scholarly journals Mouse-Adapted Ovine Scrapie Prion Strains Are Characterized by Different Conformers of PrPSc

2007 ◽  
Vol 81 (22) ◽  
pp. 12119-12127 ◽  
Author(s):  
Alana M. Thackray ◽  
Lee Hopkins ◽  
Michael A. Klein ◽  
Raymond Bujdoso
Keyword(s):  
Biological Diversity ◽  
Guanidine Hydrochloride ◽  
Serial Passage ◽  
Biological Properties ◽  
Proteinase K ◽  
Specific Information ◽  
Strain Typing ◽  
Prion Strain ◽  
Prion Strains ◽  
Scrapie Strains

ABSTRACT The agent responsible for prion disease may exist in different forms, commonly referred to as strains, with each carrying the specific information that determines its own distinct biological properties, such as incubation period and lesion profile. Biological strain typing of ovine scrapie isolates by serial passage in conventional mice has shown some diversity in ovine prion strains. However, this biological diversity remains poorly supported by biochemical prion strain typing. The protein-only hypothesis predicts that variation between different prion strains in the same host is manifest in different conformations adopted by PrPSc. Here we have investigated the molecular properties of PrPSc associated with two principal Prnp a mouse-adapted ovine scrapie strains, namely, RML and ME7, in order to establish biochemical prion strain typing strategies that may subsequently be used to discriminate field cases of mouse-passaged ovine scrapie isolates. We used a conformation-dependent immunoassay and a conformational stability assay, together with Western blot analysis, to demonstrate that RML and ME7 PrPSc proteins show distinct biochemical and physicochemical properties. Although RML and ME7 PrPSc proteins showed similar resistance to proteolytic digestion, they differed in their glycoform profiles and levels of proteinase K (PK)-sensitive and PK-resistant isoforms. In addition, the PK-resistant core (PrP27-30) of ME7 was conformationally more stable following exposure to guanidine hydrochloride or Sarkosyl than was RML PrP27-30. Our data show that mouse-adapted ovine scrapie strains can be discriminated by their distinct conformers of PrPSc, which provides a basis to investigate their diversity at the molecular level.

scholarly journals The Region Approximately between Amino Acids 81 and 137 of Proteinase K-Resistant PrPSc Is Critical for the Infectivity of the Chandler Prion Strain

2009 ◽  
Vol 83 (8) ◽  
pp. 3852-3860 ◽  
Author(s):  
Ryo Shindoh ◽  
Chan-Lan Kim ◽  
Chang-Hyun Song ◽  
Rie Hasebe ◽  
Motohiro Horiuchi
Keyword(s):  
Amino Acids ◽  
Guanidine Hydrochloride ◽  
Conformational Stability ◽  
Terminal Region ◽  
Proteinase K ◽  
Prion Strain ◽  
Prion Infectivity ◽  
Prion Strains ◽  
Incubation Periods ◽  
The Relationship

ABSTRACT Although the major component of the prion is believed to be the oligomer of PrPSc, little information is available concerning regions on the PrPSc molecule that affect prion infectivity. During the analysis of PrPSc molecules from various prion strains, we found that PrPSc of the Chandler strain showed a unique property in the conformational-stability assay, and this property appeared to be useful for studying the relationship between regions of the PrPSc molecule and prion infectivity. Thus, we analyzed PrPSc of the Chandler strain in detail and analyzed the infectivities of the N-terminally denatured and truncated forms of proteinase K-resistant PrP. The N-terminal region of PrPSc of the Chandler strain showed region-dependent resistance to guanidine hydrochloride (GdnHCl) treatment. The region approximately between amino acids (aa) 81 and 137 began to be denatured by treatment with 1.5 M GdnHCl. Within this stretch, the region comprising approximately aa 81 to 90 was denatured almost completely by 2 M GdnHCl. Furthermore, the region approximately between aa 90 and 137 was denatured completely by 3 M GdnHCl. However, the C-terminal region thereafter was extremely resistant to the GdnHCl treatment. This property was not observed in PrPSc molecules of other prion strains. Denaturation of the region between aa 81 and 137 by 3 M GdnHCl significantly prolonged the incubation periods in mice compared to that for the untreated control. More strikingly, the denaturation and removal of this region nearly abolished the infectivity. This finding suggests that the conformation of the region between aa 81 and 137 of the Chandler strain PrPSc molecule is directly associated with prion infectivity.

scholarly journals Emergence of multiple prion strains from single isolates of ovine scrapie

2011 ◽  
Vol 92 (6) ◽  
pp. 1482-1491 ◽  
Author(s):  
Alana M. Thackray ◽  
Lee Hopkins ◽  
Richard Lockey ◽  
John Spiropoulos ◽  
Raymond Bujdoso
Keyword(s):  
Transgenic Mice ◽  
Incubation Time ◽  
Infectious Agent ◽  
Serial Passage ◽  
Molecular Profile ◽  
Strain Typing ◽  
Wild Type ◽  
Diverse Range ◽  
Prion Strain ◽  
Prion Strains

The infectious agent associated with prion diseases such as ovine scrapie shows strain diversity. Ovine prion strains have typically been identified by their transmission properties in wild-type mice. However, strain typing of ovine scrapie isolates in wild-type mice may not reveal properties of the infectious prion agent as they exist in the original host. This could be circumvented if ovine scrapie isolates are passaged in ovine prion protein (PrP)-transgenic mice. This study used incubation time, lesion profile, immunohistochemistry of the disease-associated PrP (PrPSc) and molecular profile to compare the range of ovine prion strains that emerged from sheep scrapie isolates following serial passage in wild-type and ovine PrP transgenic mice. It was found that a diverse range of ovine prion strains emerged from homozygous ARQ and VRQ scrapie isolates passaged in wild-type and ovine PrP transgenic mice. However, strain-specific PrPSc deposition and PrP27–30 molecular profile patterns were identified in ovine PrP transgenic mice that were not detected in wild-type mice. Significantly, it was established that the individual mouse brain selected for transmission during prion strain typing had a significant influence on strain definition. Serial passage of short- and long-incubation-time animals from the same group of scrapie-inoculated mice revealed different prion strain phenotypes. These observations are consistent with the possibility that some scrapie isolates contain more than one prion strain.

scholarly journals Prions from Sporadic Creutzfeldt-Jakob Disease Patients Propagate as Strain Mixtures

mBio ◽  
2020 ◽  
Vol 11 (3) ◽  
Author(s):  
Hervé Cassard ◽  
Alvina Huor ◽  
Juan-Carlos Espinosa ◽  
Jean-Yves Douet ◽  
Severine Lugan ◽  
...  
Keyword(s):  
Relative Proportion ◽  
Significant Proportion ◽  
Proteinase K ◽  
Single Patient ◽  
Prion Strain ◽  
Prion Strains ◽  
Jakob Disease ◽  
The Brain

ABSTRACT Sporadic Creutzfeldt-Jakob disease (sCJD) cases are currently classified according to the methionine/valine polymorphism at codon 129 of the PRNP gene and the proteinase K-digested abnormal prion protein (PrPres) isoform identified by Western blotting (type 1 or type 2). Converging evidence led to the view that MM/MV1, VV/MV2, and VV1 and MM2 sCJD cases are caused by distinct prion strains. However, in a significant proportion of sCJD patients, both type 1 and type 2 PrPres were reported to accumulate in the brain, which raised questions about the diversity of sCJD prion strains and the coexistence of two prion strains in the same patient. In this study, a panel of sCJD brain isolates (n = 29) that displayed either a single or mixed type 1/type 2 PrPres were transmitted into human-PrP-expressing mice (tgHu). These bioassays demonstrated that two distinct prion strains (M1CJD and V2CJD) were associated with the development of sCJD in MM1/MV1 and VV2/MV2 patients. However, in about 35% of the investigated VV and MV cases, transmission results were consistent with the presence of both M1CJD and V2CJD strains, including in patients who displayed a “pure” type 1 or type 2 PrPres. The use of a highly sensitive prion in vitro amplification technique that specifically probes the V2CJD strain revealed the presence of the V2CJD prion in more than 80% of the investigated isolates, including isolates that propagated as a pure M1CJD strain in tgHu. These results demonstrate that at least two sCJD prion strains can be present in a single patient. IMPORTANCE sCJD occurrence is currently assumed to result from spontaneous and stochastic formation of a misfolded PrP nucleus in the brains of affected patients. This original nucleus then recruits and converts nascent PrPC into PrPSc, leading to the propagation of prions in the patient’s brain. Our study demonstrates the coexistence of two prion strains in the brains of a majority of the 23 sCJD patients investigated. The relative proportion of these sCJD strains varied both between patients and between brain areas in a single patient. These findings strongly support the view that the replication of an sCJD prion strain in the brain of a patient can result in the propagation of different prion strain subpopulations. Beyond its conceptual importance for our understanding of prion strain properties and evolution, the sCJD strain mixture phenomenon and its frequency among patients have important implications for the development of therapeutic strategies for prion diseases.

scholarly journals The effects of prion protein proteolysis and disaggregation on the strain properties of hamster scrapie

2008 ◽  
Vol 89 (10) ◽  
pp. 2642-2650 ◽  
Author(s):  
Andrea M. Deleault ◽  
Nathan R. Deleault ◽  
Brent T. Harris ◽  
Judy R. Rees ◽  
Surachai Supattapone
Keyword(s):  
Physical Properties ◽  
Conformational Stability ◽  
Proteinase K ◽  
Prion Strain ◽  
Biochemical Characteristics ◽  
Strain Diversity ◽  
Protease Digestion ◽  
Scrapie Strains ◽  
Strain Dependent ◽  
Mammalian Prions

Native mammalian prions exist in self-propagating strains that exhibit distinctive clinical, pathological and biochemical characteristics. Prion strain diversity is associated with variations in PrPSc conformation, but it remains unknown precisely which physical properties of the PrPSc molecules are required to encipher mammalian prion strain phenotypes. In this study, we subjected prion-infected brain homogenates derived from three different hamster scrapie strains to either (i) proteinase K digestion or (ii) sonication, and inoculated the modified samples into normal hamsters. The results show that the strain-specific clinical features and neuropathological profiles of inoculated animals were not affected by either treatment. Similarly, the strain-dependent biochemical characteristics of the PrPSc molecules (including electrophoretic mobility, glycoform composition, conformational stability and susceptibility to protease digestion) in infected animals were unaffected by either proteolysis or sonication of the original inocula. These results indicate that the infectious strain properties of native prions do not appear to be altered by PrPSc disaggregation, and that maintenance of such properties does not require the N-domain (approximately residues 23–90) of the protease-resistant PrPSc molecules or protease-sensitive PrPSc molecules.

scholarly journals Coinfecting Prion Strains Compete for a Limiting Cellular Resource

2010 ◽  
Vol 84 (11) ◽  
pp. 5706-5714 ◽  
Author(s):  
Ronald A. Shikiya ◽  
Jacob I. Ayers ◽  
Charles R. Schutt ◽  
Anthony E. Kincaid ◽  
Jason C. Bartz
Keyword(s):  
Protein Misfolding ◽  
Neuronal Loss ◽  
Infectious Agent ◽  
Prion Strain ◽  
Prion Strains ◽  
The Central Nervous System ◽  
Transmissible Mink Encephalopathy ◽  
Prion Conversion ◽  
Cellular Components

ABSTRACT Prion strain interference can influence the emergence of a dominant strain from a mixture; however, the mechanisms underlying prion strain interference are poorly understood. In our model of strain interference, inoculation of the sciatic nerve with the drowsy (DY) strain of the transmissible mink encephalopathy (TME) agent prior to superinfection with the hyper (HY) strain of TME can completely block HY TME from causing disease. We show here that the deposition of PrPSc, in the absence of neuronal loss or spongiform change, in the central nervous system corresponds with the ability of DY TME to block HY TME infection. This suggests that DY TME agent-induced damage is not responsible for strain interference but rather prions compete for a cellular resource. We show that protein misfolding cyclic amplification (PMCA) of DY and HY TME maintains the strain-specific properties of PrPSc and replicates infectious agent and that DY TME can interfere, or completely block, the emergence of HY TME. DY PrPSc does not convert all of the available PrPC to PrPSc in PMCA, suggesting the mechanism of prion strain interference is due to the sequestering of PrPC and/or other cellular components required for prion conversion. The emergence of HY TME in PMCA was controlled by the initial ratio of the TME agents. A higher ratio of DY to HY TME agent is required for complete blockage of HY TME in PMCA compared to several previous in vivo studies, suggesting that HY TME persists in animals coinfected with the two strains. This was confirmed by PMCA detection of HY PrPSc in animals where DY TME had completely blocked HY TME from causing disease.

scholarly journals Synergistic and strain-specific effects of bovine spongiform encephalopathy and scrapie prions in the cell-free conversion of recombinant prion protein

2006 ◽  
Vol 87 (12) ◽  
pp. 3753-3761 ◽  
Author(s):  
Martin Eiden ◽  
Gottfried J. Palm ◽  
Winfried Hinrichs ◽  
Ulrich Matthey ◽  
Ralph Zahn ◽  
...  
Keyword(s):  
Bovine Spongiform Encephalopathy ◽  
Prion Protein ◽  
De Novo ◽  
Proteinase K ◽  
Spongiform Encephalopathy ◽  
Cooperative Action ◽  
Specific Effects ◽  
Scrapie Strains ◽  
Recombinant Prion Protein

This study describes the conversion of murine PrPC by PrPSc from three different mouse scrapie strains (ME7, 87V and 22A) and from a mouse-passaged bovine spongiform encephalopathy (BSE) strain (BSE/Bl6). This was demonstrated by a modified, non-radioactive, cell-free conversion assay using bacterial prion protein, which was converted into a proteinase K (PK)-resistant fragment designated PrPres. Using this assay, newly formed PrPres could be detected by an antibody that discriminated de novo PrPres and the original PrPSc seed. The results suggested that PrPres formation occurs in three phases: the first 48 h when PrPres formation is delayed, followed by a period of substantially accelerated PrPres formation and a plateau phase when a maximum concentration of PrPres is reached after 72 h. The conversion of prokaryotically expressed PrPC by ME7 and BSE prions led to unglycosylated, PK-digested, abnormal PrPres fragments, which differed in molecular mass by 1 kDa. Therefore, prion strain phenotypes were retained in the cell-free conversion, even when recombinant PrPC was used as the substrate. Moreover, co-incubation of ME7 and BSE prions resulted in equal amounts of both ME7- and BSE-derived PrPres fragments (as distinguished by their different molecular sizes) and also in a significantly increased total amount of de novo-generated PrPres. This was found to be more than twice the amount of either strain when incubated separately. This result indicates a synergistic effect of both strains during cell-free conversion. It is not yet known whether such a cooperative action between BSE and scrapie prions also occurs in vivo.

scholarly journals Prion Strains and Transmission Barrier Phenomena

Pathogens ◽  
2018 ◽  
Vol 7 (1) ◽  
pp. 5 ◽  
Author(s):  
◽  
◽  
◽  
Keyword(s):  
Human Health ◽  
Current Knowledge ◽  
Species Barrier ◽  
Barrier Crossing ◽  
Prion Strain ◽  
Differential Selection ◽  
Physical Support ◽  
Prion Strains ◽  
Selection Of ◽  
Host Characteristics

Several experimental evidences show that prions are non-conventional pathogens, which physical support consists only in proteins. This finding raised questions regarding the observed prion strain-to-strain variations and the species barrier that happened to be crossed with dramatic consequences on human health and veterinary policies during the last 3 decades. This review presents a focus on a few advances in the field of prion structure and prion strains characterization: from the historical approaches that allowed the concept of prion strains to emerge, to the last results demonstrating that a prion strain may in fact be a combination of a few quasi species with subtle biophysical specificities. Then, we will focus on the current knowledge on the factors that impact species barrier strength and species barrier crossing. Finally, we present probable scenarios on how the interaction of strain properties with host characteristics may account for differential selection of new conformer variants and eventually species barrier crossing.

scholarly journals Molecular and Transmission Characteristics of Primary-Passaged Ovine Scrapie Isolates in Conventional and Ovine PrP Transgenic Mice

2008 ◽  
Vol 82 (22) ◽  
pp. 11197-11207 ◽  
Author(s):  
Alana M. Thackray ◽  
Lee Hopkins ◽  
John Spiropoulos ◽  
Raymond Bujdoso
Keyword(s):  
Transgenic Mice ◽  
Transmission Characteristics ◽  
Prion Strain ◽  
Prion Strains ◽  
True Diversity ◽  
Incubation Periods ◽  
Primary Transmission ◽  
Molecular Nature ◽  
Sheep Scrapie ◽  
Mouse Lines

ABSTRACT A more complete assessment of ovine prion strain diversity will be achieved by complementing biological strain typing in conventional and ovine PrP transgenic mice with a biochemical analysis of the resultant PrPSc. This will provide a correlation between ovine prion strain phenotype and the molecular nature of different PrP conformers associated with particular prion strains. Here, we have compared the molecular and transmission characteristics of ovine ARQ/ARQ and VRQ/VRQ scrapie isolates following primary passage in tg338 (VRQ) and tg59 (ARQ) ovine PrP transgenic mice and the conventional mouse lines C57BL/6 (Prnp a ), RIII (Prnp a ), and VM (Prnp b ). Our data show that these different genotypes of scrapie isolates display similar incubation periods of >350 days in conventional and tg59 mice. Facilitated transmission of sheep scrapie isolates occurred in tg338 mice, with incubation times reduced to 64 days for VRQ/VRQ inocula and to ≤210 days for ARQ/ARQ samples. Distinct genotype-specific lesion profiles were seen in the brains of conventional and tg59 mice with prion disease, which was accompanied by the accumulation of more conformationally stable PrPSc, following inoculation with ARQ/ARQ compared to VRQ/VRQ scrapie isolates. In contrast, the lesion profiles, quantities, and stability of PrPSc induced by the same inocula in tg338 mice were more similar than in the other mouse lines. Our data show that primary transmission of different genotypes of ovine prions is associated with the formation of different conformers of PrPSc with distinct molecular properties and provide the basis of a molecular approach to identify the true diversity of ovine prion strains.

scholarly journals PrPSc-Specific Antibody Reveals C-Terminal Conformational Differences between Prion Strains

2016 ◽  
Vol 90 (10) ◽  
pp. 4905-4913 ◽  
Author(s):  
Eri Saijo ◽  
Andrew G. Hughson ◽  
Gregory J. Raymond ◽  
Akio Suzuki ◽  
Motohiro Horiuchi ◽  
...  
Keyword(s):  
Specific Antibody ◽  
Enzyme Linked Immunosorbent Assay ◽  
Strain Variation ◽  
Prion Strains ◽  
Disease Biology ◽  
Folded Structure ◽  
C Terminus ◽  
Wide Range ◽  
Scrapie Strains ◽  
Strain Dependent

ABSTRACTUnderstanding the structure of PrPScand its strain variation has been one of the major challenges in prion disease biology. To study the strain-dependent conformations of PrPSc, we purified proteinase-resistant PrPSc(PrPRES) from mouse brains with three different murine-adapted scrapie strains (Chandler, 22L, and Me7) and systematically tested the accessibility of epitopes of a wide range of anti-PrP and anti-PrPScspecific antibodies by indirect enzyme-linked immunosorbent assay (ELISA). We found that epitopes of most anti-PrP antibodies were hidden in the folded structure of PrPRES, even though these epitopes are revealed with guanidine denaturation. However, reactivities to a PrPSc-specific conformational C-terminal antibody showed significant differences among the three different prion strains. Our results provide evidence for strain-dependent conformational variation near the C termini of molecules within PrPScmultimers.IMPORTANCEIt has long been apparent that prion strains can have different conformations near the N terminus of the PrPScprotease-resistant core. Here, we show that a C-terminal conformational PrPSc-specific antibody reacts differently to three murine-adapted scrapie strains. These results suggest, in turn, that conformational differences in the C terminus of PrPScalso contribute to the phenotypic distinction between prion strains.

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