Human stem cell-derived astrocytes replicate human prions in a PRNP genotype-dependent manner.

Prions are infectious agents that cause neurodegenerative diseases such as Creutzfeldt-Jakob disease (CJD). The absence of a human cell culture model that replicates human prions has hampered prion disease research for decades. In this paper, we show that astrocytes derived from human induced pluripotent stem cells (iPSCs) support the replication of prions from brain samples of CJD patients. For experimental exposure of astrocytes to variant CJD (vCJD), the kinetics of prion replication occur in a prion protein codon 129 genotype-dependent manner, reflecting the genotype-dependent susceptibility to clinical vCJD found in patients. Furthermore, iPSC-derived astrocytes can replicate prions associated with the major sporadic CJD strains found in human patients. Lastly, we demonstrate the subpassage of prions from infected to naive astrocyte cultures, indicating the generation of prion infectivity in vitro. Our study addresses a long-standing gap in the repertoire of human prion disease research, providing a new in vitro system for accelerated mechanistic studies and drug discovery

Mapping the Beta-Sheet Structure of the Yeast Prion Sup35 through Creation of Targeted Mutant Forms

Proteins with an aggregated form rich in beta-sheet structure are known as amyloids, of which a subset are infectious. These infectious proteins are known as prions and cause diseases including bovine spongiform encephalopathy (“Mad Cow” disease). Several prions have been identified in the baker’s yeast, Saccharomyces cerevisiae. One of the most well-studied yeast prions is the protein Sup35. To understand the fine protein structure of Sup35 better, we used PCR-based mutagenesis to introduce a lysine residue (a charged amino acid) at five defined places in the protein sequence of Sup35. We describe our process for creating these mutant versions and the results of DNA sequencing of each mutant version. The next step will be to assess prion formation and stability of clones with the correct sequences. Understanding the behavior of yeast prions has proven helpful in understanding human amyloid diseases and further studies on these yeast prions, including Sup35, will expand our knowledge further

Targeted Mutagenesis of the Oligopeptide Repeat Domain of the Yeast Prion Sup35

The formation of prions in the baker’s yeast Saccharomyces cerevisiae is determined by amino acid composition rather than the primary sequence of amino acids. The infectious amyloid proteins known as prions undergo nucleation and propagation, two distinct activities critical for prion formation. The ability for prions to be transferred from cell to cell, or propagate, is of interest not only in yeast prions but also in prion diseases such as the mammalian spongiform encephalopathies. Prion formation has been widely studied in yeast prions, however, the fundamental mechanisms behind the specific process of propagation of prions from cell to cell are not yet understood. In the most well-studied yeast prion, the prion form [PSI+] of Sup35, a domain of 5 ½ degenerate oligopeptide repeats called the oligopeptide repeat domain (ORD) has been shown to be important for prion propagation and to have a distinct amino acid composition as compared to the nucleation domain region. A library mutagenesis experiment has identified amino acids that favor or disfavor prion propagation in yeast cells. To confirm the results of the random library mutagenesis experiment, we generated several clones in which a portion of the ORD (the fourth oligopeptide repeat) was replaced with defined sequences expected to propagate or fail to propagate

Cellular Prion Protein Mediates Toxic Signaling of Amyloid Beta

Prion diseases in humans and animals comprise a group of invariably fatal neurodegenerative diseases characterized by the formation of a pathogenic protein conformer designated PrPSc and infectious particles denoted prions. The cellular prion protein (PrPC) has a central role in the pathogenesis of prion disease. First, it is the precursor of PrPSc and infectious prions and second, its expression on neuronal cells is required to mediate toxic effects of prions. To specifically study the role of PrPC as a mediator of toxic signaling, we have developed novel cell culture models, including primary neurons prepared from PrP-deficient mice. Using these approaches we have been able to show that PrPC can interact with and mediate toxic signaling of various beta-sheet-rich conformers of different origins, including amyloid beta, suggesting a pathophysiological role of the prion protein beyond prion diseases. Copyright (C) 2011 S. Karger AG, Base

Water and the Biology of Prions and Plaques

This is an attempt to account for the insolubility and/or aggregation of prions and plaques in terms of a model of water consisting of an equilibrium between high 
density and low density microdomains. Hydrophobic molecules, including proteins, 
accumulate selectively into stable populations, enriched in high density water, at 
charged sites on biopolymers. In enriched high density water, proteins are probably 
partially unfolded and may precipitate out when released. All extracellular matrices 
contain such charged polymers. Prions, which have been shown to accumulate in soils 
and clays containing silicates and aluminates also probably accumulate in 
extracellular matrices. 
 
Release of proteins follows hydrolysis of the charged groups by highly reactive high 
density water. This is normally a slow process but is greatly accelerated by urea. 
Plaques may form with age and disease because of accumulation of urea and, perhaps, 
glucose in the blood. This favours precipitation of proteins emerging from matrices, 
rather than refolding and solution. Dialysis should, therefore, interfere with plaque 
formation and impede the development of some age-related diseases

Half a world apart? overlap in nonbreeding distributions of Atlantic and Indian ocean thin-billed prions

Distant populations of animals may share their non-breeding grounds or migrate to distinct areas, and this may have important consequences for population differentiation and dynamics. Small burrow-nesting seabirds provide a suitable case study, as they are often restricted to safe breeding sites on islands, resulting in a patchy breeding distribution. For example, Thin-billed prions Pachyptila belcheri have two major breeding colonies more than 8,000 km apart, on the Falkland Islands in the south-western Atlantic and in the Kerguelen Archipelago in the Indian Ocean. We used geolocators and stable isotopes to compare at-sea movements and trophic levels of these two populations during their non-breeding season, and applied ecological niche models to compare environmental conditions in the habitat. Over three winters, birds breeding in the Atlantic showed a high consistency in their migration routes. Most individuals migrated more than 3000 km eastwards, while very few remained over the Patagonian Shelf. In contrast, all Indian Ocean birds migrated westwards, resulting in an overlapping nonbreeding area in the eastern Atlantic sector of the Southern Ocean. Geolocators and isotopic signature of feathers indicated that prions from the Falklands moulted at slightly higher latitudes than those from Kerguelen Islands. All birds fed on low trophic level prey, most probably crustaceans. The phenology differed notably between the two populations. Falkland birds returned to the Patagonian Shelf after 2-3 months, while Kerguelen birds remained in the nonbreeding area for seven months, before returning to nesting grounds highly synchronously and at high speed. Habitat models identified sea surface temperature and chlorophyll a concentration as important environmental parameters. In summary, we show that even though the two very distant populations migrate to roughly the same area to moult, they have distinct wintering strategies: They had significantly different realized niches and timing which may contribute to spatial niche partitioning