Review
Prion diseases of yeast: Amyloid structure and biology

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Abstract

Prion “variants” or “strains” are prions with the identical protein sequence, but different characteristics of the prion infection: e.g. different incubation periods for scrapie strains or different phenotype intensities for yeast prion variants. We have shown that infectious amyloids of the yeast prions [PSI+], [URE3] and [PIN+] each have an in-register parallel β-sheet architecture. Moreover, we have pointed out that this amyloid architecture can explain how one protein can faithfully transmit any of several conformations to new protein monomers. This explains how proteins can be genes.

Introduction

‘Prion’ means ‘infectious protein’, a protein which can transmit a disease or trait horizontally without the need for an accompanying nucleic acid. Although the self-activating yeast vacuolar protease B also can be a prion [1], most prions are amyloids, the filamentous protein polymer rich in β-sheet with the β-strands perpendicular to the filament long axis. Here we restrict our attention to the amyloid-based prions.

Perhaps the greatest mystery about prions, those of yeast as well as those of mammals, is the fact that a single protein sequence can stably propagate any of several prion variants. It is not particularly amazing that one protein can have several conformations; but that either of these conformations can be faithfully transmitted from molecule to molecule is certainly surprising. How is this conformational information transmitted? This “variant mystery” was one of the reasons for early skepticism of the prion hypothesis, independent of the absence of clear proof. We have shown that infectious amyloids of the prion domains of Ure2p, Sup35p, and Rnq1p each have an in-register parallel β-sheet architecture [2], [3], [4], with folds in the β-sheet along the long axis of the filaments [5]. This architecture nearly completely specifies the structure of the filament except for the locations of the folds and the precise extent of the sheet structure. “It has not escaped our attention that” this in-register parallel architecture provides a mechanism whereby the location of the folds and extent of the sheet once established, can be transmitted faithfully to each new protein monomer as it joins the end of the chain, thus explaining the variant mystery. It is not clear that other β-sheet structures (antiparallel, β-helix or out of register parallel) could explain this central mystery.

We will review the evidence for the in-register parallel β-sheet structure of these prion amyloids, and contrast it with the two-turn β-helix structure of the [Het-s] prion. It is likely that the different structural patterns reflect the different biology of these systems. [Het-s] is evolved to be a prion with a specific structure, while the yeast prions [PSI+] and [URE3] are apparently diseases.

Section snippets

The basics

The word ‘prion’ means ‘infectious protein’, a protein able to transmit a trait or disease without a required nucleic acid. Most of the prions of yeast and fungi are amyloids. Amyloid is a filamentous protein aggregate with a cross-β sheet secondary structure, meaning that the β-strands of the β-sheet are perpendicular to the long axis of the filament. We restrict our attention to the prions [PSI+], [URE3], [PIN+] and [MCA] of Saccharomyces cerevisiae, and [Het-s] of Podospora anserina, which

Prion domains

The prion properties of Ure2p, Sup35p, HET-s and Rnq1p are each determined by a restricted region of the protein. Residues 1–65 of Ure2p, 1–124 of Sup35p, 218–289 of HET-s and 153–405 of Rnq1p can propagate the prion in vivo or, as amyloid made in vitro from recombinant protein, infect yeast cells with the corresponding prion [21], [9], [34], [35]. For example, a cell expressing only Ure2p1–65 is efficiently infected by cytoplasm expressing the full length protein and carrying the [URE3] prion,

Shuffled prion domains can still form prions: suggests in-register parallel architecture

Initially with the goal of showing that the sequence of the Ure2p prion domain is important for prion formation, we found that each of 5 random shuffles of Ure2p residues 1–89, when integrated in place of the normal Ure2 prion domain, could support prion formation and propagation in vivo and amyloid formation in vitro [37]. The same result was obtained with the Sup35p prion domain [38]. These results indicate that the amino acid composition of these domains, not their sequence, determines their

Structural studies of yeast prion amyloids by solid-state NMR

X-ray crystallography and solution NMR cannot be used for studying the structure of amyloids because they cannot be crystallized and are not soluble (and are too large). Solid-state NMR [50] and electron paramagnetic resonance studies [51] are most useful, and solid-state NMR is particularly suited because the amyloid-forming peptide need not be chemically modified. Solid state NMR can measure distances between nuclei (through the efficiency of magnetization exchange), chemical environment

Structural differences between prion variants

Structural differences between prion variants of transmissible mink encephalopathy (studied in mice) were first shown by different protease – resistant cores of isolated PrPSc, the putative infectious material [31]. Genetic studies have shown that mutations in the Sup35 prion domain affect different [PSI+] variants differently [29]. Equivalently, variants of [URE3] show different degrees of propagation across a species barrier [30]. Extensive scanning mutagenesis indicates that the extent of

Mechanism of conformation inheritance inferred from parallel in-register structure

The in-register parallel architecture, and the presence of longitudinal folds in the sheets, and the requirement that each of many alternate prion protein conformations are faithfully propagated, led us to suggest that the locations of folds, as well as the extent of the β-sheet structure (see above), distinguish the amyloids that underlie different variants, and to suggest a detailed mechanism of prion conformational templating [69], [70]. In the parallel in-register structure, the register is

Are yeast prions adaptive or disease agents?

The biology of prions is determined by their structure, and the structure, in turn, reflects the biology. A functional amyloid (like a functional enzyme) is likely to have a specific structure optimized by evolution to carry out that function. However, a pathologic amyloid (like a denatured protein) is expected to have any of a variety of detailed structures. The yeast prions are known to have a variety of structures, as reflected in the prion variants, and wide peaks in 2D solid-state NMR

What is the connection of amyloid structure and prion biology?

First, a protein selected in evolution to be a prion should have a single prion variant/structure, because it is selected to do a specific task. Of course, we are thinking of [Het-s], which by one view is helping the host by its role in heterokaryon incompatibility and in another view is primarily a manifestation of a ‘parasitic’ meiotic drive gene. In either case, the HET-s protein is selected to have a specific form, and, indeed, there has not been any report of multiple [Het-s] variants.

Acknowledgement

This work was supported by the Intramural Program of the National Institute of Diabetes Digestive and Kidney Diseases.

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