Subtyping of human cellular prion proteins and their differential solubility

A human form of a prion disorder is the Creutzfeldt-Jakob disease. A hallmark of the disease is the accumulation of misfolded prion proteins (PrP^Sc), which exist as heterogeneous subtypes. PrP^Sc is formed by protein conversion from the host-encoded cellular prion (PrP^C), which is expressed and modified to various isoforms. Little is known about variation in PrP^C; however, it is assumed that PrP^C types play important roles in the formation of PrP^Sc. In this study, we separated distinct human PrP^C subtypes on the basis of differential protein solubilities in detergent solutions. Single and sequential application of the detergents Triton X-100, octyl-glucopyranoside and CHAPS facilitated high solubility of glycosylated PrP^C isoforms, whereas high proportions of nonglycosylated PrP^C remained non-soluble. Most proteins became highly soluble with laurylsarcosine and sodium dodecyl sulphate. Our findings demonstrate that the solubility characteristics of heterogeneous PrP^C overlap in human brains and convey distinct solubility subtypes. Differentiation by solubility experiments can therefore provide valuable information on prion protein composition, facilitate the separation of subtypes, and offer new prospects for conversion specificity of distinct isoforms.     

Persistent retroviral infection with MoMuLV influences neuropathological signature and phenotype of prion disease

A fundamental step in pathophysiology of prion diseases is the conversion of the host encoded prion protein (PrP^C) into a misfolded isoform (PrP^Sc) that accumulates mainly in neuronal but also non-neuronal tissues. Prion diseases are transmissible within and between species. In a subset of prion diseases, peripheral prion uptake and subsequent transport to the central nervous system are key to disease initiation. The involvement of retroviruses in this process has been postulated based on the findings that retroviral infections enhance the spread of prion infectivity and PrP^Sc from cell to cell in vitro. To study whether retroviral infection influences the phenotype of prion disease or the spread of prion infectivity and PrP^Sc in vivo, we developed a murine model with persistent Moloney murine leukemia retrovirus (MoMuLV) infection with and without additional prion infection. We investigated the pathophysiology of prion disease in MoMuLV and prion-infected mice, monitoring temporal kinetics of PrP^Sc spread and prion infectivity, as well as clinical presentation. Unexpectedly, infection of MoMuLV challenged mice with prions did not change incubation time to clinical prion disease. However, clinical presentation of prion disease was altered in mice infected with both pathogens. This was paralleled by remarkably enhanced astrogliosis and pathognomonic astrocyte morphology in the brain of these mice. Therefore, we conclude that persistent viral infection might act as a disease modifier in prion disease.     

Novel Compounds Identified by Structure-Based Prion Disease Drug Discovery Using In Silico Screening Delay the Progression of an Illness in Prion-Infected Mice

The accumulation of abnormal prion protein (PrP^Sc) produced by the structure conversion of PrP (PrP^C) in the brain induces prion disease. Although the conversion process of the protein is still not fully elucidated, it has been known that the intramolecular chemical bridging in the most fragile pocket of PrP, known as the “hot spot,” stabilizes the structure of PrP^C and inhibits the conversion process. Using our original structure-based drug discovery algorithm, we identified the low molecular weight compounds that predicted binding to the hot spot. NPR-130 and NPR-162 strongly bound to recombinant PrP in vitro, and fragment molecular orbital (FMO) analysis indicated that the high affinity of those candidates to the PrP is largely dependent on nonpolar interactions, such as van der Waals interactions. Those NPRs showed not only significant reduction of the PrP^Sc levels but also remarkable decrease of the number of aggresomes in persistently prion-infected cells. Intriguingly, treatment with those candidate compounds significantly prolonged the survival period of prion-infected mice and suppressed prion disease-specific pathological damage, such as vacuole degeneration, PrP^Sc accumulation, microgliosis, and astrogliosis in the brain, suggesting their possible clinical use. Our results indicate that in silico drug discovery using NUDE/DEGIMA may be widely useful to identify candidate compounds that effectively stabilize the protein.Electronic supplementary materialThe online version of this article (10.1007/s13311-020-00903-9) contains supplementary material, which is available to authorized users.     

Molecular biology and pathology of prion strains in sporadic human prion diseases

Prion diseases are believed to propagate by the mechanism involving self-perpetuating conformational conversion of the normal form of the prion protein, PrP^C, to the misfolded, pathogenic state, PrP^Sc. One of the most intriguing aspects of these disorders is the phenomenon of prion strains. It is believed that strain properties are fully encoded in distinct conformations of PrP^Sc. Strains are of practical relevance to human prion diseases as their diversity may explain the unusual heterogeneity of these disorders. The first insight into the molecular mechanisms underlying heterogeneity of human prion diseases was provided by the observation that two distinct disease phenotypes and their associated PrP^Sc conformers co-distribute with distinct PrP genotypes as determined by the methionine/valine polymorphism at codon 129 of the PrP gene. Subsequent studies identified six possible combinations of the three genotypes (determined by the polymorphic codon 129) and two common PrP^Sc conformers (named types 1 and 2) as the major determinants of the phenotype in sporadic human prion diseases. This scenario implies that each 129 genotype–PrP^Sc type combination would be associated with a distinct disease phenotype and prion strain. However, notable exceptions have been found. For example, two genotype–PrP^Sc type combinations are linked to the same phenotype, and conversely, the same combination was found to be associated with two distinct phenotypes. Furthermore, in some cases, PrP^Sc conformers naturally associated with distinct phenotypes appear, upon transmission, to lose their phenotype-determining strain characteristics. Currently it seems safe to assume that typical sporadic prion diseases are associated with at least six distinct prion strains. However, the intrinsic characteristics that distinguish at least four of these strains remain to be identified.     

Virus-induced alterations of membrane lipids affect the incorporation of PrP Sc into cells

Prion diseases are fatal neurodegenerative disorders characterized by long incubation periods. To investigate whether concurrent diseases can modify the clinical outcome of prion‐affected subjects, we tested the effect of viral infection on the binding and internalization of PrP^Sc, essential steps of prion propagation. To this effect, we added scrapie brain homogenate or purified PrP^Sc to fibroblasts previously infected with minute virus of mice (MVM), a mouse parvovirus. We show here that the rate of incorporation of PrP^Sc into MVM‐infected cells was significantly higher than that observed for naïve cells. Immunostaining of cells and immunoblotting of subcellular fractions using antibodies recognizing PrP and LysoTracker, a lysosomal marker, revealed that in both control and MVM‐infected cells the incorporated PrP^Sc was associated mostly with lysosomes. Interestingly, floatation gradient analysis revealed that the majority of the PrP^Sc internalized into MVM‐infected cells shifted toward raft‐containing low‐density fractions. Concomitantly, the MVM‐infected cells demonstrated increased levels of the glycosphingolipid GM1 (an essential raft lipid component) throughout the gradient and a shift in caveolin 1 (a raft protein marker) toward lighter membrane fractions compared with noninfected cells. Our results suggest that the effect of viral infection on membrane lipid composition may promote the incorporation of exogenous PrP^Sc into rafts. Importantly, membrane rafts are believed to be the conversion site of PrP^C to PrP^Sc; therefore, the association of exogenous PrP^Sc with such membrane microdomains may facilitate prion infection. © 2008 Wiley‐Liss, Inc.     

Distribution of the cellular prion protein (PrPC) in brains of livestock and domesticated species

In transmissible spongiform encephalopathies (TSEs) the prion protein (PrP) plays a central role in pathogenesis. The PrP gene (Prnp) has been described in a number of mammalian and avian species and its expression product, the cellular prion protein (PrP^C), has been mapped in brains of different laboratory animals (rodent and non-human primates). However, mapping of PrP^C expression in mammalian species suffering from natural (bovine and ovine) and experimental (swine) TSE or in species in which prion disease has never been reported (equine and canine) deserves further attention. Thus, localising the cellular prion protein (PrP^C) distribution in brain may be noteworthy for the understanding of prion disease pathogenesis since lesions seem to be restricted to particular brain areas. In the present work, we analysed the distribution of PrP^C expression among several brain structures of the above species. Our results suggest that the expression of PrP^C, within the same species, differs depending on the brain structure studied, but no essential differences between the PrP^C distribution patterns among the studied species could be established. Positive immunoreaction was found mainly in the neuropil and to a lesser extent in neuronal bodies which occasionally appeared strongly stained in discrete regions. Overall, the expression of PrP^C in the brain was significantly higher in grey matter areas than in white matter, where accumulation of PrP^Sc is first observed in prion diseases. Therefore, other factors besides the level of expression of cellular PrP may account for the pathogenesis of TSEs     

Recombinant prion protein induces a new transmissible prion disease in wild-type animals

Prion disease is a neurodegenerative malady, which is believed to be transmitted via a prion protein in its abnormal conformation (PrP^Sc). Previous studies have failed to demonstrate that prion disease could be induced in wild-type animals using recombinant prion protein (rPrP) produced in Escherichia coli. Here, we report that prion infectivity was generated in Syrian hamsters after inoculating full-length rPrP that had been converted into the cross-β-sheet amyloid form and subjected to annealing. Serial transmission gave rise to a disease phenotype with highly unique clinical and neuropathological features. Among them were the deposition of large PrP^Sc plaques in subpial and subependymal areas in brain and spinal cord, very minor lesioning of the hippocampus and cerebellum, and a very slow progression of disease after onset of clinical signs despite the accumulation of large amounts of PrP^Sc in the brain. The length of the clinical duration is more typical of human and large animal prion diseases, than those of rodents. Our studies establish that transmissible prion disease can be induced in wild-type animals by inoculation of rPrP and introduce a valuable new model of prion diseases.     

Neurodevelopmental expression and localization of the cellular prion protein in the central nervous system of the mouse

Transmissible spongiform encephalopathies (TSEs) are neurodegenerative disorders caused by PrP^Sc, or prion, an abnormally folded form of the cellular prion protein (PrP^C). The abundant expression of PrP^C in the central nervous system (CNS) is a requirement for prion replication, yet despite years of intensive research the physiological function of PrP^C still remains unclear. Several routes of investigation point out a potential role for PrP^C in axon growth and neuronal development. Thus, we undertook a detailed analysis of the spatial and temporal expression of PrP^C during mouse CNS development. Our findings show regional differences of the expression of PrP, with some specific white matter structures showing the earliest and highest expression of PrP^C. Indeed, all these regions are part of the thalamolimbic neurocircuitry, suggesting a potential role of PrP^C in the development and functioning of this specific brain system. J. Comp. Neurol. 518:1879–1891, 2010. © 2010 Wiley‐Liss, Inc.     

Interplay between 20S proteasomes and prion proteins in scrapie disease

Scrapie is a transmissible spongiform encephalopathy affecting the central nervous system in sheep. The key event in such neurodegeneration is the conversion of the normal prion protein (PrP^C) into the pathological isoform (PrP^Sc). Misfolded prion proteins are normally degraded by the proteasome. This work, analyzing models of scrapie disease, describes the in vivo relationship between the proteasome and prions. We report that the disease is associated with an increase of proteasome functionality, most likely as a means of counteracting the increased levels of oxidative stress. Here, we show that prions coprecipitate with the 20S proteasome and that they colocalize within the same neuron, thus raising the possibility that PrP interacts with the proteasome in both normal and diseased brain, affecting substrate trafficking and proteasome functionality. This interaction, inducing proteasome activation, leads to different neuronal alterations and triggers apoptosis. Furthermore, testing the effects of isolated PrP^C on purified 20S proteasomes, we obtain a concentration‐ and proteasome composition‐dependent decrease in the complex activity. © 2009 Wiley‐Liss, Inc.     

On the biology of prions

Summary Prions cause scrapie and Creutzfeldt-Jakob disease (CJD); these infectious pathogens are composed largely, if not entirely, of protein molecules. No prion-specific polynucleotide has been identified. Purified preparations of scrapie prions contain high titers (10^9.5 ID₅₀/ml), one protein (PrP 27-30) and amyloid rods (10–20 nm in diameter ×100–200 nm in length). Considerable evidence indicates that PrP 27-30 is required for and inseparable from scrapie infectivity. PrP 27-30 is encoded by a cellular gene and is derived from a larger protein, denoted PrP^Sc or PrP 33-35^Sc, by protease digestion. A cellular isoform, designated PrP^C or PrP 33-35^C, is encoded by the same gene as PrP^Sc and both proteins appear to be translated from the same 2.1 kb mRNA. Monoclonal antibodies to PrP 27-30, as well as antisera to PrP synthetic peptides, specifically react with both PrP^C and PrP^Sc, establishing their relatedness. PrP^C is digested by proteinase K, while PrP^Sc is converted to PrP 27-30 under the same conditions. Prion proteins are synthesized with signal peptides and are integrated into membranes. Detergent extraction of microsomal membranes isolated from scrapie-infected hamster brains solubilizes PrP^C but induces PrP^Sc to polymerize into amyloid rods. This procedure allows separation of the two prion protein isoforms and the demonstration that PrP^Sc accumulates during scrapie infection, while the level of PrP^C does not change. The prion amyloid rods generated by detergent extraction are identical morphologically, except for length, to extracellular collections of prion amyloid filaments which form plaques in scrapie- and CJD-infected brains. The prion amyloid plaques stain with antibodies to PrP 27-30 and PrP peptides. PrP 33-35^C does not accumulate in the extracellular space. Prion rods composed of PrP 27-30 can be dissociated into phospholipid vesicles with full retention of scrapie infectivity. The murine PrP gene (Prn-p) is linked to thePrn-i gene which controls the length of the scrapie incubation period. Prolonged incubation times are a cardinal feature of scrapie and CJD. While the central role of PrP^Sc in scrapie pathogenesis is well established, the chemical as well as conformational differences between PrP^C and PrP^Sc are unknown but probably arise from post-translational modifications.Supported by research grants from the National Institutes of Health (AG 02132 and NS 14069) and a Senator Jacob Javits Center of Excellence in Neuroscience (NS 22786) as well as by gifts from RJR-Nabisco, Inc. and Sherman Fairchild FoundationThis review is based upon a plenary lecture entitled Biology and Neuropathology of Prions presented at the Xth International Congress of Neuropathology, Stockholm, Sweden, September 11, 1986, and is dedicated to the memory of Peter Wilhelm Lampert (1929–1986)     

Intracellular mechanisms mediating the neuronal death and astrogliosis induced by the prion protein fragment 106–126

Prion encephalopathies include fatal diseases of the central nervous system of men and animals characterized by nerve cell loss, glial proliferation and deposition of amyloid fibrils into the brain. During these diseases a cellular glycoprotein (the prion protein, PrP^C) is converted, through a not yet completely clear mechanism, in an altered isoform (the prion scrapie, PrP^Sc) that accumulates within the brain tissue by virtue of its resistance to the intracellular catabolism. PrP^Sc is believed to be responsible for the neuronal loss that is observed in the prion disease. The PrP 106–126, a synthetic peptide that has been obtained from the amyloidogenic portion of the prion protein, represents a suitable model for studying the pathogenic role of the PrP^Sc, retaining, in vitro, some characteristics of the entire protein, such as the capability to aggregate in fibrils, and the neurotoxicity. In this work we present the results we have recently obtained regarding the action of the PrP 106–126 in different cellular models. We report that the PrP 106–126 induces proliferation of cortical astrocytes, as well as degeneration of primary cultures of cortical neurons or of neuroectodermal stable cell lines (GH₃ cells). In particular, these two opposite effects are mediated by the same attitude of the peptide to interact with the L-type calcium channels: in the astrocytes, the activity of these channels seems to be activated by PrP 106–126, while, in the cortical neurons and in the GH₃ cells, the same treatment causes a blockade of these channels causing a toxic effect.     

Molecular pathogenesis of prion diseases

Prion diseases such as Creutzfeldt-Jakob disease in humans, scrapie in sheep, and BSE in cattle are transmissible and fatal neurodegenerative diseases. The infectious agent of these diseases has been designated as “prion”. It consists mainly and perhaps exclusively of a conformational variant of a physiological glycoprotein, the cellular prion, protein, PrP^C, which is a copper-binding protein of the cell surface. In spite of the wealth of biochemical and biophysical information, the conformational transition from PrP^C to PrP^Sc, the infectious isoform of the prion protein, is not well understood. Nerve cell loss in prion diseases may be caused by neurotoxic effects of the prion protein. Certain properties of the prion protein such as the apparent form of its glycosylation and conformational properties reflected by the preferential site of digestion with proteinase K are associated with particular phenotypes of prion disease. The appearance of a new variant of Creutzfeldt-Jakob disease in humans, which is most likely caused by the consumption of BSE-infected food in the UK, is cause for major concern particularly since there is no known effective treatment of prion diseases.     

Incidence and spectrum of sporadic Creutzfeldt–Jakob disease variants with mixed phenotype and co-occurrence of PrPSc types: an updated classification

Six subtypes of sporadic Creutzfeldt–Jakob disease with distinctive clinico-pathological features have been identified largely based on two types of the abnormal prion protein, PrP^Sc, and the methionine (M)/valine (V) polymorphic codon 129 of the prion protein. The existence of affected subjects showing mixed phenotypic features and concurrent PrP^Sc types has been reported but with inconsistencies among studies in both results and their interpretation. The issue currently complicates diagnosis and classification of cases and also has implications for disease pathogenesis. To explore the issue in depth, we carried out a systematic regional study in a large series of 225 cases. PrP^Sc types 1 and 2 concurrence was detected in 35% of cases and was higher in MM than in MV or VV subjects. The deposition of either type 1 or 2, when concurrent, was not random and always characterized by the coexistence of phenotypic features previously described in the pure subtypes. PrP^Sc type 1 accumulation and related pathology predominated in MM and MV cases, while the type 2 phenotype prevailed in VVs. Neuropathological examination best identified the mixed types 1 and 2 features in MMs and most MVs, and also uniquely revealed the co-occurrence of pathological variants sharing PrP^Sc type 2. In contrast, molecular typing best detected the concurrent PrP^Sc types in VV subjects and MV cases with kuru plaques. The present data provide an updated disease classification and are of importance for future epidemiologic and transmission studies aimed to identify etiology and extent of strain variation in sporadic Creutzfeldt–Jakob disease.     

Quantification of surviving cerebellar granule neurones and abnormal prion protein (PrPSc) deposition in sporadic Creutzfeldt-Jakob disease supports a pathogenic role for small PrPSc deposits common to the various molecular subtypes

B. A. Faucheux, E. Morain, V. Diouron, J.‐P. Brandel, D. Salomon, V. Sazdovitch, N. Privat, J.‐L. Laplanche, J.‐J. Hauw and S. Haïk (2011) Neuropathology and Applied Neurobiology 37, 500–512 Quantification of surviving cerebellar granule neurones and abnormal prion protein (PrP^Sc) deposition in sporadic Creutzfeldt–Jakob disease supports a pathogenic role for small PrP^Sc deposits common to the various molecular subtypes Aims: Neuronal death is a major neuropathological hallmark in prion diseases. The association between the accumulation of the disease‐related prion protein (PrP^Sc) and neuronal loss varies within the wide spectrum of prion diseases and their experimental models. In this study, we investigated the relationships between neuronal loss and PrP^Sc deposition in the cerebellum from cases of the six subtypes of sporadic Creutzfeldt–Jakob disease (sCJD; n = 100) that can be determined according to the M129V polymorphism of the human prion protein gene (PRNP) and PrP^Sc molecular types. Methods: The numerical density of neurones was estimated with a computer‐assisted image analysis system and the accumulation of PrP^Sc deposits was scored. Results: The scores of PrP^Sc immunoreactive deposits of the punctate type (synaptic type) were correlated with neurone counts – the higher the score the higher the neuronal loss – in all sCJD subtypes. Large 5‐ to 50‐µm‐wide deposits (focal type) were found in sCJD‐MV2 and sCJD‐VV2 subtypes, and occasionally in a few cases of the other studied groups. By contrast, the highest scores for 5‐ to 50‐µm‐wide deposits observed in sCJD‐MV2 subtype were not associated with higher neuronal loss. In addition, these scores were inversely correlated with neuronal counts in the sCJD‐VV2 subtype. Conclusions: These results support a putative pathogenic role for small PrP^Sc deposits common to the various sCJD subtypes. Furthermore, the observation of a lower loss of neurones associated with PrP^Sc type‐2 large deposits is consistent with a possible ‘protective’ role of aggregated deposits in both sCJD‐MV2 and sCJD‐VV2 subtypes.     

Accumulation of prion protein in the vagus nerve in creutzfeldt–jakob disease

Disease‐associated proteins are thought to propagate along neuronal processes in neurodegenerative diseases. To detect disease‐associated prion protein (PrP^Sc) in the vagus nerve in different forms and molecular subtypes of Creutzfeldt–Jakob disease (CJD), we applied 3 different anti‐PrP antibodies. We screened the vagus nerve in 162 sporadic and 30 genetic CJD cases. Four of 31 VV‐2 type sporadic CJD and 7 of 30 genetic CJD cases showed vagal PrP^Sc immunodeposits with distinct morphology. Thus, PrP^Sc in CJD affects the vagus nerve analogously to α‐synuclein in Parkinson disease. The morphologically diverse deposition of PrP^Sc in genetic and sporadic CJD argues against uniform mechanisms of propagation of PrP^Sc. Ann Neurol 2019;85:782–787     

Abnormal Activation of Microglia Accompanied with Disrupted CX3CR1/CX3CL1 Pathway in the Brains of the Hamsters Infected with Scrapie Agent 263K

Microglial cells are resident mononuclear phagocytes of the central nervous system (CNS). Active proliferation of microglia in the brain has been identified in neurodegenerative disorders, including some kinds of prion disease. However, the detailed regional distribution between microglia and PrP^Sc deposition has not been presented, and investigation of fractalkine signaling which is involved in the regulation of activation of microglia in prion disease is not well documented. In this study, the disease phenomenon of microglial accumulation in the CNS was thoroughly analyzed using a scrapie-infected experimental model. Western blots of microglia-specific markers Iba1 and CD68, immunohistochemical and immunofluorescent assays demonstrated obviously activation of microglia in almost whole brain regions in the infected animals. Under the dynamic analysis on hallmarks of activation of microglia, a time-dependent increase of Iba1 and CD68 was detected, accompanied by accumulation of PrP^Sc and progression of neurodegenerative symptoms. With serial brain sections and double staining of Iba1 and PrP^Sc, we observed that the microglia distributed around PrP^Sc deposits in 263K-infected hamsters’ brains, proposing PrP^Sc phagocytosis. Flow cytometry assays with the single-cell suspensions prepared from the cortical region of the infected brains verified an activation of microglial population. ELISA assays of the cytokines in brain homogenates revealed significant upregulations of interleukin (IL)-1β, IL-6 and TNF-α when infected. Evaluation of fractalkine signaling in the infected hamsters’ brains showed progressively downregulation of CX3CL1 during the incubation. Prion peptide PrP106-126 also disrupted fractalkine and evoked microglial activation in rat primary neuron–glia mixed cultures. Our data here demonstrate an activated status of microglia in CNS tissues of infectious prion disease, possibly through fractalkine signaling deficiency.     

Anti-prion Drugs Targeting the Protein Folding Activity of the Ribosome Reduce PABPN1 Aggregation

Prion diseases are caused by the propagation of PrP^Sc, the pathological conformation of the PrP^C prion protein. The molecular mechanisms underlying PrP^Sc propagation are still unsolved and no therapeutic solution is currently available. We thus sought to identify new anti-prion molecules and found that flunarizine inhibited PrP^Sc propagation in cell culture and significantly prolonged survival of prion-infected mice. Using an in silico therapeutic repositioning approach based on similarities with flunarizine chemical structure, we tested azelastine, duloxetine, ebastine, loperamide and metixene and showed that they all have an anti-prion activity. Like flunarizine, these marketed drugs reduced PrP^Sc propagation in cell culture and in mouse cerebellum organotypic slice culture, and inhibited the protein folding activity of the ribosome (PFAR). Strikingly, some of these drugs were also able to alleviate phenotypes due to PABPN1 nuclear aggregation in cell and Drosophila models of oculopharyngeal muscular dystrophy (OPMD). These data emphasize the therapeutic potential of anti-PFAR drugs for neurodegenerative and neuromuscular proteinopathies.Supplementary InformationThe online version contains supplementary material available at 10.1007/s13311-020-00992-6.Key Words: Prions, PrP^Sc, drug repositioning, PFAR, OPMD, PABPN1     

Using Protein Misfolding Cyclic Amplification Generates a Highly Neurotoxic PrP Dimer Causing Neurodegeneration

Under the “protein-only” hypothesis, prion-based diseases are proposed to result from an infectious agent that is an abnormal isoform of the prion protein in the scrapie form, PrP^Sc. However, since PrP^Sc is highly insoluble and easily aggregates in vivo, this view appears to be overly simplistic, implying that the presence of PrP^Sc may indirectly cause neurodegeneration through its intermediate soluble form. We generated a neurotoxic PrP dimer with partial pathogenic characteristics of PrP^Sc by protein misfolding cyclic amplification in the presence of 1-palmitoyl-2-oleoylphosphatidylglycerol consisting of recombinant hamster PrP (23–231). After intracerebral injection of the PrP dimer, wild-type hamsters developed signs of neurodegeneration. Clinical symptoms, necropsy findings, and histopathological changes were very similar to those of transmissible spongiform encephalopathies. Additional investigation showed that the toxicity is primarily related to cellular apoptosis. All results suggested that we generated a new neurotoxic form of PrP, PrP dimer, which can cause neurodegeneration. Thus, our study introduces a useful model for investigating PrP-linked neurodegenerative mechanisms.