35 as the sole copy of Sup35 in the cell. Mutants were selected for prion formation by plating on medium lacking adenine. Isolates from the naïve library and the prion-forming subset were sequenced to determine which amino acids were over/under-represented among the prion-forming isolates.

Figure 2 Prion propensity maps. Ure2p (A) and Sup35 (B) were scanned using a window size of 41 amino acids, calculating for each window the average order propensity using FoldIndex and prion propensity as the sum of the experimentally determined prion propensities for each amino acid across the window. The prion domain (PFD) is shaded.

Table 1 Prion propensity, order propensity and prevalence for each amino acid