Distinct Amino Acid Compositional Requirements for Formation and Maintenance of the [PSI⁺] Prion in Yeast

Abstract

bMultiple yeast prions have been identified that result from the structural conversion of proteins into a self-propagating amyloid form. Amyloid-based prion activity in yeast requires a series of discrete steps. First, the prion protein must form an amyloid nucleus that can recruit and structurally convert additional soluble proteins. Subsequently, maintenance of the prion during cell division requires fragmentation of these aggregates to create new heritable propagons. For the Saccharomyces cerevisiae prion protein Sup35, these different activities are encoded by different regions of the Sup35 prion domain. An N-terminal glutamine/ asparagine-rich nucleation domain is required for nucleation and fiber growth, while an adjacent oligopeptide repeat domain is largely dispensable for prion nucleation and fiber growth but is required for chaperone-dependent prion maintenance. Although prion activity of glutamine/asparagine-rich proteins is predominantly determined by amino acid composition, the nucleation and oligopeptide repeat domains of Sup35 have distinct compositional requirements. Here, we quantitatively define these compositional requirements in vivo. We show that aromatic residues strongly promote both prion formation and chaperone-dependent prion maintenance. In contrast, nonaromatic hydrophobic residues strongly promote prion formation but inhibit prion propagation. These results provide insight into why some aggregation-prone proteins are unable to propagate as prions. Misfolding of a wide range of proteins leads to formation of amyloid fibrils, which are ordered, ␤-sheet-rich protein aggregates. Many human diseases are associated with the formation of amyloid fibrils, including Alzheimer's disease, type II diabetes, and the transmissible spongiform encephalopathies (TSEs) (1). However, only a small subset of amyloids are infectious (called prions), including the causative agents of TSEs in mammals (2-4) and, and others in Saccharomyces cerevisiae (5-9).Most of the known yeast prion proteins contain glutamine/ asparagine (Q/N)-rich domains that drive amyloid formation. Q/N-rich domains are found in 1 to 4% of the proteins in most eukaryotic proteomes (10), but very few of these proteins have been shown to undergo amyloid structural conversion. Bioinformatics screens for prions in yeast have had some notable successes (reviewed in reference 11); however, despite advances in predicting which Q/N-rich domains may turn out to be bona fide prions (12, 13), predictions remain imperfect.A well-studied model prion from yeast (S. cerevisiae) is [PSI ϩ ], the prion form of the translational terminator protein Sup35 (5). Like other yeast prion proteins, Sup35 is modular, as it contains a distinct prion-forming domain (PFD), middle domain (M), and C-terminal domain (C) (Fig. 1A) (14-17). The PFD (amino acids 1 to 114) drives the conversion of Sup35 into its amyloid form (15), the charged M domain has no known function other than its ability to stabilize [PSI ϩ ] fibers, and the C domain is an essential component resp...

Amino acid composition predicts prion activity

“…Another amino acid where a big difference is observed is tryptophan: it has the second lowest negative log-odds value, but is assigned a weight that is close to zero in the genome-wide pRANK evaluation. There is experimental support for the non-deleterious effect of tryptophan on prion formation: MacLea et al showed that replacing tyrosines in Sup35 with tryptophan increases prion formation [45], and similar results were reported by Ohhashi et al [46]. Although tryptophan is known to strongly promote aggregation, it has been suggested that it too strongly promotes prion formation, and therefore is not well tolerated in disordered segments [22]; this observation is in agreement with the analysis of Buck et al [47].…”

Distinct Prion Domain Sequences Ensure Efficient Amyloid Propagation by Promoting Chaperone Binding or Processing In Vivo

“…In this model, the requirement for a minimum number of repeats would be interpreted as a threshold of tyrosine residues necessary to promote efficient fragmentation. Consistent with this idea, replacement of the tyrosines in repeats 3, 4 and 5 with non-aromatic residues leads to [ PSI + ] loss in vivo , although progressive effects and the mechanism by which this loss occurs were not assessed [95]. …”

Natural and Pathogenic Protein Sequence Variation Affecting Prion-Like Domains Within and Across Human Proteomes

“…Using centralized PTM databases, we mapped PTMs to human PrLDs. While the contribution of each of the canonical amino acids to aggregation of PrLDs has been fairly wellcharacterized (7,80), consistent effects of each type of PTM on aggregation of PrLDs have not been defined. Therefore, we mapped PTMs to PrLDs using a relaxed aggregation propensity threshold (PAPA cutoff=0.0, rather than the standard 0.05 threshold), which accounts for the possibility that PTMs could increase aggregation propensity or regulate the solubility of proteins whose aggregation propensity is near the standard 0.05 aggregation threshold.…”