Pα-syn* mitotoxicity is linked to MAPK activation and involves tau phosphorylation and aggregation at the mitochondria.

We recently identified a truncated and phosphorylated form of α-synuclein, pα-syn*, as a key neurotoxic α-synuclein species found in cultured neurons, as well as in mouse and Parkinson's disease patients' brains. Small pα-syn* aggregates localize to mitochondria and induce mitochondrial damage and fragmentation. Herein, we investigated the molecular basis of pα-syn*-induced toxicity.

Identification of a highly neurotoxic α-synuclein species inducing mitochondrial damage and mitophagy in Parkinson's disease.

Exposure of cultured primary neurons to preformed α-synuclein fibrils (PFFs) leads to the recruitment of endogenous α-synuclein and its templated conversion into fibrillar phosphorylated α-synuclein (pα-synF) aggregates resembling those involved in Parkinson's disease (PD) pathogenesis. Pα-synF was described previously as inclusions morphologically similar to Lewy bodies and Lewy neurites in PD patients. We discovered the existence of a conformationally distinct, nonfibrillar, phosphorylated α-syn species that we named "pα-syn*.

Neuronal death induced by misfolded prion protein is due to NAD+ depletion and can be relieved in vitro and in vivo by NAD+ replenishment.

The mechanisms of neuronal death in protein misfolding neurodegenerative diseases such as Alzheimer's, Parkinson's and prion diseases are poorly understood. We used a highly toxic misfolded prion protein (TPrP) model to understand neurotoxicity induced by prion protein misfolding. We show that abnormal autophagy activation and neuronal demise is due to severe, neuron-specific, nicotinamide adenine dinucleotide (NAD(+)) depletion.

Unfolded protein response-induced ERdj3 secretion links ER stress to extracellular proteostasis.

The Unfolded Protein Response (UPR) indirectly regulates extracellular proteostasis through transcriptional remodeling of endoplasmic reticulum (ER) proteostasis pathways. This remodeling attenuates secretion of misfolded, aggregation-prone proteins during ER stress. Through these activities, the UPR has a critical role in preventing the extracellular protein aggregation associated with numerous human diseases.

Unique drug screening approach for prion diseases identifies tacrolimus and astemizole as antiprion agents.

Prion diseases such as Creutzfeldt-Jakob disease (CJD) are incurable and rapidly fatal neurodegenerative diseases. Because prion protein (PrP) is necessary for prion replication but dispensable for the host, we developed the PrP-FRET-enabled high throughput assay (PrP-FEHTA) to screen for compounds that decrease PrP expression. We screened a collection of drugs approved for human use and identified astemizole and tacrolimus, which reduced cell-surface PrP and inhibited prion replication in neuroblastoma cells.

Strain-specific role of RNAs in prion replication.

Several lines of evidence suggest that various cofactors may be required for prion replication. PrP binds to polyanions, and RNAs were shown to promote the conversion of PrP(C) into PrP(Sc) in vitro. In the present study, we investigated strain-specific differences in RNA requirement during in vitro conversion and the potential role of RNA as a strain-specifying component of infectious prions.

Highly neurotoxic monomeric α-helical prion protein.

Prion diseases are infectious and belong to the group of protein misfolding neurodegenerative diseases. In these diseases, neuronal dysfunction and death are caused by the neuronal toxicity of a particular misfolded form of their cognate protein. The ability to specifically target the toxic protein conformer or the neuronal death pathway would provide powerful therapeutic approaches to these diseases.

Prion strain discrimination based on rapid in vivo amplification and analysis by the cell panel assay.

Prion strain identification has been hitherto achieved using time-consuming incubation time determinations in one or more mouse lines and elaborate neuropathological assessment. In the present work, we make a detailed study of the properties of PrP-overproducing Tga20 mice. We show that in these mice the four prion strains examined are rapidly and faithfully amplified and can subsequently be discriminated by a cell-based procedure, the Cell Panel Assay.

Atypical BSE (BASE) transmitted from asymptomatic aging cattle to a primate.

Background: Human variant Creutzfeldt-Jakob Disease (vCJD) results from foodborne transmission of prions from slaughtered cattle with classical Bovine Spongiform Encephalopathy (cBSE). Atypical forms of BSE, which remain mostly asymptomatic in aging cattle, were recently identified at slaughterhouses throughout Europe and North America, raising a question about human susceptibility to these new prion strains.

Methodology/principal Findings: Brain homogenates from cattle with classical BSE and atypical (BASE) infections were inoculated intracerebrally into cynomolgus monkeys (Macacca fascicularis), a non-human primate model previously demonstrated to be susceptible to the original strain of cBSE.

In vitro and in vivo neurotoxicity of prion protein oligomers.

The mechanisms underlying prion-linked neurodegeneration remain to be elucidated, despite several recent advances in this field. Herein, we show that soluble, low molecular weight oligomers of the full-length prion protein (PrP), which possess characteristics of PrP to PrPsc conversion intermediates such as partial protease resistance, are neurotoxic in vitro on primary cultures of neurons and in vivo after subcortical stereotaxic injection. Monomeric PrP was not toxic.

Toxic effects of intracerebral PrP antibody administration during the course of BSE infection in mice.

The absence of specific immune response is a hallmark of prion diseases. However, in vitro and in vivo experiments have provided evidence that an anti-PrP humoral response could have beneficial effects. Prophylactic passive immunization performed at the time of infection delayed or prevented disease.

The 37-kDa/67-kDa laminin receptor acts as a receptor for infectious prions and is inhibited by polysulfated glycanes.

Background: Recently, we showed that the 37-kDa/67-kDa laminin receptor (LRP/LR) acts as the receptor of the cellular prion protein.

Methods: For the present study, we investigated the binding of the murine scrapie prion protein (moPrP27-30) to baby hamster kidney (BHK) cells, using the Semliki Forest virus system.

Results: The enhanced binding of moPrP27-30 to BHK cells expressing moLRP::FLAG was inhibited by the LRP/LR-specific antibody W3, which suggests that LRP/LR acts as a receptor for the scrapie form of the prion protein, PrP(Sc).

PrPTSE distribution in a primate model of variant, sporadic, and iatrogenic Creutzfeldt-Jakob disease.

Human prion diseases, such as Creutzfeldt-Jakob disease (CJD), are neurodegenerative and fatal. Sporadic CJD (sCJD) can be transmitted between humans through medical procedures involving highly infected organs, such as the central nervous system. However, in variant CJD (vCJD), which is due to human contamination with the bovine spongiform encephalopathy (BSE) agent, lymphoreticular tissue also harbors the transmissible spongiform encephalopathy-associated prion protein (PrP(TSE)), which poses a particularly acute risk for iatrogenic transmission.

The metabolism of glycosaminoglycans is impaired in prion diseases.

It is well established that the conversion of PrP(C) to PrP(Sc) is the key event in prion disease biology. In addition, several lines of evidence suggest that glycosaminoglycans (GAGs) and in particular heparan sulfate (HS) may play a role in the PrP(C) to PrP(Sc) conversion process. It has been proposed that PrP(Sc) accumulation in prion diseases may induce aberrant activation of lysosomal activity, which has been shown to result in neurodegeneration in a number of diseases, especially lysosomal storage disorders.

Risk of oral infection with bovine spongiform encephalopathy agent in primates.

The uncertain extent of human exposure to bovine spongiform encephalopathy (BSE)--which can lead to variant Creutzfeldt-Jakob disease (vCJD)--is compounded by incomplete knowledge about the efficiency of oral infection and the magnitude of any bovine-to-human biological barrier to transmission. We therefore investigated oral transmission of BSE to non-human primates. We gave two macaques a 5 g oral dose of brain homogenate from a BSE-infected cow.

Novel methods for disinfection of prion-contaminated medical devices.

Background: The unique resistance of prions to classic methods of decontamination, and evidence that prion diseases can be transmitted iatrogenically by medical devices pose a serious infection control challenge to health-care facilities. In view of the widespread tissue distribution of the variant Creutzfeldt-Jakob disease agent in human beings, new practicable decontamination procedures are urgently needed.

Methods: We adapted an in-vivo method using stainless steel wires contaminated with prions to the hamster-adapted scrapie strain 263K.

Knock-down of the 37-kDa/67-kDa laminin receptor in mouse brain by transgenic expression of specific antisense LRP RNA.

The 37-kDa/67-kDa laminin receptor (LRP/LR) plays a major role in the propagation of PrPSc, the abnormal form of the prion protein. In order to ablate the expression of LRP/LR in mouse brain we generated transgenic mice ectopically expressing antisense LRP RNA in the brain under control of the neuron-specific enolase (NSE) promoter. Hemizygous transgenic mice TgN(NSEasLRP)2 showed a significant reduction of LRP/LR protein levels in hippocampal and cerebellar brain regions.

Putative functions of PrP(C).

While the exact function of the cellular prion protein (PrP(C)) remains unknown, there are several leads due to increasing knowledge on the localisation and interaction of PrP(C) with other molecules. This chapter will concentrate on these aspects. Identified ligands of PrP(C) mainly belong to the categories of heat-shock proteins, membrane-bound receptors, or heparan sulphates.

A novel generation of heparan sulfate mimetics for the treatment of prion diseases.

The accumulation of PrP(res), the protease-resistant abnormal form of the host-encoded cellular prion protein, PrP(C), plays a central role in transmissible spongiform encephalopathies. Human contamination by bovine spongiform encephalopathy (BSE) has propelled many scientific teams on a highway for anti-prion drug development. This study reports that heparan sulfate mimetics (HMs), developed originally for their effect on tissue regeneration, abolish prion propagation in scrapie-infected GT1 cells.

The 37 kDa/67 kDa laminin receptor is required for PrP(Sc) propagation in scrapie-infected neuronal cells.

The accumulation of PrP(Sc) in scrapie-infected neuronal cells has been prevented by three approaches: (i) transfection of ScMNB cells with an antisense laminin receptor precursor (LRP) RNA-expression plasmid, (ii) transfection of ScN2a cells and ScGT1 cells with small interfering RNAs (siRNAs) specific for the LRP mRNA, and (iii) incubation of ScN2a cells with an anti-LRP/LR antibody. LRP antisense RNA and LRP siRNAs reduced LRP/LR expression and inhibited the accumulation of PrP(Sc) in these cells. The treatments also reduced PrP(c) levels.