Early structural features in mammalian prion conformation conversion

The conversion to a disease-associated conformer (PrP (Sc) ) of the cellular prion protein (PrP (C) ) is the central event in prion diseases. Wild-type PrPC converts to PrP (Sc) in the sporadic forms of the disorders through an unknown mechanism. These forms account for up to 85% of all human (Hu) occurrences; the infectious types contribute for less than 1%, while genetic incidence of the disease is about 15%. Familial Hu prion diseases are associated with about forty point mutations of the gene coding for the PrP denominated PRNP. Most of the variants associated with these mutations are located in the globular domain of the protein. In a recent work in collaboration with the German Research School for Simulation Science, in Jülich, Germany, we performed molecular dynamics simulations for each of these mutants to investigate their structure in aqueous solution. Structural analysis of the various point mutations present in the globular domain unveiled common folding traits that may allow to a better understanding of the early conformational changes leading to the formation of monomeric PrP (Sc) . Recent experimental data support these findings, thus opening novel approaches to determine initial structural determinants of prion formation. © 2012 Landes Bioscience

Molecular pathogenesis of prion diseases

Introduction: Prion diseases or transmissible spongiform encephalopathies (TSEs) are rare, fatal and incurable neurodegenerative disorders of humans and animals (Prusiner, 1998). In humans, prion diseases occur with unique aetiology as sporadic, genetic or infectious disorders. Sporadic cases of prion diseases, which account for the majority of casualties (up to 85% of all cases), are of unknown origin; the genetic forms are less frequent (up to 15%), while the infectious cases are extremely rare with an incidence of less than 1% (Prusiner, 2001). Creutzfeldt-Jakob disease (CJD), Gerstmann-Str\ue4ussler-Scheinker (GSS) syndrome, Fatal Familial Insomnia (FFI) are examples of human prion diseases. In animals the disease is mostly infectious and the mode of transmission is horizontal. Prion diseases include scrapie in sheep and goats, bovine spongiform encephalopathy (BSE) in cattle, and chronic wasting disease of deer, elk, and moose (Williams, 2005). The agents responsible for prion diseases are infectious proteins named prions. The term \u2018prion\u2019 was coined when Stanley B. Prusiner introduced the concept of proteinaceous infectious particles (Prusiner, 1982). Since the introduction of this once heretical notion, mounting evidence has strengthened its validity. In the next sections of this chapter we present and discuss the peculiar complexity of the molecular pathogenesis of prion diseases in humans and animals

Prion protein and aging

The cellular prion protein (PrP(C)) has been widely investigated ever since its conformational isoform, the prion (or PrP(Sc)), was identified as the etiological agent of prion disorders. The high homology shared by the PrP(C)-encoding gene among mammals, its high turnover rate and expression in every tissue strongly suggest that PrP(C) may possess key physiological functions. Therefore, defining PrP(C) roles, properties and fate in the physiology of mammalian cells would be fundamental to understand its pathological involvement in prion diseases. Since the incidence of these neurodegenerative disorders is enhanced in aging, understanding PrP(C) functions in this life phase may be of crucial importance. Indeed, a large body of evidence suggests that PrP(C) plays a neuroprotective and antioxidant role. Moreover, it has been suggested that PrP(C) is involved in Alzheimer disease, another neurodegenerative pathology that develops predominantly in the aging population. In prion diseases, PrP(C) function is likely lost upon protein aggregation occurring in the course of the disease. Additionally, the aging process may alter PrP(C) biochemical properties, thus influencing its propensity to convert into PrP(Sc). Both phenomena may contribute to the disease development and progression. In Alzheimer disease, PrP(C) has a controversial role because its presence seems to mediate β-amyloid toxicity, while its down-regulation correlates with neuronal death. The role of PrP(C) in aging has been investigated from different perspectives, often leading to contrasting results. The putative protein functions in aging have been studied in relation to memory, behavior and myelin maintenance. In aging mice, PrP(C) changes in subcellular localization and post-translational modifications have been explored in an attempt to relate them to different protein roles and propensity to convert into PrP(Sc). Here we provide an overview of the most relevant studies attempting to delineate PrP(C) functions and fate in aging

Transcriptome analysis of prion disease animal models

Prion diseases are incurable and fatal neurodegenerative disorders that affect both humans and animals. The causative agent is an infectious protein called prion (PrPSc), which is the pathological form of a normal protein (PrPC) present on the cell membrane. The molecular mechanisms underlying prion replication and subsequent degeneration of the Central Nervous System (CNS) are still poorly understood and therefore innovative approaches are needed to build diagnostic, therapeutic, taxonomic, and disease surveillance tools. We are going to adopt an unbiased genomic approach and conduct whole transcriptome analyses using microarray gene expression methods in brain and/or blood of infected animals versus healthy controls. We hope to identify a set of genes that can be used for early diagnosis and/or as targets for therapeutic strategies. Within the Trans2Care project we intend to promote collaboration and exchange of knowledge to facilitate all partners’ research objectives, and possibly find a common way to accelerate the process aimed at improving our healthcare system

Aberrant ERK 1/2 complex activation and localization in scrapie-infected GT1-1 cells

<p>Abstract</p> <p>Background</p> <p>Fatal neurodegenerative disorders such as Creutzfeldt-Jakob and Gerstmann-Sträussler-Scheinker diseases in humans, scrapie and bovine spongiform encephalopathy in animals, are characterized by the accumulation in the brain of a pathological form of the prion protein (PrP) denominated PrP^Sc. The latter derives from the host cellular form, PrP^C, through a process whereby portions of its α-helical and coil structures are refolded into β-sheet structures.</p> <p>Results</p> <p>In this work, the widely known <it>in vitro </it>model of prion replication, hypothalamic GT1-1 cell line, was used to investigate cellular and molecular responses to prion infection. The MAP kinase cascade was dissected to assess the phosphorylation levels of src, MEK 1/2 and ERK 1/2 signaling molecules, both before and after prion infection. Our findings suggest that prion replication leads to a hyper-activation of this pathway. Biochemical analysis was complemented with immunofluorescence studies to map the localization of the ERK complex within the different cellular compartments. We showed how the ERK complex relocates in the cytosol upon prion infection. We correlated these findings with an impairment of cell growth in prion-infected GT1-1 cells as probed by MTT assay. Furthermore, given the persistent urgency in finding compounds able to cure prion infected cells, we tested the effects on the ERK cascade of two molecules known to block prion replication <it>in vitro</it>, quinacrine and Fab D18. We were able to show that while these two compounds possess similar effects in curing prion infection, they affect the MAP kinase cascade differently.</p> <p>Conclusions</p> <p>Taken together, our results help shed light on the molecular events involved in neurodegeneration and neuronal loss in prion infection and replication. In particular, the combination of chronic activation and aberrant localization of the ERK complex may lead to a lack of essential neuroprotective and survival factors. Interestingly, these data seem to define some common traits with other neurodegenerative disorders such as, for example, Alzheimer's disease.</p

Novel markers for neurodegeneration

Prion diseases are incurable and fatal neurodegenerative disorders that affect both humans and animals. The causative agent is an infectious protein called prion (PrPSc), which is the pathological form of a normal protein (PrPC) present on the cell membrane. The molecular mechanisms underlying prion replication and subsequent degeneration of the Central Nervous System (CNS) are still poorly understood and therefore innovative approaches are needed to build diagnostic, therapeutic, taxonomic, and disease surveillance tools. We adopted an unbiased genomic approach and conducted whole transcriptome analyses using microarray and RT-qPCR gene expression methods in brain of infected macaques versus healthy controls. We identified a set of genes that could become novel biomarkers for early diagnosis and/or therapeutic strategies for prion diseases and other neurodegenerative disorders

Structural Consequences of Copper Binding to the Prion Protein

Prion, or PrPSc, is the pathological isoform of the cellular prion protein (PrPC) and it is the etiological agent of transmissible spongiform encephalopathies (TSE) affecting humans and animal species. The most relevant function of PrPC is its ability to bind copper ions through its flexible N-terminal moiety. This review includes an overview of the structure and function of PrPC with a focus on its ability to bind copper ions. The state-of-the-art of the role of copper in both PrPC physiology and in prion pathogenesis is also discussed. Finally, we describe the structural consequences of copper binding to the PrPC structure

A system-level approach for deciphering the transcriptional response to prion infection

Motivation: Deciphering the response of a complex biological system to an insulting event, at the gene expression level, requires adopting theoretical models that are more sophisticated than a one-to-one comparison (i.e. t-test). Here, we investigate the ability of a novel reverse engineering approach (System Response Inference) to unveil non-obvious transcriptional signatures of the system response induced by prion infection. Results: To this end, we analyze previously published gene expression data, from which we extrapolate a putative full-scale model of transcriptional gene-gene dependencies in the mouse central nervous system. Then, we use this nominal model to interpret the gene expression changes caused by prion replication, aiming at selecting the genes primarily influenced by this perturbation. Our method sheds light on the mode of action of prions by identifying key transcripts that are the most likely to be responsible for the overall transcriptional rearrangement from a nominal regulatory network. As a first result of our inference, we have been able to predict known targets of prions (i.e. PrPC) and to unveil the potential role of previously unsuspected genes. Contact: [email protected] Supplementary Information: Supplementary data are available at Bioinformatics onlin

Probing Early Misfolding Events in Prion Protein Mutants by NMR Spectroscopy

The post-translational conversion of the ubiquitously expressed cellular form of the prion protein, PrPC, into its misfolded and pathogenic isoform, known as prion or PrPSc, plays a key role in prion diseases. These maladies are denoted transmissible spongiform encephalopathies (TSEs) and affect both humans and animals. A prerequisite for understanding TSEs is unraveling the molecular mechanism leading to the conversion process whereby most \u3b1-helical motifs are replaced by \u3b2-sheet secondary structures. Importantly, most point mutations linked to inherited prion diseases are clustered in the C-terminal domain region of PrPC and cause spontaneous conversion to PrPSc. Structural studies with PrP variants promise new clues regarding the proposed conversion mechanism and may help identify "hot spots" in PrPC involved in the pathogenic conversion. These investigations may also shed light on the early structural rearrangements occurring in some PrPC epitopes thought to be involved in modulating prion susceptibility. Here we present a detailed overview of our solution-state NMR studies on human prion protein carrying different pathological point mutations and the implications that such findings may have for the future of prion research

New insights into structural determinants of prion protein folding and stability

Prions are the etiological agent of fatal neurodegenerative diseases called prion diseases or transmissible spongiform encephalopathies. These maladies can be sporadic, genetic or infectious disorders. Prions are due to post-translational modifications of the cellular prion protein leading to the formation of a \u3b2-sheet enriched conformer with altered biochemical properties. The molecular events causing prion formation in sporadic prion diseases are still elusive. Recently, we published a research elucidating the contribution of major structural determinants and environmental factors in prion protein folding and stability. Our study highlighted the crucial role of octarepeats in stabilizing prion protein; the presence of a highly enthalpically stable intermediate state in prion-susceptible species; and the role of disulfide bridge in preserving native fold thus avoiding the misfolding to a \u3b2-sheet enriched isoform. Taking advantage from these findings, in this work we present new insights into structural determinants of prion protein folding and stability