Membrane Toxicity of Abnormal Prion Protein in Adrenal Chromaffin Cells of Scrapie Infected Sheep

Transmissible spongiform encephalopathies (TSEs) or prion diseases are associated with accumulations of disease specific PrP (PrP^d) in the central nervous system (CNS) and often the lymphoreticular system (LRS). Accumulations have additionally been recorded in other tissues including the peripheral nervous system and adrenal gland. Here we investigate the effect of sheep scrapie on the morphology and the accumulation of PrP^d in the adrenal medulla of scrapie affected sheep using light and electron microscopy. Using immunogold electron microscopy, non-fibrillar forms of PrP^d were shown to accumulate mainly in association with chromaffin cells, occasional nerve endings and macrophages. PrP^d accumulation was associated with distinctive membrane changes of chromaffin cells including increased electron density, abnormal linearity and invaginations. Internalisation of PrP^d from the chromaffin cell plasma membrane occurred in association with granule recycling following hormone exocytosis. PrP^d accumulation and internalisation from membranes is similarly associated with perturbations of membrane structure and trafficking in CNS neurons and tingible body macrophages of the LRS. These data suggest that a major toxic effect of PrP^d is at the level of plasma membranes. However, the precise nature of PrP^d-membrane toxicity is tissue and cell specific suggesting that the normal protein may act as a multi-functional scaffolding molecule. We further suggest that the co-localisation of PrP^d with exocytic granules of the hormone trafficking system may provide an additional source of infectivity in blood.     

Scrapie-specific pathology of sheep lymphoid tissues

Transmissible spongiform encephalopathies (TSEs) or prion diseases often result in accumulation of disease-associated PrP (PrP(d)) in the lymphoreticular system (LRS), specifically in association with follicular dendritic cells (FDCs) and tingible body macrophages (TBMs) of secondary follicles. We studied the effects of sheep scrapie on lymphoid tissue in tonsils and lymph nodes by light and electron microscopy. FDCs of sheep were grouped according to morphology as immature, mature or regressing. Scrapie was associated with FDC dendrite hypertrophy and electron dense deposit or vesicles. PrP(d) was located using immunogold labelling at the plasmalemma of FDC dendrites and, infrequently, mature B cells. Abnormal electron dense deposits surrounding FDC dendrites were identified as immunoglobulins suggesting that excess immune complexes are retained and are indicative of an FDC dysfunction. Within scrapie-affected lymph nodes, macrophages outside the follicle and a proportion of germinal centre TBMs accumulated PrP(d) within endosomes and lysosomes. In addition, TBMs showed PrP(d) in association with the cell membrane, non-coated pits and vesicles, and also with discrete, large and random endoplasmic reticulum networks, which co-localised with ubiquitin. These observations suggest that PrP(d) is internalised via the caveolin-mediated pathway, and causes an abnormal disease-related alteration in endoplasmic reticulum structure. In contrast to current dogma, this study shows that sheep scrapie is associated with cytopathology of germinal centres, which we attribute to abnormal antigen complex trapping by FDCs and abnormal endocytic events in TBMs. The nature of the sub-cellular changes in FDCs and TBMs differs from those of scrapie infected neurones and glial cells suggesting that different PrP(d)/cell membrane interactions occur in different cell types.     

Evaluation of the Possible Transmission of BSE and Scrapie to Gilthead Sea Bream (Sparus aurata)

In transmissible spongiform encephalopathies (TSEs), a group of fatal neurodegenerative disorders affecting many species, the key event in disease pathogenesis is the accumulation of an abnormal conformational isoform (PrP^Sc) of the host-encoded cellular prion protein (PrP^C). While the precise mechanism of the PrP^C to PrP^Sc conversion is not understood, it is clear that host PrP^C expression is a prerequisite for effective infectious prion propagation. Although there have been many studies on TSEs in mammalian species, little is known about TSE pathogenesis in fish. Here we show that while gilthead sea bream (Sparus aurata) orally challenged with brain homogenates prepared either from a BSE infected cow or from scrapie infected sheep developed no clinical prion disease, the brains of TSE-fed fish sampled two years after challenge did show signs of neurodegeneration and accumulation of deposits that reacted positively with antibodies raised against sea bream PrP. The control groups, fed with brains from uninfected animals, showed no such signs. Remarkably, the deposits developed much more rapidly and extensively in fish inoculated with BSE-infected material than in the ones challenged with the scrapie-infected brain homogenate, with numerous deposits being proteinase K-resistant. These plaque-like aggregates exhibited congophilia and birefringence in polarized light, consistent with an amyloid-like component. The neurodegeneration and abnormal deposition in the brains of fish challenged with prion, especially BSE, raises concerns about the potential risk to public health. As fish aquaculture is an economically important industry providing high protein nutrition for humans and other mammalian species, the prospect of farmed fish being contaminated with infectious mammalian PrP^Sc, or of a prion disease developing in farmed fish is alarming and requires further evaluation.     

Novel Aspects of Prions,Their Receptor Molecules,and Innovative Approaches for TSE Therapy

1. Prion diseases are a group of rare, fatal neurodegenerative diseases, also known as transmissible spongiform encephalopathies (TSEs), that affect both animals and humans and include bovine spongiform encephalopathy (BSE) in cattle, scrapie in sheep, chronic wasting disease (CWD) in deer and elk, and Creutzfeldt–Jakob disease (CJD) in humans. TSEs are usually rapidly progressive and clinical symptoms comprise dementia and loss of movement coordination due to the accumulation of an abnormal isoform (PrP^Sc) of the host-encoded prion protein (PrP^c). 2. This article reviews the current knowledge on PrP^c and PrP^Sc, prion replication mechanisms, interaction partners of prions, and their cell surface receptors. Several strategies, summarized in this article, have been investigated for an effective antiprion treatment including development of a vaccination therapy and screening for potent chemical compounds. Currently, no effective treatment for prion diseases is available. 3. The identification of the 37 kDa/67 kDa laminin receptor (LRP/LR) and heparan sulfate as cell surface receptors for prions, however, opens new avenues for the development of alternative TSE therapies.     

Comparative profiling of highly enriched 22L and Chandler mouse scrapie prion protein preparations

Transmissible spongiform encephalopathies (TSEs) or prion diseases are characterized by the accumulation of an aggregated isoform of the prion protein (PrP). This pathological isoform, termed PrP^Sc, appears to be the primary component of the TSE infectious agent or prion. However, it is not clear to what extent other protein cofactors may be involved in TSE pathogenesis or whether there are PrP^Sc‐associated proteins which help to determine TSE strain‐specific disease phenotypes. We enriched PrP^Sc from the brains of mice infected with either 22L or Chandler TSE strains and examined the protein content of these samples using nanospray LC‐MS/MS. These samples were compared with “mock” PrP^Sc preparations from uninfected brains. PrP was the major component of the infected samples and ferritin was the most abundant impurity. Mock enrichments contained no detectable PrP but did contain a significant amount of ferritin. Of the total proteins identified, 32% were found in both mock and infected samples. The similarities between PrP^Sc samples from 22L and Chandler TSE strains suggest that the non‐PrP^Sc protein components found in standard enrichment protocols are not strain specific.     

Post‐translational changes to PrP alter transmissible spongiform encephalopathy strain properties

The agents responsible for transmissible spongiform encephalopathies (TSEs), or prion diseases, contain as a major component PrP^Sc, an abnormal conformer of the host glycoprotein PrP^C. TSE agents are distinguished by differences in phenotypic properties in the host, which nevertheless can contain PrP^Sc with the same amino‐acid sequence. If PrP alone carries information defining strain properties, these must be encoded by post‐translational events. Here we investigated whether the glycosylation status of host PrP affects TSE strain characteristics. We inoculated wild‐type mice with three TSE strains passaged through transgenic mice with PrP devoid of glycans at the first, second or both N‐glycosylation sites. We compared the infectious properties of the emerging isolates with TSE strains passaged in wild‐type mice by in vivo strain typing and by the standard scrapie cell assay in vitro. Strain‐specific characteristics of the 79A TSE strain changed when PrP^Sc was devoid of one or both glycans. Thus infectious properties of a TSE strain can be altered by post‐translational changes to PrP which we propose result in the selection of mutant TSE strains.     

Widespread PrPSc accumulation in muscles of hamsters orally infected with scrapie

Scrapie, bovine spongiform encephalopathy and chronic wasting disease are orally communicable, transmissible spongiform encephalopathies (TSEs). As zoonotic transmissions of TSE agents may pose a risk to human health, the identification of reservoirs for infectivity in animal tissues and their exclusion from human consumption has become a matter of great importance for consumer protection. In this study, a variety of muscles from hamsters that were orally challenged with scrapie was screened for the presence of a molecular marker for TSE infection, PrP^Sc (the pathological isoform of the prion protein PrP). Sensitive western blotting revealed consistent PrP^Sc accumulation in skeletal muscles from forelimb and hindlimb, head, back and shoulder, and in tongue. Previously, our animal model has provided substantial baseline information about the peripheral routing of infection in naturally occurring and orally acquired ruminant TSEs. Therefore, the findings described here highlight further the necessity to investigate thoroughly whether muscles of TSE-infected sheep, cattle, elk and deer contain infectious agents.     

Transgenic Rabbits Expressing Ovine PrP Are Susceptible to Scrapie

Transmissible spongiform encephalopathies (TSEs) are a group of neurodegenerative diseases affecting a wide range of mammalian species. They are caused by prions, a proteinaceous pathogen essentially composed of PrP^Sc, an abnormal isoform of the host encoded cellular prion protein PrPC. Constrained steric interactions between PrP^Sc and PrP^C are thought to provide prions with species specificity, and to control cross-species transmission into other host populations, including humans. Transgenetic expression of foreign PrP genes has been successfully and widely used to overcome the recognized resistance of mouse to foreign TSE sources. Rabbit is one of the species that exhibit a pronounced resistance to TSEs. Most attempts to infect experimentally rabbit have failed, except after inoculation with cell-free generated rabbit prions. To gain insights on the molecular determinants of the relative resistance of rabbits to prions, we generated transgenic rabbits expressing the susceptible V¹³⁶R¹⁵⁴Q¹⁷¹ allele of the ovine PRNP gene on a rabbit wild type PRNP New Zealand background and assessed their experimental susceptibility to scrapie prions. All transgenic animals developed a typical TSE 6–8 months after intracerebral inoculation, whereas wild type rabbits remained healthy more than 700 days after inoculation. Despite the endogenous presence of rabbit PrP^C, only ovine PrP^Sc was detectable in the brains of diseased animals. Collectively these data indicate that the low susceptibility of rabbits to prion infection is not enciphered within their non-PrP genetic background.     

Changes in Retinal Function and Morphology Are Early Clinical Signs of Disease in Cattle with Bovine Spongiform Encephalopathy

Bovine spongiform encephalopathy (BSE) belongs to a group of fatal, transmissible protein misfolding diseases known as transmissible spongiform encephalopathies (TSEs). All TSEs are caused by accumulation of misfolded prion protein (PrP^Sc) throughout the central nervous system (CNS), which results in neuronal loss and ultimately death. Like other protein misfolding diseases including Parkinson’s disease and Alzheimer’s disease, TSEs are generally not diagnosed until the onset of disease after the appearance of unequivocal clinical signs. As such, identification of the earliest clinical signs of disease may facilitate diagnosis. The retina is the most accessible part of the central nervous system, and retinal pathology in TSE affected animals has been previously reported. Here we describe antemortem changes in retinal function and morphology that are detectable in BSE inoculated animals several months (up to 11 months) prior to the appearance of any other signs of clinical disease. We also demonstrate that differences in the severity of these clinical signs reflect the amount of PrP^Sc accumulation in the retina and the resulting inflammatory response of the tissue. These results are the earliest reported clinical signs associated with TSE infection and provide a basis for understanding the pathology and evaluating therapeutic interventions.     

Species and Strain Glycosylation Patterns of PrPSc

Background

A key event in transmissible spongiform encephalopathies (TSEs) is the conversion of the soluble, protease-sensitive glycosylated prion protein (PrP^C) to an abnormally structured, aggregated and partially protease-resistant isoform (PrP^Sc). Both PrP isoforms bear two potential glycosylation sites and thus in a typical western blot with an anti-PrP antibody three distinct bands appear, corresponding to the di-, mono- or unglycosylated forms of the protein. The relative intensity and electrophoretic mobility of the three bands are characteristic of each TSE strain and have been used to discriminate between them.

Methodology/Principal Findings

In the present study we used lectin-based western blotting to evaluate possible variations in composition within sugar chains carried by PrP^Sc purified from subjects affected with different TSEs. Our findings indicate that in addition to the already well-documented differences in electrophoretic mobility and amounts of the glycosylated PrP^Sc forms, TSE strains also vary in the abundance of specific N-linked sugars of the PrP^Sc protein.

Conclusions/Significance

These results imply that PrP glycosylation might fine-tune the conversion of PrP^C to PrP^Sc and could play an accessory role in the appearance of some of the characteristic features of TSE strains. The differences in sugar composition could also be used as an additional tool for discrimination between the various TSEs.     

Evidence of scrapie transmission via milk

Background

The risk of scrapie infection increases with increased duration and proximity of contact between sheep at lambing. Scrapie infectivity has not been detected in milk but cellular prion protein, the precursor of disease-associated prion protein PrP^d, has been found in milk from ruminants. To determine whether milk is able to transmit scrapie, 18 lambs with a prion protein genotype associated with high susceptibility to scrapie (VRQ/VRQ) were fed milk from twelve scrapie-affected ewes of the same genotype, and 15 VRQ/VRQ sheep reared on scrapie-free dams served as controls.

Results

Three lambs fed milk from scrapie-affected ewes were culled due to intercurrent diseases at 43, 44 and 105 days of age respectively, and PrP^d was detected in the distal ileum of the first two lambs, whilst PrP^d was not found in lymphoreticular tissues in the third lamb. A control lamb, housed in a separate pen and culled at 38 days of age, was also negative for PrP^d in a range of tissues. Samples of recto-anal mucosa associated lymphoid tissue collected from the remaining 15 live lambs at seven months of age (between five to seven months after mixing) were positive for PrP^d in the scrapie milk recipients, whereas PrP^d was not detected in the remaining 14 controls at that time. A subsequent sample collected from control lambs revealed PrP^d accumulation in two of five lambs eight months after mixing with scrapie milk recipients suggestive of an early stage of infection via lateral transmission. By contrast, the control sheep housed in the same building but not mixed with the scrapie milk recipients were still negative for PrP^d.

Conclusion

The presence of PrP^d in distal ileum and rectal mucosa indicates transmission of scrapie from ewe to lamb via milk (or colostrum) although it is not yet clear if such cases would go on to develop clinical disease. The high level of infection in scrapie-milk recipients revealed by rectal mucosal testing at approximately seven months of age may be enhanced or supplemented by intra-recipient infection as these lambs were mixed together after feeding with milk from scrapie-affected ewes and we also observed lateral transmission from these animals to lambs weaned from scrapie-free ewes.

     

Prions in Milk from Ewes Incubating Natural Scrapie

Since prion infectivity had never been reported in milk, dairy products originating from transmissible spongiform encephalopathy (TSE)-affected ruminant flocks currently enter unrestricted into the animal and human food chain. However, a recently published study brought the first evidence of the presence of prions in mammary secretions from scrapie-affected ewes. Here we report the detection of consistent levels of infectivity in colostrum and milk from sheep incubating natural scrapie, several months prior to clinical onset. Additionally, abnormal PrP was detected, by immunohistochemistry and PET blot, in lacteal ducts and mammary acini. This PrP^Sc accumulation was detected only in ewes harbouring mammary ectopic lymphoid follicles that developed consequent to Maedi lentivirus infection. However, bioassay revealed that prion infectivity was present in milk and colostrum, not only from ewes with such lympho-proliferative chronic mastitis, but also from those displaying lesion-free mammary glands. In milk and colostrum, infectivity could be recovered in the cellular, cream, and casein-whey fractions. In our samples, using a Tg 338 mouse model, the highest per ml infectious titre measured was found to be equivalent to that contained in 6 µg of a posterior brain stem from a terminally scrapie-affected ewe. These findings indicate that both colostrum and milk from small ruminants incubating TSE could contribute to the animal TSE transmission process, either directly or through the presence of milk-derived material in animal feedstuffs. It also raises some concern with regard to the risk to humans of TSE exposure associated with milk products from ovine and other TSE-susceptible dairy species.     

Temporary depletion of complement component C3 or genetic deficiency of C1q significantly delays onset of scrapie

Following peripheral exposure to transmissible spongiform encephalopathies (TSEs), infectivity usually accumulates in lymphoid tissues before neuroinvasion. The host prion protein (PrPc) is critical for TSE agent replication and accumulates as an abnormal, detergent insoluble, relatively proteinase-resistant isoform (PrPSc) in diseased tissues. Early PrPSc accumulation takes place on follicular dendritic cells (FDCs) within germinal centers in lymphoid tissues of patients with variant Creutzfeldt-Jakob disease (vCJD), sheep with natural scrapie or rodents following experimental peripheral infection with scrapie. In mouse scrapie models, the absence of FDCs blocks scrapie replication and PrPSc accumulation in the spleen, and neuroinvasion is significantly impaired. The mechanisms by which the TSE agent initially localizes to lymphoid follicles and interacts with FDCs are unknown. Antigens are trapped and retained on the surface of FDCs through interactions between complement and cellular complement receptors. Here we show that in mice, both temporary depletion of complement component C3 or genetic deficiency of C1q significantly delays the onset of disease following peripheral infection, and reduces the early accumulation of PrPSc in the spleen. Thus, in the early stages of infection, C3 and perhaps C1q contribute to the localization of TSE infectivity in lymphoid tissue and may be therapeutic targets.     

Sequestration of free cholesterol in cell membranes by prions correlates with cytoplasmic phospholipase A<Subscript>2</Subscript>activation

Background  

The transmissible spongiform encephalopathies (TSEs), otherwise known as the prion diseases, occur following the conversion of the normal cellular prion protein (PrP^C) to an alternatively folded isoform (PrP^Sc). The accumulation of PrP^Sc within the brain leads to neurodegeneration through an unidentified mechanism. Since many neurodegenerative disorders including prion, Parkinson's and Alzheimer's diseases may be modified by cholesterol synthesis inhibitors, the effects of prion infection on the cholesterol balance within neuronal cells were examined.     

Prion Seeding Activities of Mouse Scrapie Strains with Divergent PrPSc Protease Sensitivities and Amyloid Plaque Content Using RT-QuIC and eQuIC

Different transmissible spongiform encephalopathy (TSE)-associated forms of prion protein (e.g. PrP^Sc) can vary markedly in ultrastructure and biochemical characteristics, but each is propagated in the host. PrP^Sc propagation involves conversion from its normal isoform, PrP^C, by a seeded or templated polymerization mechanism. Such a mechanism is also the basis of the RT-QuIC and eQuIC prion assays which use recombinant PrP (rPrP^Sen) as a substrate. These ultrasensitive detection assays have been developed for TSE prions of several host species and sample tissues, but not for murine models which are central to TSE pathogenesis research. Here we have adapted RT-QuIC and eQuIC to various murine prions and evaluated how seeding activity depends on glycophosphatidylinositol (GPI) anchoring and the abundance of amyloid plaques and protease-resistant PrP^Sc (PrP^Res). Scrapie brain dilutions up to 10⁻⁸ and 10⁻¹³ were detected by RT-QuIC and eQuIC, respectively. Comparisons of scrapie-affected wild-type mice and transgenic mice expressing GPI anchorless PrP showed that, although similar concentrations of seeding activity accumulated in brain, the heavily amyloid-laden anchorless mouse tissue seeded more rapid reactions. Next we compared seeding activities in the brains of mice with similar infectivity titers, but widely divergent PrP^Res levels. For this purpose we compared the 263K and 139A scrapie strains in transgenic mice expressing P101L PrP^C. Although the brains of 263K-affected mice had little immunoblot-detectable PrP^Res, RT-QuIC indicated that seeding activity was comparable to that associated with a high-PrP^Res strain, 139A. Thus, in this comparison, RT-QuIC seeding activity correlated more closely with infectivity than with PrP^Res levels. We also found that eQuIC, which incorporates a PrP^Sc immunoprecipitation step, detected seeding activity in plasma from wild-type and anchorless PrP transgenic mice inoculated with 22L, 79A and/or RML scrapie strains. Overall, we conclude that these new mouse-adapted prion seeding assays detect diverse types of PrP^Sc.     

Horse Prion Protein NMR Structure and Comparisons with Related Variants of the Mouse Prion Protein

The NMR structure of the horse (Equus caballus) cellular prion protein at 25 °C exhibits the typical PrP^C [cellular form of prion protein (PrP)] global architecture, but in contrast to most other mammalian PrP^Cs, it contains a well-structured loop connecting the β2 strand with the α2 helix. Comparison with designed variants of the mouse prion protein resulted in the identification of a single amino acid exchange within the loop, D167S, which correlates with the high structural order of this loop in the solution structure at 25 °C and is unique to the PrP sequences of equine species. The β2-α2 loop and the α3 helix form a protein surface epitope that has been proposed to be the recognition area for a hypothetical chaperone, “protein X,” which would promote conversion of PrP^C into the disease-related scrapie form and thus mediate intermolecular interactions related to the transmission barrier for transmissible spongiform encephalopathies (TSEs) between different species. The present results are evaluated in light of recent indications from in vivo experiments that the local β2-α2 loop structure affects the susceptibility of transgenic mice to TSEs and the fact that there are no reports on TSE in horses.     

Proteomic profiling of PrP27‐30‐enriched preparations extracted from the brain of hamsters with experimental scrapie

Transmissible spongiform encephalopathies (TSEs) are neurodegenerative disorders characterized by the accumulation in the CNS of a pathological conformer (PrP^TSE) of the host‐encoded cellular prion protein (PrP^C). PrP^TSE has a central role in the pathogenesis of the disease but other factors are likely involved in the pathological process. In this work we employed a multi‐step proteomic approach for the identification of proteins that co‐purify with the protease‐resistant core of PrP^TSE (PrP27‐30) extracted from brains of hamsters with experimental scrapie. We identified ferritin, calcium/calmodulin‐dependent protein kinase α type II, apolipoprotein E, and tubulin as the major components associated with PrP27‐30 but also trace amounts of actin, cofilin, Hsp90α, the γ subunit of the T‐complex protein 1, glyceraldehyde 3‐phosphate dehydrogenase, histones, and keratins. Whereas some of these proteins (tubulin and ferritin) are known to bind PrP, other proteins (calcium/calmodulin‐dependent protein kinase α type II, Hsp90α) may associate with PrP^TSE fibrils during disease. Apolipoprotein E and actin have been previously observed in association with PrP^TSE, whereas cofilin and actin were shown to form abnormal rods in the brain of patients with Alzheimer disease. The roles of these proteins in the development of brain lesions are still unclear and further work is needed to explain their involvement in the pathogenesis of TSEs.     

Anti-Prion Activity of a Panel of Aromatic Chemical Compounds: In Vitro and In Silico Approaches

The prion protein (PrP) is implicated in the Transmissible Spongiform Encephalopathies (TSEs), which comprise a group of fatal neurodegenerative diseases affecting humans and other mammals. Conversion of cellular PrP (PrP^C) into the scrapie form (PrP^Sc) is the hallmark of TSEs. Once formed, PrP^Sc aggregates and catalyzes PrP^C misfolding into new PrP^Sc molecules. Although many compounds have been shown to inhibit the conversion process, so far there is no effective therapy for TSEs. Besides, most of the previously evaluated compounds failed in vivo due to poor pharmacokinetic profiles. In this work we propose a combined in vitro/in silico approach to screen for active anti-prion compounds presenting acceptable drugability and pharmacokinetic parameters. A diverse panel of aromatic compounds was screened in neuroblastoma cells persistently infected with PrP^Sc (ScN2a) for their ability to inhibit PK-resistant PrP (PrP^Res) accumulation. From ∼200 compounds, 47 were effective in decreasing the accumulation of PrP^Res in ScN2a cells. Pharmacokinetic and physicochemical properties were predicted in silico, allowing us to obtain estimates of relative blood brain barrier permeation and mutagenicity. MTT reduction assays showed that most of the active compounds were non cytotoxic. Compounds that cleared PrP^Res from ScN2a cells, were non-toxic in the MTT assay, and presented a good pharmacokinetic profile were investigated for their ability to inhibit aggregation of an amyloidogenic PrP peptide fragment (PrP^109–149). Molecular docking results provided structural models and binding affinities for the interaction between PrP and the most promising compounds. In summary, using this combined in vitro/in silico approach we have identified new small organic anti-scrapie compounds that decrease the accumulation of PrP^Res in ScN2a cells, inhibit the aggregation of a PrP peptide, and possess pharmacokinetic characteristics that support their drugability. These compounds are attractive candidates for prion disease therapy.     

Peroxiredoxin 6 promotes upregulation of the prion protein (PrP) in neuronal cells of prion-infected mice

Background

It has been widely established that the conversion of the cellular prion protein (PrP^C) into its abnormal isoform (PrP^Sc) is responsible for the development of transmissible spongiform encephalopathies (TSEs). However, the knowledge of the detailed molecular mechanisms and direct functional consequences within the cell is rare. In this study, we aimed at the identification of deregulated proteins which might be involved in prion pathogenesis.

Findings

Apolipoprotein E and peroxiredoxin 6 (PRDX6) were identified as upregulated proteins in brains of scrapie-infected mice and cultured neuronal cell lines. Downregulation of PrP gene expression using specific siRNA did not result in a decrease of PRDX6 amounts. Interestingly, selective siRNA targeting PRDX6 or overexpression of PRDX6 controlled PrP^C and PrP^Sc protein amounts in neuronal cells.

Conclusions

Besides its possible function as a novel marker protein in the diagnosis of TSEs, PDRX6 represents an attractive target molecule in putative pharmacological intervention strategies in the future.     

RuNAway Disease: A two cycle model for transmissible spongiform encephalopathies (TSEs) wherein SINE proliferation drives PrP overproduction

Background

Despite decades of research, the agent responsible for transmitting spongiform encephalopathies (TSEs) has not been identified. The Prion hypothesis, which dominates the field, supposes that modified host PrP protein, termed PrP^Sc, acts as the transmissible agent. This model fits the observation that TSE diseases elicit almost no immune reaction. Prion transmission has not been verified, however, as it has not been possible to produce pure PrP^Sc aggregates. One long-standing objection to the Prion model is the observation that TSE disease agents show classical genetic behaviours, such as reproducible strain variation, while also responding to selection for novel traits such as adaptation to new hosts. Moreover, evidence has been steadily accumulating that infectious titre is decoupled from the quantity (or even the presence) of PrP^Sc deposits. Rather awkwardly for the Prion hypothesis, PrP^0/0 knockout mice have been found to incubate and transmit TSE agents (despite themselves being refractory to TSE disease).

Hypothesis

In this article, a new scheme, RuNAway, is proposed whereby uncontrolled proliferation of a type of parasitic gene, the small dispersed repeat sequences (SINEs), in somatic cells induces overproduction of PrP with pathogenic consequences. The RuNAway scheme involves twin tandem positive feedback loops: triggering the second loop leads to the pathogenic disease. This model is consistent with the long latency period and much shorter visible disease progression typical of TSEs.