Heterogeneous Seeding of a Prion Structure by a Generic Amyloid Form of the Fungal Prion-forming Domain HET-s(218–289)

William Wan,Wen Bian,Michele Mcdonald,Aleksandra Kijac,David E Wemmer,Gerald Stubbs

doi:10.1074/jbc.m113.505511

Abstract

The fungal prion-forming domain HET-s(218-289) forms infectious amyloid fibrils at physiological pH that were shown by solid-state NMR to be assemblies of a two-rung β-solenoid structure. Under acidic conditions, HET-s(218-289) has been shown to form amyloid fibrils that have very low infectivity in vivo, but structural information about these fibrils has been very limited. We show by x-ray fiber diffraction that the HET-s(218-289) fibrils formed under acidic conditions have a stacked β-sheet architecture commonly found in short amyloidogenic peptides and denatured protein aggregates. At physiological pH, stacked β-sheet fibrils nucleate the formation of the infectious β-solenoid prions in a process of heterogeneous seeding, but do so with kinetic profiles distinct from those of spontaneous or homogeneous (seeded with infectious β-solenoid fibrils) fibrillization. Several serial passages of stacked β-sheet-seeded solutions lead to fibrillization kinetics similar to homogeneously seeded solutions. Our results directly show that structural mutation can occur between substantially different amyloid architectures, lending credence to the suggestion that the processes of strain adaptation and crossing species barriers are facilitated by structural mutation.

Highlights

Prions are aberrantly folded infectious proteins whose biological activity is determined by their distinct folds
We show by x-ray fiber diffraction that the HET-s(218 – 289) fibrils formed under acidic conditions have a stacked ␤-sheet architecture commonly found in short amyloidogenic peptides and denatured protein aggregates
The pH 7.5 diffraction patterns are in good qualitative agreement with the two-rung ␤-solenoid solid state NMR (ssNMR) structure, and we have used ssNMR to verify that our pH 7.5 HET-s(218 –289) sample has the same chemical shift assignments as those published [13] (Fig. 2)

Summary

Background

Prions are aberrantly folded infectious proteins whose biological activity is determined by their distinct folds. Accumulated amyloids are associated with a number of diseases in addition to transmissible spongiform encephalopathies, including Alzheimer disease, Parkinson disease, and type II diabetes [5] These amyloid diseases do not appear to be communicable between individuals, there is increasing evidence that the associated aberrantly folded proteins share self-propagating infectious properties similar to those of the transmissible spongiform encephalopathy-associated prions [6, 7]. Functional amyloids in yeast and fungi, have been shown to be prions by their propagation of phenotype through inheritance, by contact with prion-infected individuals, and by inoculation of prion fibrils prepared in vitro [11] Among these functional prions are the glutamine/asparagine-rich prions of Saccharomyces cerevisiae, including Sup35p and Ure3p, and HET-s from the fungus Podospora anserina. HET-s(218 –289) can be fibrillized into prions or noninfectious amyloids by controlling the pH of the fibrillization buffer

The abbreviations used are

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION