Research Article: Synthetic prions with novel strain-specified properties

Date Published: December 31, 2015

Publisher: Public Library of Science

Author(s): Fabio Moda, Thanh-Nhat T. Le, Suzana Aulić, Edoardo Bistaffa, Ilaria Campagnani, Tommaso Virgilio, Antonio Indaco, Luisa Palamara, Olivier Andréoletti, Fabrizio Tagliavini, Giuseppe Legname, David Westaway.

http://doi.org/10.1371/journal.ppat.1005354

Abstract

Prions are infectious proteins that possess multiple self-propagating structures. The information for strains and structural specific barriers appears to be contained exclusively in the folding of the pathological isoform, PrPSc. Many recent studies determined that de novo prion strains could be generated in vitro from the structural conversion of recombinant (rec) prion protein (PrP) into amyloidal structures. Our aim was to elucidate the conformational diversity of pathological recPrP amyloids and their biological activities, as well as to gain novel insights in characterizing molecular events involved in mammalian prion conversion and propagation. To this end we generated infectious materials that possess different conformational structures. Our methodology for the prion conversion of recPrP required only purified rec full-length mouse (Mo) PrP and common chemicals. Neither infected brain extracts nor amplified PrPSc were used. Following two different in vitro protocols recMoPrP converted to amyloid fibrils without any seeding factor. Mouse hypothalamic GT1 and neuroblastoma N2a cell lines were infected with these amyloid preparations as fast screening methodology to characterize the infectious materials. Remarkably, a large number of amyloid preparations were able to induce the conformational change of endogenous PrPC to harbor several distinctive proteinase-resistant PrP forms. One such preparation was characterized in vivo habouring a synthetic prion with novel strain specified neuropathological and biochemical properties.

Partial Text

Prion diseases or transmissible spongiform encephalopathies are fatal neurodegenerative disorders of humans and animals. During the course of these maladies the cellular prion protein (PrPC) converts into an abnormally folded isoform, PrPSc, which accumulates in the central nervous system (CNS), ultimately leading to the host death [1, 2]. The key molecular event of these disorders is therefore the conformational change of PrPC from its physiological form into the pathological structural isoform noted as prion or PrPSc. The physiological PrPC is a glycosylphosphatidylinositol (GPI)-anchored protein present in all cell types. Unlike PrPSc, PrPC has a high content of α-helix in its secondary structure. Differences in structural conformation lead to different biological characteristics: while PrPC is soluble in detergents and sensitive to proteinase K (PK) digestion, PrPSc is partially PK-resistant and insoluble in non-ionic detergents [3]. Nevertheless, a sizeable fraction of PrPSc is sensitive to PK digestion [4] and is dispersed with detergent and/or sonication [5]. Moreover, Colby et al. reported that protease-sensitive synthetic prions could be generated in vitro during polymerization of recombinant PrP (recPrP) into amyloid fibers [6]. Recently, PK-sensitive and PK-resistant PrPSc were shown to share a common structure and phenotype despite the differences in resistance to PK-digestion, sediment and distribution of multimers [7, 8].

The production of synthetic prions was introduced in 2004, via a simple in vitro induction of misfolding and aggregation of bacterially expressed recPrP [15]. PrP amyloids possessing different conformation stability were generated by altering the conditions for their formation, including urea concentration, pH and temperature. After passaging in mice, a large ensemble of synthetic prions showed a direct relationship between stability and incubation time of novel prion strains [6]. The recent, impressive progress in this technique has spurred the renewed investigation of prion structural biology [41–45].

 

Source:

http://doi.org/10.1371/journal.ppat.1005354

 

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