Abstract
Prions are unconventional infectious agents thought to be primarily composed of PrPSc, a multimeric misfolded conformer of the ubiquitously expressed host-encoded prion protein (PrPC). They cause fatal neurodegenerative diseases in both animals and humans. The disease phenotype is not uniform within species, and stable, self-propagating variations in PrPSc conformation could encode this ‘strain’ diversity. However, much remains to be learned about the physical relationship between the infectious agent and PrPSc aggregation state, and how this varies according to the strain. We applied a sedimentation velocity technique to a panel of natural, biologically cloned strains obtained by propagation of classical and atypical sheep scrapie and BSE infectious sources in transgenic mice expressing ovine PrP. Detergent-solubilized, infected brain homogenates were used as starting material. Solubilization conditions were optimized to separate PrPSc aggregates from PrPC. The distribution of PrPSc and infectivity in the gradient was determined by immunoblotting and mouse bioassay, respectively. As a general feature, a major proteinase K-resistant PrPSc peak was observed in the middle part of the gradient. This population approximately corresponds to multimers of 12–30 PrP molecules, if constituted of PrP only. For two strains, infectivity peaked in a markedly different region of the gradient. This most infectious component sedimented very slowly, suggesting small size oligomers and/or low density PrPSc aggregates. Extending this study to hamster prions passaged in hamster PrP transgenic mice revealed that the highly infectious, slowly sedimenting particles could be a feature of strains able to induce a rapidly lethal disease. Our findings suggest that prion infectious particles are subjected to marked strain-dependent variations, which in turn could influence the strain biological phenotype, in particular the replication dynamics.
Highlights
Transmissible spongiform encephalopathies (TSE), such as human Creutzfeldt-Jakob disease, sheep scrapie, bovine spongiform encephalopathy (BSE) and chronic wasting disease of cervidae, are infectious, fatal, neurodegenerative disorders caused by prions [1]
Prions are unconventional transmissible agents causing fatal neurodegenerative diseases in human and animals. They are thought to be formed from polymers of abnormal conformations of the host-encoded prion protein (PrP), but little is known about the physical organization of the infectious particles and any relationship between packing order and infectivity
We subjected PrP polymers from eight different ovine and hamster prion strains to sedimentation velocity centrifugation, which allows separation of macromolecular complexes according to their size, density or shape
Summary
Transmissible spongiform encephalopathies (TSE), such as human Creutzfeldt-Jakob disease, sheep scrapie, bovine spongiform encephalopathy (BSE) and chronic wasting disease of cervidae, are infectious, fatal, neurodegenerative disorders caused by prions [1]. Prions are unconventional pathogens primarily composed of PrPSc, a rearranged conformer of the ubiquitously expressed prion protein (PrPC), whose precise physiological function is largely unknown. PrPSc dictates the self-perpetuating conformational conversion of PrPC into nascent PrPSc. Upon infection, PrPSc dictates the self-perpetuating conformational conversion of PrPC into nascent PrPSc This conversion involves – without any apparent posttranslational modification – the refolding of soluble, alpha-helixrich PrPC molecules into beta-sheet enriched PrPSc polymers that form deposits in TSE-infected brains [2,3] and are assumed to be responsible for the observed neurodegenerative disorders [4]. The conversion reaction may proceed through a nucleated polymerization mechanism in which PrPSc multimers recruit PrPC molecules and trigger their conformational conversion into PrPSc (for review [5]). The refolding/multimerisation process confers distinct physico-chemical properties to PrPSc, such as insolubility in non-denaturing detergents and partial resistance to proteolysis [6]
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