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1

Comparative analysis of the prion protein gene sequences in African lion  

Microsoft Academic Search

The prion protein gene of African lion (Panthera Leo) was first cloned and polymorphisms screened. The results suggest that the prion protein gene of eight African lions is highly\\u000a homogenous. The amino acid sequences of the prion protein (PrP) of all samples tested were identical. Four single nucleotide\\u000a polymorphisms (C42T, C81A, C420T, T600C) in the prion protein gene (Prnp) of

Chang-De Wu; Wan-Yong Pang; De-Ming Zhao

2006-01-01

2

Comparative analysis of the prion protein gene sequences in African lion.  

PubMed

The prion protein gene of African lion (Panthera Leo) was first cloned and polymorphisms screened. The results suggest that the prion protein gene of eight African lions is highly homogenous. The amino acid sequences of the prion protein (PrP) of all samples tested were identical. Four single nucleotide polymorphisms (C42T, C81A, C420T, T600C) in the prion protein gene (Prnp) of African lion were found, but no amino acid substitutions. Sequence analysis showed that the higher homology is observed to felis catus AF003087 (96.7%) and to sheep number M31313.1 (96.2%) Genbank accessed. With respect to all the mammalian prion protein sequences compared, the African lion prion protein sequence has three amino acid substitutions. The homology might in turn affect the potential intermolecular interactions critical for cross species transmission of prion disease. PMID:16972036

Wu, Chang-De; Pang, Wan-Yong; Zhao, De-Ming

2006-10-01

3

Prions and prion-like proteins.  

PubMed

Prions are self-replicating protein aggregates and are the primary causative factor in a number of neurological diseases in mammals. The prion protein (PrP) undergoes a conformational transformation leading to aggregation into an infectious cellular pathogen. Prion-like protein spreading and transmission of aggregates between cells have also been demonstrated for other proteins associated with Alzheimer disease and Parkinson disease. This protein-only phenomenon may therefore have broader implications in neurodegenerative disorders. The minireviews in this thematic series highlight the recent advances in prion biology and the roles these unique proteins play in disease. PMID:24860092

Fraser, Paul E

2014-07-18

4

Prion protein structural features indicate possible relations to signal peptidases  

E-print Network

Hypothesis Prion protein structural features indicate possible relations to signal peptidases Rudi are believed to be caused by an oligomeric isoform, PrP , of the cellular prion protein, PrPg . One of the key prion protein; Signal peptidase; Three-dimensional structure; Sequence similarity 1. Introduction Prions

Wider, Gerhard

5

A Systematic Survey Identifies Prions and Illuminates Sequence Features of Prionogenic Proteins  

E-print Network

Prions are proteins that convert between structurally and functionally distinct states, one or more of which is transmissible. In yeast, this ability allows them to act as non-Mendelian elements of phenotypic inheritance. ...

Kapila, Atul

6

Prions, protein homeostasis, and phenotypic diversity  

E-print Network

Prions are fascinating but often misunderstood protein aggregation phenomena. The traditional association of the mammalian prion protein with disease has overshadowed a potentially more interesting attribute of prions: ...

Lindquist, Susan

7

Signal sequence insufficiency contributes to neurodegeneration caused by transmembrane prion protein  

PubMed Central

Protein translocation into the endoplasmic reticulum is mediated by signal sequences that vary widely in primary structure. In vitro studies suggest that such signal sequence variations may correspond to subtly different functional properties. Whether comparable functional differences exist in vivo and are of sufficient magnitude to impact organism physiology is unknown. Here, we investigate this issue by analyzing in transgenic mice the impact of signal sequence efficiency for mammalian prion protein (PrP). We find that replacement of the average efficiency signal sequence of PrP with more efficient signals rescues mice from neurodegeneration caused by otherwise pathogenic PrP mutants in a downstream hydrophobic domain (HD). This effect is explained by the demonstration that efficient signal sequence function precludes generation of a cytosolically exposed, disease-causing transmembrane form of PrP mediated by the HD mutants. Thus, signal sequences are functionally nonequivalent in vivo, with intrinsic inefficiency of the native PrP signal being required for pathogenesis of a subset of disease-causing PrP mutations. PMID:20156965

Rane, Neena S.; Chakrabarti, Oishee; Feigenbaum, Lionel

2010-01-01

8

Discriminant analysis of prion sequences for prediction of susceptibility  

PubMed Central

Prion diseases, including ovine scrapie, bovine spongiform encephalopathy (BSE), human kuru and Creutzfeldt–Jakob disease (CJD), originate from a conformational change of the normal cellular prion protein (PrPC) into abnormal protease-resistant prion protein (PrPSc). There is concern regarding these prion diseases because of the possibility of their zoonotic infections across species. Mutations and polymorphisms of prion sequences may influence prion-disease susceptibility through the modified expression and conformation of proteins. Rapid determination of susceptibility based on prion-sequence polymorphism information without complex structural and molecular biological analyses may be possible. Information regarding the effects of mutations and polymorphisms on prion-disease susceptibility was collected based on previous studies to classify the susceptibilities of sequences, whereas the BLOSUM62 scoring matrix and the position-specific scoring matrix were utilised to determine the distance of target sequences. The k-nearest neighbour analysis was validated with cross-validation methods. The results indicated that the number of polymorphisms did not influence prion-disease susceptibility, and three and four k-objects showed the best accuracy in identifying the susceptible group. Although sequences with negative polymorphisms showed relatively high accuracy for determination, polymorphisms may still not be an appropriate factor for estimating variation in susceptibility. Discriminant analysis of prion sequences with scoring matrices was attempted as a possible means of determining susceptibility to prion diseases. Further research is required to improve the utility of this method. PMID:24113272

Lee, Ji-Hae; Bae, Se-Eun; Jung, Sunghoon; Ahn, Insung; Son, Hyeon Seok

2013-01-01

9

Recombinant Human Prion Protein Inhibits Prion Propagation in vitro  

E-print Network

Recombinant Human Prion Protein Inhibits Prion Propagation in vitro Jue Yuan1,3 *, Yi-An Zhan1, Ohio, USA, 3 National Prion Disease Pathology Surveillance Center, Case Western Reserve University), Cairo, Egypt. Prion diseases are associated with the conformational conversion of the cellular prion

10

Prion protein and aging  

PubMed Central

The cellular prion protein (PrPC) has been widely investigated ever since its conformational isoform, the prion (or PrPSc), was identified as the etiological agent of prion disorders. The high homology shared by the PrPC-encoding gene among mammals, its high turnover rate and expression in every tissue strongly suggest that PrPC may possess key physiological functions. Therefore, defining PrPC 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 PrPC functions in this life phase may be of crucial importance. Indeed, a large body of evidence suggests that PrPC plays a neuroprotective and antioxidant role. Moreover, it has been suggested that PrPC is involved in Alzheimer disease, another neurodegenerative pathology that develops predominantly in the aging population. In prion diseases, PrPC function is likely lost upon protein aggregation occurring in the course of the disease. Additionally, the aging process may alter PrPC biochemical properties, thus influencing its propensity to convert into PrPSc. Both phenomena may contribute to the disease development and progression. In Alzheimer disease, PrPC has a controversial role because its presence seems to mediate ?-amyloid toxicity, while its down-regulation correlates with neuronal death. The role of PrPC 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, PrPC 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 PrPSc. Here we provide an overview of the most relevant studies attempting to delineate PrPC functions and fate in aging. PMID:25364751

Gasperini, Lisa; Legname, Giuseppe

2014-01-01

11

Metal Binding to Prion Protein  

Microsoft Academic Search

\\u000a The number of original research articles, review articles, book chapters, and monographs published annually on prion is staggering.\\u000a The experienced reader in this field might ask: “Do we really need another review on prion protein?” This chapter attempts\\u000a to be different from other recent reviews by focusing on the chemical aspects of metal binding of prion and some of its

R. P. Bonomo; D. Grasso; G. Grasso; V. Guantieri; G. Impellizzeri; C. Rosa; D. Milardi; G. Pappalardo; G. Tabbì; E. Rizzarelli

12

Prion Variants and Species Barriers Among Saccharomyces Ure2 Proteins  

Microsoft Academic Search

As hamster scrapie cannot infect mice, due to sequence differences in their PrP proteins, we find ''species barriers'' to transmission of the (URE3) prion in Saccharomyces cerevisiae among Ure2 proteins of S. cerevisiae, paradoxus, bayanus, cariocanus, and mikatae on the basis of differences among their Ure2p prion domain sequences. The rapid variation of the N-terminal Ure2p prion domains results in

Herman K. Edskes; Lindsay M. McCann; Andrea M. Hebert; Reed B. Wickner

2009-01-01

13

Ultrasensitive detection of scrapie prion protein using seeded conversion of recombinant prion protein  

E-print Network

Ultrasensitive detection of scrapie prion protein using seeded conversion of recombinant prion A Onwubiko1, Suzette A Priola1 & Byron Caughey1 The scrapie prion protein isoform, PrPSc, is a prion-associated marker that seeds the conformational conversion and polymerization of normal protease-sensitive prion

Cai, Long

14

Prion variants and species barriers among Saccharomyces Ure2 proteins.  

PubMed

As hamster scrapie cannot infect mice, due to sequence differences in their PrP proteins, we find "species barriers" to transmission of the [URE3] prion in Saccharomyces cerevisiae among Ure2 proteins of S. cerevisiae, paradoxus, bayanus, cariocanus, and mikatae on the basis of differences among their Ure2p prion domain sequences. The rapid variation of the N-terminal Ure2p prion domains results in protection against the detrimental effects of infection by a prion, just as the PrP residue 129 Met/Val polymorphism may have arisen to protect humans from the effects of cannibalism. Just as spread of bovine spongiform encephalopathy prion variant is less impaired by species barriers than is sheep scrapie, we find that some [URE3] prion variants are infectious to another yeast species while other variants (with the identical amino acid sequence) are not. The species barrier is thus prion variant dependent as in mammals. [URE3] prion variant characteristics are maintained even on passage through the Ure2p of another species. Ure2p of Saccharomyces castelli has an N-terminal Q/N-rich "prion domain" but does not form prions (in S. cerevisiae) and is not infected with [URE3] from Ure2p of other Saccharomyces. This implies that conservation of its prion domain is not for the purpose of forming prions. Indeed the Ure2p prion domain has been shown to be important, though not essential, for the nitrogen catabolism regulatory role of the protein. PMID:19124570

Edskes, Herman K; McCann, Lindsay M; Hebert, Andrea M; Wickner, Reed B

2009-03-01

15

Cellular prion protein neuroprotective function: implications in prion diseases  

Microsoft Academic Search

Prion protein can display two conformations: a normal cellular conformation (PrP) and a pathological conformation associated with prion diseases (PrP Sc). Three complementary strategies are used by researchers investigating how PrP is involved in the pathogenesis of prion diseases: elucidation of the normal function of PrP, determination of how PrP Sc is toxic to neurons, and unraveling the mechanism for

Xavier Roucou; Andréa C. LeBlanc

2005-01-01

16

PrPSc-like prion protein peptide inhibits the function of cellular prion protein.  

PubMed Central

Mice lacking expression of the prion protein are protected against infection with prion disease. Neurodegeneration in prion disease requires the formation of the abnormal isoform of the prion protein (PrP(Sc)) from host prion protein. Therefore expression of normal host prion protein is necessary for prion disease. In the present investigation, it was demonstrated that PrP(Sc) and a peptide resembling PrP(Sc), PrP106-126, both bind to cellular prion protein at amino acid residues 112-119. Interaction between PrP106-126 and the prion protein strips the prion protein from cells. Direct interaction of PrP106-126 with the prion protein was found to make cells more susceptible to copper toxicity, inhibited copper uptake into cells and inhibited the superoxide dismutase-like activity of the prion protein. Direct inhibition of prion protein function by PrP(Sc) may be necessary for neurodegeneration in prion disease. PMID:11085945

Brown, D R

2000-01-01

17

Healthy goats naturally devoid of prion protein.  

PubMed

Prion diseases such as scrapie in small ruminants, bovine spongiform encephalopathy (BSE) in cattle and Creutzfeldt-Jakob disease (CJD) in man, are fatal neurodegenerative disorders. These diseases result from the accumulation of misfolded conformers of the host-encoded prion protein (PrP) in the central nervous system. To date naturally-occurring PrP free animals have not been reported. Here we describe healthy non-transgenic animals, Norwegian Dairy Goats, lacking prion protein due to a nonsense mutation early in the gene. These animals are predicted to be resistant to prion disease and will be valuable for research and for production of prion-free products. PMID:23249298

Benestad, Sylvie L; Austbø, Lars; Tranulis, Michael A; Espenes, Arild; Olsaker, Ingrid

2012-01-01

18

Prion protein self-interaction in prion disease therapy approaches.  

PubMed

Transmissible spongiform encephalopathies (TSEs) or prion diseases are unique disorders that are not caused by infectious micro-organisms (bacteria or fungi), viruses or parasites, but rather seem to be the result of an infectious protein. TSEs are comprised of fatal neurodegenerative disorders affecting both human and animals. Prion diseases cause sponge-like degeneration of neuronal tissue and include (among others) Creutzfeldt-Jacob disease in humans, bovine spongiform encephalopathy (BSE) in cattle and scrapie in sheep. TSEs are characterized by the formation and accumulation of transmissible (infectious) disease-associated protease-resistant prion protein (PrP(Sc)), mainly in tissues of the central nervous system. The exact molecular processes behind the conversion of PrP(C) into PrP(Sc) are not clearly understood. Correlations between prion protein polymorphisms and disease have been found, however in what way these polymorphisms influence the conversion processes remains an enigma; is stabilization or destabilization of the prion protein the basis for a higher conversion propensity? Apart from the disease-associated polymorphisms of the prion protein, the molecular processes underlying conversion are not understood. There are some notions as to which regions of the prion protein are involved in refolding of PrP(C) into PrP(Sc) and where the most drastic structural changes take place. Direct interactions between PrP(C) molecules and/or PrP(Sc) are likely at the basis of conversion, however which specific amino acid domains are involved and to what extent these domains contribute to conversion resistance/sensitivity of the prion protein or the species barrier is still unknown. PMID:22029882

Rigter, Alan; Priem, Jan; Langeveld, Jan P M; Bossers, Alex

2011-09-01

19

Signal Transduction Through Prion Protein  

Microsoft Academic Search

The cellular prion protein PrPc is a glycosylphosphatidylinositol-anchored cell-surface protein whose biological function is unclear. We used the murine 1C11 neuronal differentiation model to search for PrPc-dependent signal transduction through antibody-mediated cross-linking. A caveolin-1-dependent coupling of PrPc to the tyrosine kinase Fyn was observed. Clathrin might also contribute to this coupling. The ability of the 1C11 cell line to trigger

S. Mouillet-Richard; M. Ermonval; C. Chebassier; J. L. Laplanche; S. Lehmann; J. M. Launay; O. Kellermann

2000-01-01

20

Amyloid diseases of yeast: prions are proteins acting as genes.  

PubMed

The unusual genetic properties of the non-chromosomal genetic elements [URE3] and [PSI+] led to them being identified as prions (infectious proteins) of Ure2p and Sup35p respectively. Ure2p and Sup35p, and now several other proteins, can form amyloid, a linear ordered polymer of protein monomers, with a part of each molecule, the prion domain, forming the core of this ?-sheet structure. Amyloid filaments passed to a new cell seed the conversion of the normal form of the protein into the same amyloid form. The cell's phenotype is affected, usually from the deficiency of the normal form of the protein. Solid-state NMR studies indicate that the yeast prion amyloids are in-register parallel ?-sheet structures, in which each residue (e.g. Asn35) forms a row along the filament long axis. The favourable interactions possible for aligned identical hydrophilic and hydrophobic residues are believed to be the mechanism for propagation of amyloid conformation. Thus, just as DNA mediates inheritance by templating its own sequence, these proteins act as genes by templating their conformation. Distinct isolates of a given prion have different biological properties, presumably determined by differences between the amyloid structures. Many lines of evidence indicate that the Saccharomyces cerevisiae prions are pathological disease agents, although the example of the [Het-s] prion of Podospora anserina shows that a prion can have beneficial aspects. PMID:25131596

Wickner, Reed B; Edskes, Herman K; Bateman, David A; Kelly, Amy C; Gorkovskiy, Anton; Dayani, Yaron; Zhou, Albert

2014-01-01

21

Discovering putative prion sequences in complete proteomes using probabilistic representations of Q/N-rich domains  

PubMed Central

Background Prion proteins conform a special class among amyloids due to their ability to transmit aggregative folds. Prions are known to act as infectious agents in neurodegenerative diseases in animals, or as key elements in transcription and translation processes in yeast. It has been suggested that prions contain specific sequential domains with distinctive amino acid composition and physicochemical properties that allow them to control the switch between soluble and ?-sheet aggregated states. Those prion-forming domains are low complexity segments enriched in glutamine/asparagine and depleted in charged residues and prolines. Different predictive methods have been developed to discover novel prions by either assessing the compositional bias of these stretches or estimating the propensity of protein sequences to form amyloid aggregates. However, the available algorithms hitherto lack a thorough statistical calibration against large sequence databases, which makes them unable to accurately predict prions without retrieving a large number of false positives. Results Here we present a computational strategy to predict putative prion-forming proteins in complete proteomes using probabilistic representations of prionogenic glutamine/asparagine rich regions. After benchmarking our predictive model against large sets of non-prionic sequences, we were able to filter out known prions with high precision and accuracy, generating prediction sets with few false positives. The algorithm was used to scan all the proteomes annotated in public databases for the presence of putative prion proteins. We analyzed the presence of putative prion proteins in all taxa, from viruses and archaea to plants and higher eukaryotes, and found that most organisms encode evolutionarily unrelated proteins with susceptibility to behave as prions. Conclusions To our knowledge, this is the first wide-ranging study aiming to predict prion domains in complete proteomes. Approaches of this kind could be of great importance to identify potential targets for further experimental testing and to try to reach a deeper understanding of prions’ functional and regulatory mechanisms. PMID:23663289

2013-01-01

22

Alzheimer's Disease and Prion Protein  

PubMed Central

Summary Alzheimer's disease (AD) is a devastating neurodegenerative disease with progressive loss of memory and cognitive function, pathologically hallmarked by aggregates of the amyloid-beta (A?) peptide and hyperphosphorylated tau in the brain. Aggregation of A? under the form of amyloid fibrils has long been considered central to the pathogenesis of AD. However, recent evidence has indicated that soluble A? oligomers, rather than insoluble fibrils, are the main neurotoxic species in AD. The cellular prion protein (PrPC) has newly been identified as a cell surface receptor for A? oligomers. PrPC is a cell surface glycoprotein that plays a key role in the propagation of prions, proteinaceous infectious agents that replicate by imposing their abnormal conformation to PrPC molecules. In AD, PrPC acts to transduce the neurotoxic signals arising from A? oligomers, leading to synaptic failure and cognitive impairment. Interestingly, accumulating evidence has also shown that aggregated A? or tau possesses prion-like activity, a property that would allow them to spread throughout the brain. In this article, we review recent findings regarding the function of PrPC and its role in AD, and discuss potential therapeutic implications of PrPC-based approaches in the treatment of AD. PMID:25343100

Zhou, Jiayi; Liu, Bingqian

2013-01-01

23

Protein misfolding cyclic amplification of infectious prions  

PubMed Central

Prions are proteinaceous infectious agents responsible for the transmission of prion diseases. The lack of a procedure for cultivating prions in the laboratory has been a major limitation to the study of the unorthodox nature of this infectious agent and the molecular mechanism by which the normal prion protein (PrPC) is converted into the abnormal isoform (PrPSc). Protein misfolding cyclic amplification (PMCA ), described in detail in this protocol, is a simple, fast and efficient methodology to mimic prion replication in the test tube. PMCA involves incubating materials containing minute amounts of infectious prions with an excess of PrPC and boosting the conversion by cycles of sonication to fragment the converting units, thereby leading to accelerated prion replication. PMCA is able to detect the equivalent of a single molecule of infectious PrPSc and propagate prions that maintain high infectivity, strain properties and species specificity. A single PMCA assay takes little more than 3 d to replicate a large amount of prions, which could take years in an in vivo situation. Since its invention 10 years ago, PMCA has helped to answer fundamental questions about this intriguing infectious agent and has been broadly applied in research areas that include the food industry, blood bank safety and human and veterinary disease diagnosis. PMID:22743831

Morales, Rodrigo; Duran-Aniotz, Claudia; Diaz-Espinoza, Rodrigo; Camacho, Manuel V; Soto, Claudio

2014-01-01

24

Thermodynamics of Model Prions and its Implications for the Problem of Prion Protein Folding  

E-print Network

Thermodynamics of Model Prions and its Implications for the Problem of Prion Protein Folding Paul M University of California San Francisco, 513 Parnassus Ave, San Francisco CA 94143, USA Prion disease is caused by the propagation of a particle containing PrPSc , a misfolded form of the normal cellular prion

Chan, Hue Sun

25

Application of quantitative real-time PCR in the detection of prion-protein gene species-specific DNA sequences in animal meals and feedstuffs.  

PubMed

This study describes a method for quantitative and species-specific detection of animal DNA from different species (cattle, sheep, goat, swine, and chicken) in animal feed and feed ingredients, including fish meals. A quantitative real-time PCR approach was carried out to characterize species-specific sequences based on the amplification of prion-protein sequence. Prion-protein species-specific primers and TaqMan probes were designed, and amplification protocols were optimized in order to discriminate the different species with short PCR amplicons. The real-time quantitative PCR approach was also compared to conventional species-specific PCR assays. The real-time quantitative assay allowed the detection of 10 pg of ruminant, swine, and poultry DNA extracted from meat samples processed at 130 degrees C for 40 min, 200 kPa. The origin of analyzed animal meals was characterized by the quantitative estimation of ruminant, swine, and poultry DNA. The TaqMan assay was used to quantify ruminant DNA in feedstuffs with 0.1% of meat and bone meal. In conclusion, the proposed molecular approach allowed the detection of species-specific DNA in animal meals and feedstuffs. PMID:16629035

Bellagamba, Federica; Comincini, Sergio; Ferretti, Luca; Valfrè, Franco; Moretti, Vittorio M

2006-04-01

26

Horse prion protein NMR structure and comparisons with related variants of the mouse prion protein.  

PubMed

The NMR structure of the horse (Equus caballus) cellular prion protein at 25 degrees C exhibits the typical PrP(C) [cellular form of prion protein (PrP)] global architecture, but in contrast to most other mammalian PrP(C)s, it contains a well-structured loop connecting the beta2 strand with the alpha2 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 degrees C and is unique to the PrP sequences of equine species. The beta2-alpha2 loop and the alpha3 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 beta2-alpha2 loop structure affects the susceptibility of transgenic mice to TSEs and the fact that there are no reports on TSE in horses. PMID:20460128

Pérez, Daniel R; Damberger, Fred F; Wüthrich, Kurt

2010-07-01

27

Spontaneous Variants of the [RNQ+] Prion in Yeast Demonstrate the Extensive Conformational Diversity Possible with Prion Proteins  

PubMed Central

Prion strains (or variants) are structurally distinct amyloid conformations arising from a single polypeptide sequence. The existence of prion strains has been well documented in mammalian prion diseases. In many cases, prion strains manifest as variation in disease progression and pathology, and in some cases, these prion strains also show distinct biochemical properties. Yet, the underlying basis of prion propagation and the extent of conformational possibilities available to amyloidogenic proteins remain largely undefined. Prion proteins in yeast that are also capable of maintaining multiple self-propagating structures have provided much insight into prion biology. Here, we explore the vast structural diversity of the yeast prion [RNQ+] in Saccharomyces cerevisiae. We screened for the formation of [RNQ+] in vivo, allowing us to calculate the rate of spontaneous formation as ~2.96x10-6, and successfully isolate several different [RNQ+] variants. Through a comprehensive set of biochemical and biological analyses, we show that these prion variants are indeed novel. No individual property or set of properties, including aggregate stability and size, was sufficient to explain the physical basis and range of prion variants and their resulting cellular phenotypes. Furthermore, all of the [RNQ+] variants that we isolated were able to facilitate the de novo formation of the yeast prion [PSI+], an epigenetic determinant of translation termination. This supports the hypothesis that [RNQ+] acts as a functional amyloid in regulating the formation of [PSI+] to produce phenotypic diversity within a yeast population and promote adaptation. Collectively, this work shows the broad spectrum of available amyloid conformations, and thereby expands the foundation for studying the complex factors that interact to regulate the propagation of distinct aggregate structures. PMID:24205387

Huang, Vincent J.; Stein, Kevin C.; True, Heather L.

2013-01-01

28

Yeast prion architecture explains how proteins can be genes  

NASA Astrophysics Data System (ADS)

Prions (infectious proteins) transmit information without an accompanying DNA or RNA. Most yeast prions are self-propagating amyloids that inactivate a normally functional protein. A single protein can become any of several prion variants, with different manifestations due to different amyloid structures. We showed that the yeast prion amyloids of Ure2p, Sup35p and Rnq1p are folded in-register parallel beta sheets using solid state NMR dipolar recoupling experiments, mass-per-filament-length measurements, and filament diameter measurements. The extent of beta sheet structure, measured by chemical shifts in solid-state NMR and acquired protease-resistance on amyloid formation, combined with the measured filament diameters, imply that the beta sheets must be folded along the long axis of the filament. We speculate that prion variants of a single protein sequence differ in the location of these folds. Favorable interactions between identical side chains must hold these structures in-register. The same interactions must guide an unstructured monomer joining the end of a filament to assume the same conformation as molecules already in the filament, with the turns at the same locations. In this way, a protein can template its own conformation, in analogy to the ability of a DNA molecule to template its sequence by specific base-pairing.

Wickner, Reed

2013-03-01

29

Prion protein in health and disease  

E-print Network

The prion protein (PrP) is a conserved glycoprotein tethered to cell membranes by a glycosylphosphatidylinositol anchor. In mammals, PrP is expressed in many tissues, most abundantly in brain, heart, and muscle. Importantly, ...

Steele, Andrew D., Ph. D. Massachusetts Institute of Technology

2008-01-01

30

Regional Mapping of Prion Proteins in Brain  

Microsoft Academic Search

Scrapie is characterized by the accumulation of a protease-resistant isoform of the prion protein PrPSc. Limited proteolysis and chaotropes were used to map the distribution of PrPSc in cryostat sections of scrapie-infected brain blotted onto nitrocellulose membranes, designated histoblots. Proteolysis was omitted in order to map the cellular isoform of the prion protein (PrP^C) in uninfected brains. Compared with immunohistochemistry,

Albert Taraboulos; Klaus Jendroska; Dan Serban; Shu-Lian Yang; Stephen J. Dearmond

1992-01-01

31

Polymorphisms and variants in the prion protein sequence of European moose (Alces alces), reindeer (Rangifer tarandus), roe deer (Capreolus capreolus) and fallow deer (Dama dama) in Scandinavia.  

PubMed

The prion protein (PrP) sequence of European moose, reindeer, roe deer and fallow deer in Scandinavia has high homology to the PrP sequence of North American cervids. Variants in the European moose PrP sequence were found at amino acid position 109 as K or Q. The 109Q variant is unique in the PrP sequence of vertebrates. During the 1980s a wasting syndrome in Swedish moose, Moose Wasting Syndrome (MWS), was described. SNP analysis demonstrated a difference in the observed genotype proportions of the heterozygous Q/K and homozygous Q/Q variants in the MWS animals compared with the healthy animals. In MWS moose the allele frequencies for 109K and 109Q were 0.73 and 0.27, respectively, and for healthy animals 0.69 and 0.31. Both alleles were seen as heterozygotes and homozygotes. In reindeer, PrP sequence variation was demonstrated at codon 176 as D or N and codon 225 as S or Y. The PrP sequences in roe deer and fallow deer were identical with published GenBank sequences. PMID:22441661

Wik, Lotta; Mikko, Sofia; Klingeborn, Mikael; Stéen, Margareta; Simonsson, Magnus; Linné, Tommy

2012-07-01

32

Prion proteins: a biological role beyond prion diseases.  

PubMed

The biological role of the scrapie isoform of prion protein (PrP(Sc)) as an infectious agent in numerous human and non-human disorders of the central nervous system is well established. In contrast, and despite decades of intensive research, the physiological function of the endogenous cellular form of the prion protein (PrP(C)) remains elusive. In mammals, the ubiquitous expression of PrP(C) suggests biological functions other than its pathological role in propagating the accumulation of its misfolded isotype. Other functions that have been attributed to PrP(C) include signal transduction, synaptic transmission and protection against cell death through the apoptotic pathway. More recently, immunoregulatory properties of PrP(C) have been reported. We review accumulating in vitro and in vivo evidence regarding physiological functions of PrP(C). PMID:17661791

Hu, W; Rosenberg, R N; Stüve, O

2007-08-01

33

Role of microglia and host prion protein in neurotoxicity of a prion protein fragment  

Microsoft Academic Search

THE prion protein PrPc is a glycoprotein of unknown function1 normally found in neurons2 and glia3. It is involved in diseases such as bovine spongiform encephalopathy (BSE), scrapie and Creutzfeldt-Jakob disease4. PrPSc, an altered isoform of PrPc that is associated with disease, shows greater protease resistance and is part of the infectious agent, the prion5,6. Prion diseases are characterized by

David R. Brown; Bernhard Schmidt; Hans A. Kretzschmar

1996-01-01

34

Conformational variations in an infectious protein determine prion strain differences  

Microsoft Academic Search

A remarkable feature of prion biology is the strain phenomenon wherein prion particles apparently composed of the same protein lead to phenotypically distinct transmissible states. To reconcile the existence of strains with the `protein-only' hypothesis of prion transmission, it has been proposed that a single protein can misfold into multiple distinct infectious forms, one for each different strain. Several studies

Motomasa Tanaka; Peter Chien; Nariman Naber; Roger Cooke; Jonathan S. Weissman

2004-01-01

35

Structural studies of the scrapie prion protein by electron crystallography  

E-print Network

Structural studies of the scrapie prion protein by electron crystallography Holger Wille* , Melissa. Prusiner, December 27, 2001 Because the insolubility of the scrapie prion protein (PrPSc) has frustrated to characterize the structure of two infectious variants of the prion protein. Isomor- phous two

Agard, David

36

Original Research Communication Prion Protein Expression and Functional Importance in  

E-print Network

1 1 Original Research Communication Prion Protein Expression and Functional Importance illustrations: 7 Color illustrations: 2 (online 2) Page 1 of 48 Antioxidants&RedoxSignaling PrionProtein prion protein (PrPC ), a GPI-anchored glycoprotein, which we reported to be highly expressed in human

Paris-Sud XI, Université de

37

Unusual Property of Prion Protein Unfolding in Neutral Salt Solution  

Microsoft Academic Search

The unfolding of cellular prion protein and its refolding to the scrapie isoform are related to prion diseases. Studies in the literature have shown that structures of proteins, either acidic or basic, are stabilized against denaturation by certain neutral salts, for example, sulfate and fluoride. Contrary to these observations, the full-length recombinant prion protein (amino acid residues 23 -231) is

P. K. Nandi; E. Leclerc; D. Marc

2002-01-01

38

Cellular Prion Protein: From Physiology to Pathology  

PubMed Central

The human cellular prion protein (PrPC) is a glycosylphosphatidylinositol (GPI) anchored membrane glycoprotein with two N-glycosylation sites at residues 181 and 197. This protein migrates in several bands by Western blot analysis (WB). Interestingly, PNGase F treatment of human brain homogenates prior to the WB, which is known to remove the N-glycosylations, unexpectedly gives rise to two dominant bands, which are now known as C-terminal (C1) and N-terminal (N1) fragments. This resembles the ?-amyloid precursor protein (APP) in Alzheimer disease (AD), which can be physiologically processed by ?-, ?-, and ?-secretases. The processing of APP has been extensively studied, while the identity of the cellular proteases involved in the proteolysis of PrPC and their possible role in prion biology has remained limited and controversial. Nevertheless, there is a strong correlation between the neurotoxicity caused by prion proteins and the blockade of their normal proteolysis. For example, expression of non-cleavable PrPC mutants in transgenic mice generates neurotoxicity, even in the absence of infectious prions, suggesting that PrPC proteolysis is physiologically and pathologically important. As many mouse models of prion diseases have recently been developed and the knowledge about the proteases responsible for the PrPC proteolysis is accumulating, we examine the historical experimental evidence and highlight recent studies that shed new light on this issue. PMID:23202518

Yusa, Sei-ichi; Oliveira-Martins, José B.; Sugita-Konishi, Yoshiko; Kikuchi, Yutaka

2012-01-01

39

Multiple folding pathways for heterologously expressed human prion protein  

E-print Network

Multiple folding pathways for heterologously expressed human prion protein Graham S. Jackson , Anthony R. Clarke a , John Collinge aY * a Prion Disease Group, Department of Neurogenetics, Imperial-conformation in free solution. The data we present here shows that the human prion protein can exist in multiple

Hosszu, Laszlo

40

Chronic wasting disease prions are not transmissible to transgenic mice overexpressing human prion protein.  

PubMed

Chronic wasting disease (CWD) is a prion disease that affects free-ranging and captive cervids, including mule deer, white-tailed deer, Rocky Mountain elk and moose. CWD-infected cervids have been reported in 14 USA states, two Canadian provinces and in South Korea. The possibility of a zoonotic transmission of CWD prions via diet is of particular concern in North America where hunting of cervids is a popular sport. To investigate the potential public health risks posed by CWD prions, we have investigated whether intracerebral inoculation of brain and spinal cord from CWD-infected mule deer transmits prion infection to transgenic mice overexpressing human prion protein with methionine or valine at polymorphic residue 129. These transgenic mice have been utilized in extensive transmission studies of human and animal prion disease and are susceptible to BSE and vCJD prions, allowing comparison with CWD. Here, we show that these mice proved entirely resistant to infection with mule deer CWD prions arguing that the transmission barrier associated with this prion strain/host combination is greater than that observed with classical BSE prions. However, it is possible that CWD may be caused by multiple prion strains. Further studies will be required to evaluate the transmission properties of distinct cervid prion strains as they are characterized. PMID:20610667

Sandberg, Malin K; Al-Doujaily, Huda; Sigurdson, Christina J; Glatzel, Markus; O'Malley, Catherine; Powell, Caroline; Asante, Emmanuel A; Linehan, Jacqueline M; Brandner, Sebastian; Wadsworth, Jonathan D F; Collinge, John

2010-10-01

41

Impaired Motor Coordination in Mice Lacking Prion Protein  

Microsoft Academic Search

1. Prion protein (PrPC) is a host-encoded glycoprotein constitutively expressed on the neuronal cell surface. Accumulation of its protease-resistant isoform is closely related to pathologic changes and prion propagation in the brain tissue of a series of prion diseases. However, the physiological role of PrPC remains to be elucidated.

Shigeru Katamine; Noriyuki Nishida; Tetsuo Sugimoto; Tetsuo Noda; Suehiro Sakaguchi; Kazuto Shigematsu; Yasufumi Kataoka; Akira Nakatani; Sumitaka Hasegawa; Ryozo Moriuchi; Tsutomu Miyamoto

1998-01-01

42

Copper Binding to the N-Terminal Tandem Repeat Regions of Mammalian and Avian Prion Protein  

Microsoft Academic Search

Mammalian prion protein (PrP) is a normal cellular protein (PrPc) which through post-translational modification produces the infectious prion protein (PrPsc). We have shown, using mass spectrometry, that synthetic peptides containing three or four copies of an octapeptide repeat sequence (PHGGGWGQ), found in a highly conserved N-terminal domain of PrP, preferentially bind copper over other metals. Peptides from the analogous region

M. P. Hornshaw; J. R. Mcdermott; J. M. Candy

1995-01-01

43

Prions and protein-folding diseases.  

PubMed

Prions represent a group of proteins with a unique capacity to fold into different conformations. One isoform is rich in beta-pleated sheets and can aggregate into amyloid that may be pathogenic. This abnormal form propagates itself by imposing its confirmation on the homologous normal host cell protein. Pathogenic prions have been shown to cause lethal neurodegenerative diseases in humans and animals. These diseases are sometimes infectious and hence referred to as transmissible spongiform encephalopathies. In the present review, the remarkable evolution of the heterodox prion concept is summarized. The origin of this phenomenon is based on information transfer between homologous proteins, without the involvement of nucleic acid-encoded mechanisms. Historically, kuru and Creutzfeldt-Jakob disease (CJD) were the first infectious prion diseases to be identified in man. It was their relationship to scrapie in sheep and experimental rodents that allowed an unravelling of the particular molecular mechanism that underlie the disease process. Transmission between humans has been documented to have occurred in particular contexts, including ritual cannibalism, iatrogenic transmission because of pituitary gland-derived growth hormone or the use in neurosurgical procedures of dura mater from cadavers, and the temporary use of a prion-contaminated protein-rich feed for cows. The latter caused a major outbreak of bovine spongiform encephalopathy, which spread to man by human consumption of contaminated meat, causing approximately 200 cases of variant CJD. All these epidemics now appear to be over because of measures taken to curtail further spread of prions. Recent studies have shown that the mechanism of protein aggregation may apply to a wider range of diseases in and possibly also outside the brain, some of which are relatively common such as Alzheimer's and Parkinson's diseases. Furthermore, it has become apparent that the phenomenon of prion aggregation may have a wider physiological importance, but a full understanding of this remains to be defined. It may involve maintaining neuronal functions and possibly contributing to the establishment of long-term memory. PMID:21481020

Norrby, E

2011-07-01

44

Neurotoxicity of a prion protein fragment  

Microsoft Academic Search

THE cellular prion protein (PrPc) is a sialoglycoprotein of Mr 33-35K that is expressed predominantly in neurons1-3. In transmissible and genetic neurodegenerative disorders such as scrapie of sheep, spongiform encephalopathy of cattle and Creutzfeldt-Jakob or Gerstmann-Sträussler-Scheinker diseases of humans4,5, PrPc is converted into an altered form (termed PrPSc) which is distinguishable from its normal homologue by its relative resistance to

Gianluigi Forloni; Nadia Angeretti; Roberto Chiesa; Enrico Monzani; Mario Salmona; Orso Bugiani; Fabrizio Tagliavini

1993-01-01

45

The Complete CDS of the Prion Protein ( PRNP ) Gene of African Lion ( Panthera leo )  

Microsoft Academic Search

We provide the complete PRNP CDS sequence for the African lion, which is different from the previously published sequence and more similar to other carnivore\\u000a sequences. The newly obtained prion protein sequence differs from the domestic cat sequence at three amino acid positions\\u000a and contains only four octapeptide repeats. We recommend that this sequence be used as the reference sequence

Andrzej Maj; Garth M. Spellman; Shane K. Sarver

2008-01-01

46

The complete CDS of the prion protein (PRNP) gene of African lion (Panthera leo).  

PubMed

We provide the complete PRNP CDS sequence for the African lion, which is different from the previously published sequence and more similar to other carnivore sequences. The newly obtained prion protein sequence differs from the domestic cat sequence at three amino acid positions and contains only four octapeptide repeats. We recommend that this sequence be used as the reference sequence for future studies of the PRNP gene for this species. PMID:18256917

Maj, Andrzej; Spellman, Garth M; Sarver, Shane K

2008-04-01

47

Sialylation of prion protein controls the rate of prion amplification, the cross-species barrier, the ratio of PrPSc glycoform and prion infectivity.  

PubMed

The central event underlying prion diseases involves conformational change of the cellular form of the prion protein (PrP(C)) into the disease-associated, transmissible form (PrP(Sc)). Pr(PC) is a sialoglycoprotein that contains two conserved N-glycosylation sites. Among the key parameters that control prion replication identified over the years are amino acid sequence of host PrP(C) and the strain-specific structure of PrPSc. The current work highlights the previously unappreciated role of sialylation of PrP(C) glycans in prion pathogenesis, including its role in controlling prion replication rate, infectivity, cross-species barrier and PrP(Sc) glycoform ratio. The current study demonstrates that undersialylated PrP(C) is selected during prion amplification in Protein Misfolding Cyclic Amplification (PMCAb) at the expense of oversialylated PrP(C). As a result, PMCAb-derived PrP(Sc) was less sialylated than brain-derived PrP(Sc). A decrease in PrPSc sialylation correlated with a drop in infectivity of PMCAb-derived material. Nevertheless, enzymatic de-sialylation of PrP(C) using sialidase was found to increase the rate of PrP(Sc) amplification in PMCAb from 10- to 10,000-fold in a strain-dependent manner. Moreover, de-sialylation of PrP(C) reduced or eliminated a species barrier of for prion amplification in PMCAb. These results suggest that the negative charge of sialic acid controls the energy barrier of homologous and heterologous prion replication. Surprisingly, the sialylation status of PrP(C) was also found to control PrP(Sc) glycoform ratio. A decrease in Pr(PC) sialylation levels resulted in a higher percentage of the diglycosylated glycoform in PrP(Sc). 2D analysis of charge distribution revealed that the sialylation status of brain-derived PrP(C) differed from that of spleen-derived PrP(C). Knocking out lysosomal sialidase Neu1 did not change the sialylation status of brain-derived PrP(C), suggesting that Neu1 is not responsible for desialylation of PrP(C). The current work highlights previously unappreciated role of PrP(C) sialylation in prion diseases and opens multiple new research directions, including development of new therapeutic approaches. PMID:25211026

Katorcha, Elizaveta; Makarava, Natallia; Savtchenko, Regina; D'Azzo, Alessandra; Baskakov, Ilia V

2014-09-01

48

Mutant prion protein acquires resistance to protease in mouse neuroblastoma cells  

Microsoft Academic Search

Conversion of the cellular isoform of the prion protein (PrPC) into the pathogenic isoform (PrPSc )i s thought to be the causative event in prion diseases. Biochemically, PrPSc differs from PrPC in its partial resistance to proteinase K (PK). The amino acid sequence AGAAAAGA, comprising residues 112-119 of the murine PrPC, has been shown to be amyloidogenic and evolutionarily conserved.

C. Wegner; A. Ro; R. Schmalzbauer; H. Lorenz; O. Windl; H. A. Kretzschmar

2002-01-01

49

Spontaneous Neurodegeneration in Transgenic Mice with Mutant Prion Protein  

Microsoft Academic Search

Transgenic mice were created to assess genetic linkage between Gerstmann-Straussler-Scheinker syndrome and a leucine substitution at codon 102 of the human prion protein gene. Spontaneous neurologic disease with spongiform degeneration and gliosis similar to that in mouse scrapie developed at a mean age of 166 days in 35 mice expressing mouse prion protein with the leucine substitution. Thus, many of

Karen K. Hsiao; Michael Scott; Dallas Foster; Darlene F. Groth; Stephen J. Dearmond

1990-01-01

50

Prion Protein Misfolding, Strains, and Neurotoxicity: An Update from Studies on Mammalian Prions  

PubMed Central

Prion diseases, also known as transmissible spongiform encephalopathies (TSEs), are a group of fatal neurodegenerative disorders affecting humans and other mammalian species. The central event in TSE pathogenesis is the conformational conversion of the cellular prion protein, PrPC, into the aggregate, ?-sheet rich, amyloidogenic form, PrPSc. Increasing evidence indicates that distinct PrPSc conformers, forming distinct ordered aggregates, can encipher the phenotypic TSE variants related to prion strains. Prion strains are TSE isolates that, after inoculation into syngenic hosts, cause disease with distinct characteristics, such as incubation period, pattern of PrPSc distribution, and regional severity of histopathological changes in the brain. In analogy with other amyloid forming proteins, PrPSc toxicity is thought to derive from the existence of various intermediate structures prior to the amyloid fiber formation and/or their specific interaction with membranes. The latter appears particularly relevant for the pathogenesis of TSEs associated with GPI-anchored PrPSc, which involves major cellular membrane distortions in neurons. In this review, we update the current knowledge on the molecular mechanisms underlying three fundamental aspects of the basic biology of prions such as the putative mechanism of prion protein conversion to the pathogenic form PrPSc and its propagation, the molecular basis of prion strains, and the mechanism of induced neurotoxicity by PrPSc aggregates. PMID:24454379

Poggiolini, Ilaria; Parchi, Piero

2013-01-01

51

Prion protein induced signaling cascades in monocytes  

SciTech Connect

Prion proteins play a central role in transmission and pathogenesis of transmissible spongiform encephalopathies. The cellular prion protein (PrP{sup C}), whose physiological function remains elusive, is anchored to the surface of a variety of cell types including neurons and cells of the lymphoreticular system. In this study, we investigated the response of a mouse monocyte/macrophage cell line to exposure with PrP{sup C} fusion proteins synthesized with a human Fc-tag. PrP{sup C} fusion proteins showed an attachment to the surface of monocyte/macrophages in nanomolar concentrations. This was accompanied by an increase of cellular tyrosine phosphorylation as a result of activated signaling pathways. Detailed investigations exhibited activation of downstream pathways through a stimulation with PrP fusion proteins, which include phosphorylation of ERK{sub 1,2} and Akt kinase. Macrophages opsonize and present antigenic structures, contact lymphocytes, and deliver cytokines. The findings reported here may become the basis of understanding the molecular function of PrP{sup C} in monocytes and macrophages.

Krebs, Bjarne [Center for Neuropathology and Prion Research, Ludwig-Maximilians-University Munich, Muenchen (Germany); Dorner-Ciossek, Cornelia [CNS Research III, Boehringer Ingelheim Pharma GmbH and Co KG, Biberach/Riss (Germany); Schmalzbauer, Ruediger [Center for Neuropathology and Prion Research, Ludwig-Maximilians-University Munich, Munich (Germany); Vassallo, Neville [Department of Physiology and Biochemistry, University of Malta, Msida (Malta); Herms, Jochen [Center for Neuropathology and Prion Research, Ludwig-Maximilians-University Munich, Munich (Germany); Kretzschmar, Hans A. [Center for Neuropathology and Prion Research, Ludwig-Maximilians-University Munich, Munich (Germany)]. E-mail: Hans.Kretzschmar@med.uni-muenchen.de

2006-02-03

52

Prions: protein only or something more? Overview of potential prion cofactors.  

PubMed

Transmissible spongiform encephalopathies (TSEs) in humans and animals are attributed to protein-only infectious agents, called prions. Prions have been proposed to arise from the conformational conversion of the cellular protein PrP(C) into a misfolded form (e.g., PrP(Sc) for scrapie), which precipitates into aggregates and fibrils. It has been proposed that the conversion process is triggered by the interaction of the infectious form (PrP(Sc)) with the cellular form (PrP(C)) or might result from a mutation in the gene for PrP(C). However, until recently, all efforts to reproduce this process in vitro had failed, suggesting that host factors are necessary for prion replication. In this review we discuss recent findings such as the cellular factors that might be involved in the conformational conversion of prion proteins and the potential mechanisms by which they could operate. PMID:17085779

Fasano, Carlo; Campana, Vincenza; Zurzolo, Chiara

2006-01-01

53

Context Dependent Neuroprotective Properties of Prion Protein (Prp)  

E-print Network

Although it has been known for more than twenty years that an aberrant conformation of the prion protein (PrP) is the causative agent in prion diseases, the role of PrP in normal biology is undetermined. Numerous studies ...

Steele, Andrew D.

54

Prion neuropathology follows the accumulation of alternate prion protein isoforms after infective titre has peaked  

PubMed Central

Prions are lethal infectious agents thought to consist of multi-chain forms (PrPSc) of misfolded cellular prion protein (PrPC). Prion propagation proceeds in two distinct mechanistic phases: an exponential phase 1, which rapidly reaches a fixed level of infectivity irrespective of PrPC expression level, and a plateau (phase 2), which continues until clinical onset with duration inversely proportional to PrPC expression level. We hypothesized that neurotoxicity relates to distinct neurotoxic species produced following a pathway switch when prion levels saturate. Here we show a linear increase of proteinase K-sensitive PrP isoforms distinct from classical PrPSc at a rate proportional to PrPC concentration, commencing at the phase transition and rising until clinical onset. The unaltered level of total PrP during phase 1, when prion infectivity increases a million-fold, indicates that prions comprise a small minority of total PrP. This is consistent with PrPC concentration not being rate limiting to exponential prion propagation and neurotoxicity relating to critical concentrations of alternate PrP isoforms whose production is PrPC concentration dependent. PMID:25005024

Sandberg, Malin K.; Al-Doujaily, Huda; Sharps, Bernadette; De Oliveira, Michael Wiggins; Schmidt, Christian; Richard-Londt, Angela; Lyall, Sarah; Linehan, Jacqueline M.; Brandner, Sebastian; Wadsworth, Jonathan D. F.; Clarke, Anthony R.; Collinge, John

2014-01-01

55

Prion neuropathology follows the accumulation of alternate prion protein isoforms after infective titre has peaked.  

PubMed

Prions are lethal infectious agents thought to consist of multi-chain forms (PrP(Sc)) of misfolded cellular prion protein (PrP(C)). Prion propagation proceeds in two distinct mechanistic phases: an exponential phase 1, which rapidly reaches a fixed level of infectivity irrespective of PrP(C) expression level, and a plateau (phase 2), which continues until clinical onset with duration inversely proportional to PrP(C) expression level. We hypothesized that neurotoxicity relates to distinct neurotoxic species produced following a pathway switch when prion levels saturate. Here we show a linear increase of proteinase K-sensitive PrP isoforms distinct from classical PrP(Sc) at a rate proportional to PrP(C) concentration, commencing at the phase transition and rising until clinical onset. The unaltered level of total PrP during phase 1, when prion infectivity increases a million-fold, indicates that prions comprise a small minority of total PrP. This is consistent with PrP(C) concentration not being rate limiting to exponential prion propagation and neurotoxicity relating to critical concentrations of alternate PrP isoforms whose production is PrP(C) concentration dependent. PMID:25005024

Sandberg, Malin K; Al-Doujaily, Huda; Sharps, Bernadette; De Oliveira, Michael Wiggins; Schmidt, Christian; Richard-Londt, Angela; Lyall, Sarah; Linehan, Jacqueline M; Brandner, Sebastian; Wadsworth, Jonathan D F; Clarke, Anthony R; Collinge, John

2014-01-01

56

Stability and conformational properties of doppel, a prion-like protein, and its single-disulphide mutant.  

PubMed Central

Both prion protein and the structurally homologous protein doppel are associated with neurodegenerative disease by mechanisms which remain elusive. We have prepared murine doppel, and a mutant with one of the two disulphide bonds removed, in the expectation of increasing the similarity of doppel to prion protein in terms of conformation and stability. Unfolding studies of doppel and the mutant have been performed using far-UV CD over a range of solution conditions known to favour the alpha-->beta transformation of recombinant prion protein. Only partial unfolding of doppel or the mutant occurs at elevated temperature, but both exhibit full and reversible unfolding in chemical denaturation with urea. Doppel is significantly less stable than prion protein, and this stability is further reduced by removal of the disulphide bond between residues 95-148. Both doppel and the mutant are observed to unfold by a two-state mechanism, even under the mildly acidic conditions where prion protein forms an equilibrium intermediate with enhanced beta-structure, potentially analogous to the conversion of the cellular form of the prion protein into the infectious form (PrP(C)-->PrP(Sc)). Furthermore, no direct interaction of either doppel protein with prion protein, either in the alpha-form or the beta-rich conformation, was detectable spectroscopically. These studies indicate that, in spite of the similarity in secondary structure between the doppel and prion protein, there are significant differences in their solution properties. The fact that neither doppel nor its mutant exhibited the alpha-->beta transformation of the prion protein suggests that this conversion property may be dependent on unique sequences specific to the prion protein. PMID:12665426

Whyte, Sheena M; Sylvester, Ian D; Martin, Stephen R; Gill, Andrew C; Wopfner, Franziska; Schatzl, Hermann M; Dodson, Guy G; Bayley, Peter M

2003-01-01

57

Computational Studies of the Structural Stability of Rabbit Prion Protein Compared to Human and Mouse Prion Proteins  

E-print Network

Prion diseases are invariably fatal and highly infectious neurodegenerative diseases affecting humans and animals. The neurodegenerative diseases such as Creutzfeldt-Jakob disease, variant Creutzfeldt-Jakob diseases, Gerstmann-Str$\\ddot{a}$ussler-Scheinker syndrome, Fatal Familial Insomnia, Kuru in humans, scrapie in sheep, bovine spongiform encephalopathy (or 'mad-cow' disease) and chronic wasting disease in cattle belong to prion diseases. By now there have not been some effective therapeutic approaches to treat all these prion diseases. Dogs, rabbits and horses were reported to be resistant to prion diseases. By the end of year 2010 all the NMR structures of dog, rabbit and horse prion proteins (X-ray for rabbits too) had been finished to release into protein data bank. Thus, at this moment it is very worth studying the NMR and X-ray molecular structures of horse, dog and rabbit prion proteins to obtain insights into their immunity prion diseases. The author found that dog and horse prion proteins have sta...

Zhang, Jiapu

2011-01-01

58

SSCP analysis and sequencing of the human prion protein gene (PRNP) detects two different 24 bp deletions in an atypical Alzheimer`s disease family  

SciTech Connect

Alzheimer`s disease (AD) is a progressive, degenerative neurological disorder of the central nervous system. AD is the fourth leading cause of death in elderly persons 65 years or older in Western industrialized societies. The etiology of AD is unknown, but clinical, pathological, epidemiological, and molecular investigations suggest it is etiologically heterogeneous. Mutations in the amyloid protein are rare and segregate with the disease in a few early-onset familial AD (FAD) families. Similarities between AD and the unconventional viral (UCV) diseases, and between the amyloid and prion proteins, implicate the human prion protein gene (PRNP) as another candidate gene. Single strand conformation polymorphism (SSCP) analysis was used to screen for mutations at this locus in 82 AD patients from 54 families (30 FAD), vs. 39 age-matched controls. A 24-bp deletion around codon 68 that codes for one of five Gly-Pro rich octarepeats was identified in two affected sibs and one offspring of one late-onset FAD family. Two other affected sibs, three unaffected sibs, and three offspring from this family, in addition to one sporadic AD patient and three age-matched controls, were heterozygous for another octarepeat deletion located around codon 82. Two of the four affected sibs had features of PD, including one who was autopsy-verified AD and PD. Although these deletions were found infrequently in other AD patients and controls, they appear to be a rare polymorphism that is segregating in this FAD family. It does not appear that mutations at the PRNP locus are frequently associated with AD in this population. 54 refs., 4 figs.

Perry, R.T.; Go, R.C.P.; Harrell, L.E.; Acton, R.T. [Univ. of Alabama, Birmingham, AL (United States)

1995-02-27

59

Live-cell FRET imaging reveals clustering of the prion protein at the cell surface induced by infectious prions.  

PubMed

Prion diseases are associated to the conversion of the prion protein into a misfolded pathological isoform. The mechanism of propagation of protein misfolding by protein templating remains largely unknown. Neuroblastoma cells were transfected with constructs of the prion protein fused to both CFP-GPI-anchored and to YFP-GPI-anchored and directed to its cell membrane location. Live-cell FRET imaging between the prion protein fused to CFP or YFP was measured giving consistent values of 10±2%. This result was confirmed by fluorescence lifetime imaging microscopy and indicates intermolecular interactions between neighbor prion proteins. In particular, considering that a maximum FRET efficiency of 17±2% was determined from a positive control consisting of a fusion CFP-YFP-GPI-anchored. A stable cell clone expressing the two fusions containing the prion protein was also selected to minimize cell-to-cell variability. In both, stable and transiently transfected cells, the FRET efficiency consistently increased in the presence of infectious prions - from 4±1% to 7±1% in the stable clone and from 10±2% to 16±1% in transiently transfected cells. These results clearly reflect an increased clustering of the prion protein on the membrane in the presence of infectious prions, which was not observed in negative control using constructs without the prion protein and upon addition of non-infected brain. Our data corroborates the recent view that the primary site for prion conversion is the cell membrane. Since our fluorescent cell clone is not susceptible to propagate infectivity, we hypothesize that the initial event of prion infectivity might be the clustering of the GPI-anchored prion protein. PMID:24530636

Tavares, Evandro; Macedo, Joana A; Paulo, Pedro M R; Tavares, Catarina; Lopes, Carlos; Melo, Eduardo P

2014-07-01

60

REFINEMENT OF UNDER-DETERMINED LOOPS OF HUMAN PRION PROTEIN BY  

E-print Network

REFINEMENT OF UNDER-DETERMINED LOOPS OF HUMAN PRION PROTEIN BY DATABASE-DERIVED DISTANCE: http://www.ima.umn.edu #12;Refinement of Under-Determined Loops of Human Prion Protein by Database Computational simulations of the conversion from the normal cellular prion (PrPc ) to the scrapie prion (Pr

61

Evidence for Protein X Binding to a Discontinuous Epitope on the Cellular Prion Protein during Scrapie Prion Propagation  

Microsoft Academic Search

Studies on the transmission of human (Hu) prions to transgenic (Tg) mice suggested that another molecule provisionally designated protein X participates in the formation of nascent scrapie isoform of prion protein (PrPSc). We report the identification of the site at which protein X binds to the cellular isoform of PrP (PrPC) using scrapie-infected mouse (Mo) neuroblastoma cells transfected with chimeric

Kiyotoshi Kaneko; Laurence Zulianello; Michael Scott; Carol M. Cooper; Andrew C. Wallace; Thomas L. James; Fred E. Cohen

1997-01-01

62

Normal host prion protein necessary for scrapie-induced neurotoxicity  

Microsoft Academic Search

ACCUMULATION of the prion protein PrPSc, a pathological and protease-resistant isoform of the normal host protein PrPc, is a feature of prion disease such as scrapie1,2. It is still unknown whether scrapie pathology comes about by neurotoxicity of PrPSc, acute depletion of PrPc, or some other mechanism. Here we investigate this question by grafting neural tissue overexpressing PrPc into the

Sebastian Brandner; Stefan Isenmann; Alex Raeber; Marek Fischer; Andreas Sailer; Yasushi Kobayashi; Silvia Marino; Charles Weissmann; Adriano Aguzzi

1996-01-01

63

Prevalence of lymphoreticular prion protein accumulation in UK tissue samples  

Microsoft Academic Search

This study aims to provide an estimate of the number of individuals in the UK who may be incubating variant Creutzfeldt-Jakob disease and at risk of causing iatrogenic spread of the disease. Lymphoreticular accumulation of prion protein is a consistent feature of variant Creutzfeldt-Jakob at autopsy and has also been demonstrated in the pre-clinical phase. Immunohistochemical accumulation of prion protein

David A Hilton; Azra C Ghani; Lisa Conyers; Philip Edwards; Linda McCardle; Diane Ritchie; Mark Penney; Doha Hegazy; James W Ironside

2004-01-01

64

Prion disease and the 'protein-only hypothesis'.  

PubMed

Prion disease is the only naturally occurring infectious protein misfolding disorder. The chemical nature of the infectious agent has been debated for more than half a century. Early studies on scrapie suggested that the unusual infectious agent might propagate in the absence of nucleic acid. The 'protein-only hypothesis' provides a theoretical model to explain how a protein self-replicates without nucleic acid, which predicts that a prion, the proteinaceous infectious agent, propagates by converting its normal counterpart into the likeness of itself. Decades of studies have provided overwhelming evidence to support this hypothesis. The latest advances in generating infectious prions with bacterially expressed recombinant prion protein in the presence of cofactors not only provide convincing evidence supporting the 'protein-only hypothesis', but also indicate a role of cofactors in forming prion infectivity and encoding prion strains. In the present chapter, we review the literature regarding the chemical nature of the infectious agent, describe recent achievements in proving the 'protein-only hypothesis', and discuss the remaining questions in this research area. PMID:25131595

Ma, Jiyan; Wang, Fei

2014-01-01

65

Pulling rabbits to reveal the secrets of the prion protein.  

PubMed

The Prion protein (PrP) is a membrane-tethered glycoprotein that plays a central role in a unique class of neurodegenerative diseases that affect humans and other mammals. Prion diseases have genetic and sporadic origins, but their infectious nature sets them apart from other neurodegenerative disorders. According to the "protein-only" hypothesis, misfolded PrP conformers (prions) are responsible for both spongiform degeneration of the brain and disease transmissibility. Thus, understanding PrP conformational dynamics is key to developing effective therapies. Classic studies showing the different susceptibility to prion disease in mammals have recently found support in structural and transgenic studies with PrP from susceptible (mouse, hamster) and resistant (rabbit, horse, dog) animals. These studies identify key residues in PrP that determine both PrP structure and its propensity to acquire a ?-structure conformation proposed to be neurotoxic. PMID:21980555

Fernandez-Funez, Pedro; Zhang, Yan; Sanchez-Garcia, Jonatan; Jensen, Kurt; Zou, Wen-Quan; Rincon-Limas, Diego E

2011-05-01

66

Developmental expression of the prion protein gene in glial cells  

Microsoft Academic Search

Replication of prions is dependent on the presence of the host protein PrPc. During the course of disease, PrPc is converted into an abnormal isoform, PrPsc, which accumulates in the brain. Attempts to identify the cell type(s) in which prion replication and PrP conversion occur have reached conflicting results. Although PrP mRNA is present in high amounts in neurons throughout

Markus Moser; Raymond J Colello; Uwe Pott; Bruno Oesch

1995-01-01

67

Imbalance of antioxidant defense in mice lacking cellular prion protein  

Microsoft Academic Search

Prion diseases are fatal neurodegenerative disorders resulting from conformational changes in the prion protein from its normal cellular isoform, PrPC, to the infectious scrapie isoform, PrPSc. In spite of many studies, the physiological function of PrPC remains unknown. Recent work shows that PrPC binds Cu2+, internalizing it into the cytoplasm. Since many antioxidant enzymes depend on Cu2+ (e.g., Cu\\/ZnSOD), their

Fá bio Klamt; Felipe Dal-Pizzol; Mario Luiz Conte da Frota; Roger Walz; Michael Everton Andrades; Evandro Gomes da Silva; Ricardo R. Brentani; Ivá n Izquierdo; José Cláudio Fonseca Moreira

2001-01-01

68

Infectious Prion Protein Alters Manganese Transport and Neurotoxicity in a Cell Culture Model of Prion Disease  

PubMed Central

Protein misfolding and aggregation are considered key features of many neurodegenerative diseases, but biochemical mechanisms underlying protein misfolding and the propagation of protein aggregates are not well understood. Prion disease is a classical neurodegenerative disorder resulting from the misfolding of endogenously expressed normal cellular prion protein (PrPC). Although the exact function of PrPC has not been fully elucidated, studies have suggested that it can function as a metal binding protein. Interestingly, increased brain manganese (Mn) levels have been reported in various prion diseases indicating divalent metals also may play a role in the disease process. Recently, we reported that PrPC protects against Mn-induced cytotoxicity in a neural cell culture model. To further understand the role of Mn in prion diseases, we examined Mn neurotoxicity in an infectious cell culture model of prion disease. Our results show CAD5 scrapie-infected cells were more resistant to Mn neurotoxicity as compared to uninfected cells (EC50 = 428.8 ?M for CAD5 infected cells vs. 211.6 ?M for uninfected cells). Additionally, treatment with 300 ?M Mn in persistently infected CAD5 cells showed a reduction in mitochondrial impairment, caspase-3 activation, and DNA fragmentation when compared to uninfected cells. Scrapie-infected cells also showed significantly reduced Mn uptake as measured by inductively coupled plasma-mass spectrometry (ICP-MS), and altered expression of metal transporting proteins DMT1 and transferrin. Together, our data indicate that conversion of PrP to the pathogenic isoform enhances its ability to regulate Mn homeostasis, and suggest that understanding the interaction of metals with disease-specific proteins may provide further insight to protein aggregation in neurodegenerative diseases. PMID:21871919

Martin, Dustin P.; Anantharam, Vellareddy; Jin, Huajun; Witte, Travis; Houk, Robert; Kanthasamy, Arthi; Kanthasamy, Anumantha G.

2011-01-01

69

Cell Type-Specific Neuroprotective Activity of Untranslocated Prion Protein  

PubMed Central

Background A key pathogenic role in prion diseases was proposed for a cytosolic form of the prion protein (PrP). However, it is not clear how cytosolic PrP localization influences neuronal viability, with either cytotoxic or anti-apoptotic effects reported in different studies. The cellular mechanism by which PrP is delivered to the cytosol of neurons is also debated, and either retrograde transport from the endoplasmic reticulum or inefficient translocation during biosynthesis has been proposed. We investigated cytosolic PrP biogenesis and effect on cell viability in primary neuronal cultures from different mouse brain regions. Principal Findings Mild proteasome inhibition induced accumulation of an untranslocated form of cytosolic PrP in cortical and hippocampal cells, but not in cerebellar granules. A cyclopeptolide that interferes with the correct insertion of the PrP signal sequence into the translocon increased the amount of untranslocated PrP in cortical and hippocampal cells, and induced its synthesis in cerebellar neurons. Untranslocated PrP boosted the resistance of cortical and hippocampal neurons to apoptotic insults but had no effect on cerebellar cells. Significance These results indicate cell type-dependent differences in the efficiency of PrP translocation, and argue that cytosolic PrP targeting might serve a physiological neuroprotective function. PMID:21060848

Mantovani, Susanna; Airaghi, Simona; Forloni, Gianluigi; Chiesa, Roberto

2010-01-01

70

Molecular Features of the Copper Binding Sites in the Octarepeat Domain of the Prion Protein  

E-print Network

Molecular Features of the Copper Binding Sites in the Octarepeat Domain of the Prion Protein Colin evidence suggests that the prion protein (PrP) is a copper binding protein. The N-terminal region of human units that may facilitate molecular recognition between prion proteins, thereby suggesting a mechanism

Scott, William

71

Efficacy and Mechanism of a Glycoside Compound Inhibiting Abnormal Prion Protein Formation in Prion-Infected Cells: Implications of Interferon and Phosphodiesterase 4D-Interacting Protein  

PubMed Central

ABSTRACT A new type of antiprion compound, Gly-9, was found to inhibit abnormal prion protein formation in prion-infected neuroblastoma cells, in a prion strain-independent manner, when the cells were treated for more than 1 day. It reduced the intracellular prion protein level and significantly modified mRNA expression levels of genes of two types: interferon-stimulated genes were downregulated after more than 2 days of treatment, and the phosphodiesterase 4D-interacting protein gene, a gene involved in microtubule growth, was upregulated after more than 1 day of treatment. A supplement of interferon given to the cells partly restored the abnormal prion protein level but did not alter the normal prion protein level. This interferon action was independent of the Janus activated kinase-signal transducer and activator of transcription signaling pathway. Therefore, the changes in interferon-stimulated genes might be a secondary effect of Gly-9 treatment. However, gene knockdown of phosphodiesterase 4D-interacting protein restored or increased both the abnormal prion protein level and the normal prion protein level, without transcriptional alteration of the prion protein gene. It also altered the localization of abnormal prion protein accumulation in the cells, indicating that phosphodiesterase 4D-interacting protein might affect prion protein levels by altering the trafficking of prion protein-containing structures. Interferon and phosphodiesterase 4D-interacting protein had no direct mutual link, demonstrating that they regulate abnormal prion protein levels independently. Although the in vivo efficacy of Gly-9 was limited, the findings for Gly-9 provide insights into the regulation of abnormal prion protein in cells and suggest new targets for antiprion compounds. IMPORTANCE This report describes our study of the efficacy and potential mechanism underlying the antiprion action of a new antiprion compound with a glycoside structure in prion-infected cells, as well as the efficacy of the compound in prion-infected animals. The study revealed involvements of two factors in the compound's mechanism of action: interferon and a microtubule nucleation activator, phosphodiesterase 4D-interacting protein. In particular, phosphodiesterase 4D-interacting protein was suggested to be important in regulating the trafficking or fusion of prion protein-containing vesicles or structures in cells. The findings of the study are expected to be useful not only for the elucidation of cellular regulatory mechanisms of prion protein but also for the implication of new targets for therapeutic development. PMID:24453367

Nishizawa, Keiko; Oguma, Ayumi; Kawata, Maki; Sakasegawa, Yuji; Teruya, Kenta

2014-01-01

72

Low Copper and High Manganese Levels in Prion Protein Plaques  

PubMed Central

Accumulation of aggregates rich in an abnormally folded form of the prion protein characterize the neurodegeneration caused by transmissible spongiform encephalopathies (TSEs). The molecular triggers of plaque formation and neurodegeneration remain unknown, but analyses of TSE-infected brain homogenates and preparations enriched for abnormal prion protein suggest that reduced levels of copper and increased levels of manganese are associated with disease. The objectives of this study were to: (1) assess copper and manganese levels in healthy and TSE-infected Syrian hamster brain homogenates; (2) determine if the distribution of these metals can be mapped in TSE-infected brain tissue using X-ray photoelectron emission microscopy (X-PEEM) with synchrotron radiation; and (3) use X-PEEM to assess the relative amounts of copper and manganese in prion plaques in situ. In agreement with studies of other TSEs and species, we found reduced brain levels of copper and increased levels of manganese associated with disease in our hamster model. We also found that the in situ levels of these metals in brainstem were sufficient to image by X-PEEM. Using immunolabeled prion plaques in directly adjacent tissue sections to identify regions to image by X-PEEM, we found a statistically significant relationship of copper-manganese dysregulation in prion plaques: copper was depleted whereas manganese was enriched. These data provide evidence for prion plaques altering local transition metal distribution in the TSE-infected central nervous system. PMID:23435237

Johnson, Christopher J.; Gilbert, P.U.P.A.; Abrecht, Mike; Baldwin, Katherine L.; Russell, Robin E.; Pedersen, Joel A.; Aiken, Judd M.; McKenzie, Debbie

2013-01-01

73

Single nucleotide polymorphisms of the prion protein gene (PRNP) in Chinese pig breeds.  

PubMed

Prion diseases (transmissible spongiform encephalopathies, TSE), as a group of fatal neurodegenerative diseases, have affected humans and a variety of other mammals. Although no natural TSE have been documented in pigs, appropriate precautions need to be taken to prevent the iatrogenic spread of prion disease through pig-to-human xenotransplantation. Polymorphisms within the open reading frame (ORF) of the single-copy gene of prion protein (PRNP) are associated with susceptibility to scrapie in sheep and variant Creutzfeldt-Jacob disease in humans. We screened polymorphisms in the PRNP gene of 64 China Experimental Minipigs and Beijing Large White pigs. Our findings suggest that the porcine PRNP gene is highly homogenous. The amino acid sequences of the mature prion protein of all samples tested were identical. Four single nucleotide polymorphisms (G11A, G615C, G684A, T726G) in the ORF of the porcine PRNP gene were found, and the G-->C nucleotide substitution resulted in a serine to asparaginate amino acid substitution at codon 4. We conclude that pigs raised under specific pathogen-free conditions, with the exclusion of rendered mammalian material for at least two generations, will have little risk of being infected with a TSE, and even less possibility of transmitting prion disease to humans through xenotransplantation. PMID:15943782

Meng, Liping; Zhao, Deming; Liu, Hongxiang; Yang, Jianmin; Ning, Zhangyong

2005-07-01

74

Location and properties of metal-binding sites on the human prion protein  

E-print Network

Location and properties of metal-binding sites on the human prion protein Graham S. Jackson*, Ian Collinge* *Medical Research Council Prion Unit, Department of Neurogenetics, Imperial College School for the prion protein, evidence for binding at affinities character- istic of authentic metal-binding proteins

Hosszu, Laszlo

75

Electrostatics in the stability and misfolding of the prion protein: salt bridges, self energy, and  

E-print Network

Electrostatics in the stability and misfolding of the prion protein: salt bridges, self energy and mutants of the prion protein. Salt bridges and self energies play key roles in stabilizing secondary and tertiary struc- tural elements of the prion protein. The total electrostatic potential energy of each

Plotkin, Steven S.

76

Thermodynamic Characterization of the Unfolding of the Prion Protein Roumita Moulick and Jayant B. Udgaonkar*  

E-print Network

Thermodynamic Characterization of the Unfolding of the Prion Protein Roumita Moulick and Jayant B ABSTRACT The prion protein appears to be unusually susceptible to conformational change, and unlike nearly of the mouse prion protein (moPrP), the full-length moPrP (23­231) and the structured C-terminal domain, mo

77

Early Intermediate in Human Prion Protein Folding As Evidenced by Ultrarapid Mixing Experiments  

E-print Network

Early Intermediate in Human Prion Protein Folding As Evidenced by Ultrarapid Mixing ExperimentsPC -to-PrPSc conversion is to elucidate the folding pathway(s) of the prion protein. On the basis of stopped-flow measurements, we recently proposed that the prion protein folds via a transient intermediate

Roder, Heinrich

78

NMR structures of three single-residue variants of the human prion protein  

E-print Network

NMR structures of three single-residue variants of the human prion protein Luigi Calzolai single-amino acid variants of the C-terminal domain of the human prion protein, hPrP(121 ``protein X,'' and it is related to the species barrier for transmission of prion diseases. As expected

Riek, Roland

79

Binding of bovine prion protein to heparin: A fluorescence polarization study  

Microsoft Academic Search

Glycosaminoglycans (GAGs) are believed to be associated with prion disease pathology and also with metabolism of the prion protein. Fluorescence polarization assay (FPA) of binding between bovine recombinant prion protein (brecPrP) and heparin labelled with AlexaFluor488 was used in model experiments to study glycosaminoglycan–prion protein interaction. Heparin binding to brecPrP was a rapid reversible event which occurred under defined conditions.

Olga Andrievskaia; Zhanna Potetinova; Aru Balachandran; Klaus Nielsen

2007-01-01

80

Prion Protein Accumulation in Lipid Rafts of Mouse Aging Brain  

PubMed Central

The cellular form of the prion protein (PrPC) is a normal constituent of neuronal cell membranes. The protein misfolding causes rare neurodegenerative disorders known as transmissible spongiform encephalopathies or prion diseases. These maladies can be sporadic, genetic or infectious. Sporadic prion diseases are the most common form mainly affecting aging people. In this work, we investigate the biochemical environment in which sporadic prion diseases may develop, focusing our attention on the cell membrane of neurons in the aging brain. It is well established that with aging the ratio between the most abundant lipid components of rafts undergoes a major change: while cholesterol decreases, sphingomyelin content rises. Our results indicate that the aging process modifies the compartmentalization of PrPC. In old mice, this change favors PrPC accumulation in detergent-resistant membranes, particularly in hippocampi. To confirm the relationship between lipid content changes and PrPC translocation into detergent-resistant membranes (DRMs), we looked at PrPC compartmentalization in hippocampi from acid sphingomyelinase (ASM) knockout (KO) mice and synaptosomes enriched in sphingomyelin. In the presence of high sphingomyelin content, we observed a significant increase of PrPC in DRMS. This process is not due to higher levels of total protein and it could, in turn, favor the onset of sporadic prion diseases during aging as it increases the PrP intermolecular contacts into lipid rafts. We observed that lowering sphingomyelin in scrapie-infected cells by using fumonisin B1 led to a 50% decrease in protease-resistant PrP formation. This may suggest an involvement of PrP lipid environment in prion formation and consequently it may play a role in the onset or development of sporadic forms of prion diseases. PMID:24040215

Agostini, Federica; Dotti, Carlos G.; Perez-Canamas, Azucena; Ledesma, Maria Dolores; Benetti, Federico; Legname, Giuseppe

2013-01-01

81

Prions Ex Vivo: What Cell Culture Models Tell Us about Infectious Proteins  

PubMed Central

Prions are unconventional infectious agents that are composed of misfolded aggregated prion protein. Prions replicate their conformation by template-assisted conversion of the endogenous prion protein PrP. Templated conversion of soluble proteins into protein aggregates is also a hallmark of other neurodegenerative diseases. Alzheimer's disease or Parkinson's disease are not considered infectious diseases, although aggregate pathology appears to progress in a stereotypical fashion reminiscent of the spreading behavior ofmammalian prions. While basic principles of prion formation have been studied extensively, it is still unclear what exactly drives PrP molecules into an infectious, self-templating conformation. In this review, we discuss crucial steps in the life cycle of prions that have been revealed in ex vivo models. Importantly, the persistent propagation of prions in mitotically active cells argues that cellular processes are in place that not only allow recruitment of cellular PrP into growing prion aggregates but also enable the multiplication of infectious seeds that are transmitted to daughter cells. Comparison of prions with other protein aggregates demonstrates that not all the characteristics of prions are equally shared by prion-like aggregates. Future experiments may reveal to which extent aggregation-prone proteins associated with other neurodegenerative diseases can copy the replication strategies of prions. PMID:24282413

Krauss, Sybille

2013-01-01

82

Inherited Prion Disease A117V Is Not Simply a Proteinopathy but Produces Prions Transmissible to Transgenic Mice Expressing Homologous Prion Protein  

PubMed Central

Prions are infectious agents causing fatal neurodegenerative diseases of humans and animals. In humans, these have sporadic, acquired and inherited aetiologies. The inherited prion diseases are caused by one of over 30 coding mutations in the human prion protein (PrP) gene (PRNP) and many of these generate infectious prions as evidenced by their experimental transmissibility by inoculation to laboratory animals. However, some, and in particular an extensively studied type of Gerstmann-Sträussler-Scheinker syndrome (GSS) caused by a PRNP A117V mutation, are thought not to generate infectious prions and instead constitute prion proteinopathies with a quite distinct pathogenetic mechanism. Multiple attempts to transmit A117V GSS have been unsuccessful and typical protease-resistant PrP (PrPSc), pathognomonic of prion disease, is not detected in brain. Pathogenesis is instead attributed to production of an aberrant topological form of PrP, C-terminal transmembrane PrP (CtmPrP). Barriers to transmission of prion strains from one species to another appear to relate to structural compatibility of PrP in host and inoculum and we have therefore produced transgenic mice expressing human 117V PrP. We found that brain tissue from GSS A117V patients did transmit disease to these mice and both the neuropathological features of prion disease and presence of PrPSc was demonstrated in the brains of recipient transgenic mice. This PrPSc rapidly degraded during laboratory analysis, suggesting that the difficulty in its detection in patients with GSS A117V could relate to post-mortem proteolysis. We conclude that GSS A117V is indeed a prion disease although the relative contributions of CtmPrP and prion propagation in neurodegeneration and their pathogenetic interaction remains to be established. PMID:24086135

Asante, Emmanuel A.; Linehan, Jacqueline M.; Smidak, Michelle; Tomlinson, Andrew; Grimshaw, Andrew; Jeelani, Asif; Jakubcova, Tatiana; Hamdan, Shyma; Powell, Caroline; Brandner, Sebastian; Wadsworth, Jonathan D. F.; Collinge, John

2013-01-01

83

Surface charge of polyoxometalates modulates polymerization of the scrapie prion protein  

E-print Network

Surface charge of polyoxometalates modulates polymerization of the scrapie prion protein Holger 31, 2008 (sent for review October 13, 2008) Prions are composed solely of an alternatively folded isoform of the prion protein (PrP), designated PrPSc. N-terminally truncated PrPSc, denoted PrP 27

84

N Resonance assignment of parts of the HET-s prion protein in its amyloid form  

E-print Network

Article 13 C, 15 N Resonance assignment of parts of the HET-s prion protein in its amyloid form: amyloid, HET-s, prions, solid-state NMR Abstract The partial 15 N and 13 C solid-state NMR resonance assignment of the HET-s prion protein fragment 218­ 289 in its amyloid form is presented. It is based

Riek, Roland

85

Direct Observation of Protein Folding, Aggregation, and a Prion-like Conformational Conversion*  

E-print Network

Direct Observation of Protein Folding, Aggregation, and a Prion-like Conformational Conversion to -sheets precedes aggregation of proteins implicated in many diseases, including Alzheimer and prion, S. B. (1998) Proc. Natl. Acad. Sci. U. S. A. 95, 13363­ 13383) to account for prion infectivity

Dokholyan, Nikolay V.

86

Humic substances interfere with detection of pathogenic prion protein  

USGS Publications Warehouse

Studies examining the persistence of prions (the etiological agent of transmissible spongiform encephalopathies) in soil require accurate quantification of pathogenic prion protein (PrPTSE) extracted from or in the presence of soil particles. Here, we demonstrate that natural organic matter (NOM) in soil impacts PrPTSE detection by immunoblotting. Methods commonly used to extract PrPTSE from soils release substantial amounts of NOM, and NOM inhibited PrPTSE immunoblot signal. The degree of immunoblot interference increased with increasing NOM concentration and decreasing NOM polarity. Humic substances affected immunoblot detection of prion protein from both deer and hamsters. We also establish that after interaction with humic acid, PrPTSE remains infectious to hamsters inoculated intracerebrally, and humic acid appeared to slow disease progression. These results provide evidence for interactions between PrPTSE and humic substances that influence both accurate measurement of PrPTSE in soil and disease transmission.

Smith, Christen B.; Booth, Clarissa J.; Wadzinski, Tyler J.; Legname, Giuseppe; Chappell, Rick; Johnson, Christopher J.; Pedersen, Joel A.

2014-01-01

87

Prions, protein homeostasis, and phenotypic diversity  

E-print Network

that facilitate the adaptation of yeasts to stressful environments, and might speed the evolution of new traits neurodegenerative disease, variously known as kuru, scrapie and bovine spongiform encephalopathy, in humans, sheep, in fact, in some cases beneficial. In fungi, prions act as epigenetic elements that increase phenotypic

Lindquist, Susan

88

Concentration of disease-associated prion protein with silicon dioxide.  

PubMed

Reagents that can precipitate the disease-associated prion protein (PrP(Sc)) are vital for the development of high sensitivity tests to detect low levels of this disease marker in biological material. Here, a range of minerals are shown to precipitate both ovine cellular prion protein (PrP(C)) and ovine scrapie PrP(Sc). The precipitation of prion protein with silicon dioxide is unaffected by PrP(Sc) strain or host species and the method can be used to precipitate bovine BSE. This method can reliably concentrate protease-resistant ovine PrP(Sc) (PrP(res)) derived from 1.69 microg of brain protein from a clinically infected animal diluted into either 50 ml of buffer or 15 ml of plasma. The introduction of a SiO(2) precipitation step into the immunological detection of PrP(res) increased detection sensitivity by over 1,500-fold. Minerals such as SiO(2) are readily available, low cost reagents with generic application to the concentration of diseases-associated prion proteins. PMID:19058035

Rees, Helen C; Maddison, Ben C; Owen, Jonathan P; Whitelam, Garry C; Gough, Kevin C

2009-03-01

89

Characterization of the Prion Protein in Human Urine*  

PubMed Central

The presence of the prion protein (PrP) in normal human urine is controversial and currently inconclusive. This issue has taken a special relevance because prion infectivity has been demonstrated in urine of animals carrying experimental or naturally occurring prion diseases, but the actual presence and tissue origin of the infectious prion have not been determined. We used immunoprecipitation, one- and two-dimensional electrophoresis, and mass spectrometry to prove definitely the presence of PrP in human urine and its post-translational modifications. We show that urinary PrP (uPrP) is truncated mainly at residue 112 but also at other residues up to 122. This truncation makes uPrP undetectable with some commonly used antibodies to PrP. uPrP is glycosylated and carries an anchor which, at variance with that of cellular PrP, lacks the inositol-associated phospholipid moiety, indicating that uPrP is probably shed from the cell surface. The detailed characterization of uPrP reported here definitely proves the presence of PrP in human urine and will help determine the origin of prion infectivity in urine. PMID:20670940

Dagdanova, Ayuna; Ilchenko, Serguei; Notari, Silvio; Yang, Qiwei; Obrenovich, Mark E.; Hatcher, Kristen; McAnulty, Peter; Huang, Lequn; Zou, Wenquan; Kong, Qingzhong; Gambetti, Pierluigi; Chen, Shu G.

2010-01-01

90

NMR solution structure of the human prion protein  

Microsoft Academic Search

The NMR structures of the recombinant human prion protein, hPrP(23-230), and two C-terminal fragments, hPrP(90-230) and hPrP(121-230), include a globular domain extending from residues 125-228, for which a detailed structure was obtained, and an N-terminal flexibly disordered \\

Ralph Zahn; Aizhuo Liu; Thorsten Lührs; Roland Riek; Christine von Schroetter; Francisco López García; Martin Billeter; Luigi Calzolai; Gerhard Wider; Kurt Wüthrich

2000-01-01

91

Pharmacological prion protein silencing accelerates central nervous system autoimmune disease via T cell receptor signalling  

PubMed Central

The primary biological function of the endogenous cellular prion protein has remained unclear. We investigated its biological function in the generation of cellular immune responses using cellular prion protein gene-specific small interfering ribonucleic acid in vivo and in vitro. Our results were confirmed by blocking cellular prion protein with monovalent antibodies and by using cellular prion protein-deficient and -transgenic mice. In vivo prion protein gene-small interfering ribonucleic acid treatment effects were of limited duration, restricted to secondary lymphoid organs and resulted in a 70% reduction of cellular prion protein expression in leukocytes. Disruption of cellular prion protein signalling augmented antigen-specific activation and proliferation, and enhanced T cell receptor signalling, resulting in zeta-chain-associated protein-70 phosphorylation and nuclear factor of activated T cells/activator protein 1 transcriptional activity. In vivo prion protein gene-small interfering ribonucleic acid treatment promoted T cell differentiation towards pro-inflammatory phenotypes and increased survival of antigen-specific T cells. Cellular prion protein silencing with small interfering ribonucleic acid also resulted in the worsening of actively induced and adoptively transferred experimental autoimmune encephalomyelitis. Finally, treatment of myelin basic protein1–11 T cell receptor transgenic mice with prion protein gene-small interfering ribonucleic acid resulted in spontaneous experimental autoimmune encephalomyelitis. Thus, central nervous system autoimmune disease was modulated at all stages of disease: the generation of the T cell effector response, the elicitation of T effector function and the perpetuation of cellular immune responses. Our findings indicate that cellular prion protein regulates T cell receptor-mediated T cell activation, differentiation and survival. Defects in autoimmunity are restricted to the immune system and not the central nervous system. Our data identify cellular prion protein as a regulator of cellular immunological homoeostasis and suggest cellular prion protein as a novel potential target for therapeutic immunomodulation. PMID:20145049

Hu, Wei; Nessler, Stefan; Hemmer, Bernhard; Eagar, Todd N.; Kane, Lawrence P.; Leliveld, S. Rutger; Muller-Schiffmann, Andreas; Gocke, Anne R.; Lovett-Racke, Amy; Ben, Li-Hong; Hussain, Rehana Z.; Breil, Andreas; Elliott, Jeffrey L.; Puttaparthi, Krishna; Cravens, Petra D.; Singh, Mahendra P.; Petsch, Benjamin; Stitz, Lothar; Racke, Michael K.

2010-01-01

92

Pharmacological prion protein silencing accelerates central nervous system autoimmune disease via T cell receptor signalling.  

PubMed

The primary biological function of the endogenous cellular prion protein has remained unclear. We investigated its biological function in the generation of cellular immune responses using cellular prion protein gene-specific small interfering ribonucleic acid in vivo and in vitro. Our results were confirmed by blocking cellular prion protein with monovalent antibodies and by using cellular prion protein-deficient and -transgenic mice. In vivo prion protein gene-small interfering ribonucleic acid treatment effects were of limited duration, restricted to secondary lymphoid organs and resulted in a 70% reduction of cellular prion protein expression in leukocytes. Disruption of cellular prion protein signalling augmented antigen-specific activation and proliferation, and enhanced T cell receptor signalling, resulting in zeta-chain-associated protein-70 phosphorylation and nuclear factor of activated T cells/activator protein 1 transcriptional activity. In vivo prion protein gene-small interfering ribonucleic acid treatment promoted T cell differentiation towards pro-inflammatory phenotypes and increased survival of antigen-specific T cells. Cellular prion protein silencing with small interfering ribonucleic acid also resulted in the worsening of actively induced and adoptively transferred experimental autoimmune encephalomyelitis. Finally, treatment of myelin basic protein(1-11) T cell receptor transgenic mice with prion protein gene-small interfering ribonucleic acid resulted in spontaneous experimental autoimmune encephalomyelitis. Thus, central nervous system autoimmune disease was modulated at all stages of disease: the generation of the T cell effector response, the elicitation of T effector function and the perpetuation of cellular immune responses. Our findings indicate that cellular prion protein regulates T cell receptor-mediated T cell activation, differentiation and survival. Defects in autoimmunity are restricted to the immune system and not the central nervous system. Our data identify cellular prion protein as a regulator of cellular immunological homoeostasis and suggest cellular prion protein as a novel potential target for therapeutic immunomodulation. PMID:20145049

Hu, Wei; Nessler, Stefan; Hemmer, Bernhard; Eagar, Todd N; Kane, Lawrence P; Leliveld, S Rutger; Müller-Schiffmann, Andreas; Gocke, Anne R; Lovett-Racke, Amy; Ben, Li-Hong; Hussain, Rehana Z; Breil, Andreas; Elliott, Jeffrey L; Puttaparthi, Krishna; Cravens, Petra D; Singh, Mahendra P; Petsch, Benjamin; Stitz, Lothar; Racke, Michael K; Korth, Carsten; Stüve, Olaf

2010-02-01

93

Folding kinetics of the human prion protein probed by temperature jump  

E-print Network

Folding kinetics of the human prion protein probed by temperature jump Tanya Harta , Laszlo L. P. Clarkea,1 aMedical Research Council Prion Unit, Institute of Neurology, Queen Square, London WC1N 3BG) Temperature-jump perturbation was used to examine the relax- ation kinetics of folding of the human prion

Hosszu, Laszlo

94

Increasing Prion Propensity by Hydrophobic Insertion  

PubMed Central

Prion formation involves the conversion of proteins from a soluble form into an infectious amyloid form. Most yeast prion proteins contain glutamine/asparagine-rich regions that are responsible for prion aggregation. Prion formation by these domains is driven primarily by amino acid composition, not primary sequence, yet there is a surprising disconnect between the amino acids thought to have the highest aggregation propensity and those that are actually found in yeast prion domains. Specifically, a recent mutagenic screen suggested that both aromatic and non-aromatic hydrophobic residues strongly promote prion formation. However, while aromatic residues are common in yeast prion domains, non-aromatic hydrophobic residues are strongly under-represented. Here, we directly test the effects of hydrophobic and aromatic residues on prion formation. Remarkably, we found that insertion of as few as two hydrophobic residues resulted in a multiple orders-of-magnitude increase in prion formation, and significant acceleration of in vitro amyloid formation. Thus, insertion or deletion of hydrophobic residues provides a simple tool to control the prion activity of a protein. These data, combined with bioinformatics analysis, suggest a limit on the number of strongly prion-promoting residues tolerated in glutamine/asparagine-rich domains. This limit may explain the under-representation of non-aromatic hydrophobic residues in yeast prion domains. Prion activity requires not only that a protein be able to form prion fibers, but also that these fibers be cleaved to generate new independently-segregating aggregates to offset dilution by cell division. Recent studies suggest that aromatic residues, but not non-aromatic hydrophobic residues, support the fiber cleavage step. Therefore, we propose that while both aromatic and non-aromatic hydrophobic residues promote prion formation, aromatic residues are favored in yeast prion domains because they serve a dual function, promoting both prion formation and chaperone-dependent prion propagation. PMID:24586661

Petri, Michelina; Flores, Noe; Rogge, Ryan A.; Cascarina, Sean M.; Ross, Eric D.

2014-01-01

95

Role of the prion protein family in the gonads  

PubMed Central

The prion-gene family comprises four members named PRNP (PRPc), PRND (Doppel), PRNT (PRT), and SPRN (Shadoo). According to species, PRND is located 16–52 kb downstream from the PRNP locus, whereas SPRN is located on another chromosome. The fourth prion-family gene, PRNT, belongs to the same genomic cluster as PRNP and PRND in humans and bovidae. PRNT and PRND possibly resulted from a duplication event of PRND and PRNP, respectively, that occurred early during eutherian species divergence. Although most of the studies concerning the prion-family has been done on PRPc and its involvement in transmissible neurodegenerative disorders, different works report some potential roles of these proteins in the reproductive function of both sexes. Among them, a clear role of PRND, that encodes for the Doppel protein, in male fertility has been demonstrated through gene targeting studies in mice. In other species, Doppel seems to play a role in testis and ovary development but its cellular localization is variable according to the gonadal developmental stage and to the mammalian species considered. For the other three genes, their roles in reproductive function appear ill-defined and/or controversial. The present review aimed to synthesize all the available data on these prion-family members and their relations with reproductive processes, mainly in the gonad of both sexes. PMID:25364761

Allais-Bonnet, Aurelie; Pailhoux, Eric

2014-01-01

96

Kinetics of Ozone Inactivation of Infectious Prion Protein  

PubMed Central

The kinetics of ozone inactivation of infectious prion protein (PrPSc, scrapie 263K) was investigated in ozone-demand-free phosphate-buffered saline (PBS). Diluted infectious brain homogenates (IBH) (0.01%) were exposed to a predetermined ozone dose (10.8 ± 2.0 mg/liter) at three pHs (pH 4.4, 6.0, and 8.0) and two temperatures (4°C and 20°C). The inactivation of PrPSc was quantified by determining the in vitro destruction of PrPSc templating properties using the protein misfolding cyclic amplification (PMCA) assay and bioassay, which were shown to correlate well. The inactivation kinetics were characterized by both Chick-Watson (CW) and efficiency factor Hom (EFH) models. It was found that the EFH model fit the experimental data more appropriately. The efficacy of ozone inactivation of PrPSc was both pH and temperature dependent. Based on the EFH model, CT (disinfectant concentration multiplied by contact time) values were determined for 2-log10, 3-log10, and 4-log10 inactivation at the conditions under which they were achieved. Our results indicated that ozone is effective for prion inactivation in ozone-demand-free water and may be applied for the inactivation of infectious prion in prion-contaminated water and wastewater. PMID:23416994

Ding, Ning; Price, Luke M.; Braithwaite, Shannon L.; Balachandran, Aru; Mitchell, Gordon; Belosevic, Miodrag

2013-01-01

97

Prion protein degradation by lichens of the genus Cladonia  

USGS Publications Warehouse

It has recently been discovered that lichens contain a serine protease capable of degrading the pathogenic prion protein, the etiological agent of prion diseases such as sheep scrapie and cervid chronic wasting disease. Limited methods are available to degrade or inactivate prion disease agents, especially in the environment, and lichens or their serine protease could prove important for management of these diseases. Scant information is available regarding the presence or absence of the protease responsible for degrading prion protein (PrP) in lichen species and, in this study, we tested the hypothesis that PrP degradation activity in lichens is phylogenetically-based by testing 44 species of Cladonia lichens, a genus for which a significant portion of the phylogeny is well established. We categorized PrP degradation activity among the 44 species (high, moderate, low or none) and found that activity in Cladonia species did not correspond with phylogenetic position of the species. Degradation of PrP did correspond, however, with three classical taxonomic characters within the genus: species with brown apothecia, no usnic acid, and the presence of a cortex. Of the 44 species studied, 18 (41%) had either high or moderate PrP degradation activity, suggesting the protease may be frequent in this genus of lichens.

Bennett, James P.; Rodriguez, Cynthia M.; Johnson, Christopher J.

2012-01-01

98

Electrostatics in the Stability and Misfolding of the Prion Protein: Salt Bridges, Self-Energy, and Solvation  

E-print Network

Using a recently developed mesoscopic theory of protein dielectrics, we have calculated the salt bridge energies, total residue electrostatic potential energies, and transfer energies into a low dielectric amyloid-like phase for 12 species and mutants of the prion protein. Salt bridges and self energies play key roles in stabilizing secondary and tertiary structural elements of the prion protein. The total electrostatic potential energy of each residue was found to be invariably stabilizing. Residues frequently found to be mutated in familial prion disease were among those with the largest electrostatic energies. The large barrier to charged group desolvation imposes regional constraints on involvement of the prion protein in an amyloid aggregate, resulting in an electrostatic amyloid recruitment pro?le that favours regions of sequence between alpha helix 1 and beta strand 2, the middles of helices 2 and 3, and the region N-terminal to alpha helix 1. We found that the stabilization due to salt bridges is minimal among the proteins studied for disease-susceptible human mutants of prion protein.

Will Guest; Neil R. Cashman; Steven S. Plotkin

2010-04-09

99

Quaternary Structure of Pathological Prion Protein as a Determining Factor of Strain-Specific Prion Replication Dynamics  

PubMed Central

Prions are proteinaceous infectious agents responsible for fatal neurodegenerative diseases in animals and humans. They are essentially composed of PrPSc, an aggregated, misfolded conformer of the ubiquitously expressed host-encoded prion protein (PrPC). Stable variations in PrPSc conformation are assumed to encode the phenotypically tangible prion strains diversity. However the direct contribution of PrPSc quaternary structure to the strain biological information remains mostly unknown. Applying a sedimentation velocity fractionation technique to a panel of ovine prion strains, classified as fast and slow according to their incubation time in ovine PrP transgenic mice, has previously led to the observation that the relationship between prion infectivity and PrPSc quaternary structure was not univocal. For the fast strains specifically, infectivity sedimented slowly and segregated from the bulk of proteinase-K resistant PrPSc. To carefully separate the respective contributions of size and density to this hydrodynamic behavior, we performed sedimentation at the equilibrium and varied the solubilization conditions. The density profile of prion infectivity and proteinase-K resistant PrPSc tended to overlap whatever the strain, fast or slow, leaving only size as the main responsible factor for the specific velocity properties of the fast strain most infectious component. We further show that this velocity-isolable population of discrete assemblies perfectly resists limited proteolysis and that its templating activity, as assessed by protein misfolding cyclic amplification outcompetes by several orders of magnitude that of the bulk of larger size PrPSc aggregates. Together, the tight correlation between small size, conversion efficiency and duration of disease establishes PrPSc quaternary structure as a determining factor of prion replication dynamics. For certain strains, a subset of PrP assemblies appears to be the best template for prion replication. This has important implications for fundamental studies on prions. PMID:24130496

Chapuis, Jérôme; Sibille, Pierre; Herzog, Laetitia; Reine, Fabienne; Jaumain, Emilie; Laude, Hubert; Rezaei, Human; Béringue, Vincent

2013-01-01

100

Gingerol prevents prion protein-mediated neuronal toxicity by regulating HIF prolyl hydroxylase 2 and prion protein  

PubMed Central

Prion diseases are a family of progressive neurodegenerative disorders, which are fatal in the majority of cases and affect both humans and domestic animals. Prion protein (PrP) (106–126) retains the neurotoxic properties of the entire pathological PrPsc and it is generally used as a reasonable model to study the mechanisms responsible for prion diseases. In our previous studies, we demonstrated that hypoxia-inducible factor (HIF)-1? is involved in the gingerol-mediated protection of neuronal cells. HIF mediates cellular adaptations to low oxygen. Prolyl hydroxylase domain-containing protein 2 (PHD2) is an oxygen sensor that hydroxylates the HIF-?-subunit, promoting its proteasomal degradation under normoxic conditions. Thus, in the present study we wished to determine whether gingerol inhibits the catalytic activity of PHD2 and prevents HIF-1? protein proteasomal degradation, thereby preventing the occurrence of PrP (106–126)-induced neuronal apoptosis. We used the pharmacological inhibition of PHD2 by dimethyloxalylglycine (DMOG) or deferoxamine (DFO) and the genetic inhibition of HIF-1? by HIF-1? small interfering RNA (siRNA) to block the effects of gingerol against PrP (106–126)-induced neurotoxicity. Our results demonstrated that gingerol prevented PrP (106–126)-induced neuronal apoptosis by upregulating HIF-1? and inhibiting the catalytic activity of PHD2 under normoxic conditions. Moreover, the protective effects of gingerol against PrP (106–126)-induced neuronal apoptosis were associated with the upregulation of the expression of cellular prion protein (PrPc). In conclusion, our results indicate that gingerol has therapeutic potential for use in the treatment or prevention of prion diseases, and its inhibitory effects on the catalytic activity of PHD2 may be of clinical benefit. PMID:25231392

PARK, YANG-GYU; PARK, SANG-YOUEL

2014-01-01

101

Absence of the prion protein homologue Doppel causes male sterility  

PubMed Central

The agent that causes prion diseases is thought to be identical with PrPSc, a conformer of the normal prion protein PrPC. PrPC-deficient mice do not exhibit major pathologies, perhaps because they express a protein termed Dpl, which shares significant biochemical and structural homology with PrPC. To investigate the physiological function of Dpl, we generated mice harbouring a homozygous disruption of the Prnd gene that encodes Dpl. Dpl deficiency did not interfere with embryonic and postnatal development, but resulted in male sterility. Dpl protein was expressed at late stages of spermiogenesis, and spermatids of Dpl mutants were reduced in numbers, immobile, malformed and unable to fertilize oocytes in vitro. Mechanical dissection of the zona pellucida partially restored in vitro fertilization. We conclude that Dpl regulates male fertility by controlling several aspects of male gametogenesis and sperm–egg interaction. PMID:12110578

Behrens, Axel; Genoud, Nicolas; Naumann, Heike; Rulicke, Thomas; Janett, Fredi; Heppner, Frank L.; Ledermann, Birgit; Aguzzi, Adriano

2002-01-01

102

Functional Diversification of Hsp40: Distinct J-Protein Functional Requirements for Two Prions Allow for Chaperone-Dependent Prion Selection  

PubMed Central

Yeast prions are heritable amyloid aggregates of functional yeast proteins; their propagation to subsequent cell generations is dependent upon fragmentation of prion protein aggregates by molecular chaperone proteins. Mounting evidence indicates the J-protein Sis1 may act as an amyloid specificity factor, recognizing prion and other amyloid aggregates and enabling Ssa and Hsp104 to act in prion fragmentation. Chaperone interactions with prions, however, can be affected by variations in amyloid-core structure resulting in distinct prion variants or ‘strains’. Our genetic analysis revealed that Sis1 domain requirements by distinct variants of [PSI+] are strongly dependent upon overall variant stability. Notably, multiple strong [PSI+] variants can be maintained by a minimal construct of Sis1 consisting of only the J-domain and glycine/phenylalanine-rich (G/F) region that was previously shown to be sufficient for cell viability and [RNQ+] prion propagation. In contrast, weak [PSI+] variants are lost under the same conditions but maintained by the expression of an Sis1 construct that lacks only the G/F region and cannot support [RNQ+] propagation, revealing mutually exclusive requirements for Sis1 function between these two prions. Prion loss is not due to [PSI+]-dependent toxicity or dependent upon a particular yeast genetic background. These observations necessitate that Sis1 must have at least two distinct functional roles that individual prions differentially require for propagation and which are localized to the glycine-rich domains of the Sis1. Based on these distinctions, Sis1 plasmid-shuffling in a [PSI+]/[RNQ+] strain permitted J-protein-dependent prion selection for either prion. We also found that, despite an initial report to the contrary, the human homolog of Sis1, Hdj1, is capable of [PSI+] prion propagation in place of Sis1. This conservation of function is also prion-variant dependent, indicating that only one of the two Sis1-prion functions may have been maintained in eukaryotic chaperone evolution. PMID:25058638

Patel, Milan J.; Sporn, Zachary A.; Hines, Justin K.

2014-01-01

103

Functional diversification of hsp40: distinct j-protein functional requirements for two prions allow for chaperone-dependent prion selection.  

PubMed

Yeast prions are heritable amyloid aggregates of functional yeast proteins; their propagation to subsequent cell generations is dependent upon fragmentation of prion protein aggregates by molecular chaperone proteins. Mounting evidence indicates the J-protein Sis1 may act as an amyloid specificity factor, recognizing prion and other amyloid aggregates and enabling Ssa and Hsp104 to act in prion fragmentation. Chaperone interactions with prions, however, can be affected by variations in amyloid-core structure resulting in distinct prion variants or 'strains'. Our genetic analysis revealed that Sis1 domain requirements by distinct variants of [PSI+] are strongly dependent upon overall variant stability. Notably, multiple strong [PSI+] variants can be maintained by a minimal construct of Sis1 consisting of only the J-domain and glycine/phenylalanine-rich (G/F) region that was previously shown to be sufficient for cell viability and [RNQ+] prion propagation. In contrast, weak [PSI+] variants are lost under the same conditions but maintained by the expression of an Sis1 construct that lacks only the G/F region and cannot support [RNQ+] propagation, revealing mutually exclusive requirements for Sis1 function between these two prions. Prion loss is not due to [PSI+]-dependent toxicity or dependent upon a particular yeast genetic background. These observations necessitate that Sis1 must have at least two distinct functional roles that individual prions differentially require for propagation and which are localized to the glycine-rich domains of the Sis1. Based on these distinctions, Sis1 plasmid-shuffling in a [PSI+]/[RNQ+] strain permitted J-protein-dependent prion selection for either prion. We also found that, despite an initial report to the contrary, the human homolog of Sis1, Hdj1, is capable of [PSI+] prion propagation in place of Sis1. This conservation of function is also prion-variant dependent, indicating that only one of the two Sis1-prion functions may have been maintained in eukaryotic chaperone evolution. PMID:25058638

Harris, Julia M; Nguyen, Phil P; Patel, Milan J; Sporn, Zachary A; Hines, Justin K

2014-07-01

104

Identification of a Protein that Purifies with the Scrapie Prion  

NASA Astrophysics Data System (ADS)

Purification of prions from scrapie-infected hamster brain yielded a protein that was not found in a similar fraction from uninfected brain. The protein migrated with an apparent molecular size of 27,000 to 30,000 daltons in sodium dodecyl sulfate polyacrylamide gels. The resistance of this protein to digestion by proteinase K distinguished it from proteins of similar molecular weight found in normal hamster brain. Initial results suggest that the amount of this protein correlates with the titer of the agent.

Bolton, David C.; McKinley, Michael P.; Prusiner, Stanley B.

1982-12-01

105

Resistance to chronic wasting disease in transgenic mice expressing a naturally occurring allelic variant of deer prion protein  

Microsoft Academic Search

Prion protein (PrP) is a required factor for susceptibility to transmissible spongiform encephalopathy or prion diseases. In transgenic mice, expression of prion protein (PrP) from another species often confers susceptibility to prion disease from that donor species. For example, expression of deer or elk PrP in transgenic mice has induced susceptibility to chronic wasting disease (CWD), the prion disease of

Kimberly Meade-White; Brent Race; Matthew Trifilo; Alex Bossers; Cynthia Favara; Rachel Lacasse; Michael Miller; Elizabeth Williams; Michael Oldstone; Richard Race; Bruce Chesebro

2007-01-01

106

Neurotoxic Mutants of the Prion Protein Induce Spontaneous Ionic Currents in Cultured Cells*  

PubMed Central

The mechanisms by which prions kill neurons and the role of the cellular prion protein in this process are enigmatic. Insight into these questions is provided by the neurodegenerative phenotypes of transgenic mice expressing prion protein (PrP) molecules with deletions of conserved amino acids in the central region. We report here that expression in transfected cells of the most toxic of these PrP deletion mutants (?105–125) induces large, spontaneous ionic currents that can be detected by patch-clamping techniques. These currents are produced by relatively non-selective, cation-permeable channels or pores in the cell membrane and can be silenced by overexpression of wild-type PrP, as well as by treatment with a sulfated glycosaminoglycan. Similar currents are induced by PrP molecules carrying several different point mutations in the central region that cause familial prion diseases in humans. The ionic currents described here are distinct from those produced in artificial lipid membranes by synthetic peptides derived from the PrP sequence because they are induced by membrane-anchored forms of PrP that are synthesized by cells and that are found in vivo. Our results indicate that the neurotoxicity of some mutant forms of PrP is attributable to enhanced ion channel activity and that wild-type PrP possesses a channel-silencing activity. Drugs that block PrP-associated channels or pores may therefore represent novel therapeutic agents for treatment of patients with prion diseases. PMID:20573963

Solomon, Isaac H.; Huettner, James E.; Harris, David A.

2010-01-01

107

Fragmentation and dimerization of copper-loaded prion protein by copper-catalysed oxidation  

PubMed Central

Prion protein consists of an N-terminal domain containing a series of octapeptide repeats with the consensus sequence PHGGGWGQ and a C-terminal domain composed of three ?-helices and two short ?-strands. Several studies have shown that the N-terminal domain binds five Cu2+ ions. In the present study, we have investigated copper-catalysed oxidation of a recombinant mouse prion protein, PrP23–231. The copper-loaded PrP23–231 was found to be carbonylated by incubation with dopamine. Besides the formation of carbonyls, a cross-linked species with the dimeric size and C-terminally truncated species were generated. These reactions were retarded in the presence of Cu+- and Cu2+-specific copper chelators, catalase, and SOD (superoxide dismutase), but not in the presence of various bivalent metal ions. Together, these results indicate that the copper bound to prion protein undergoes catalytic cycling in the presence of catecholamines and causes the oxidation of the protein. PMID:15554874

Shiraishi, Noriyuki; Inai, Yoko; Bi, Wenxiang; Nishikimi, Morimitsu

2004-01-01

108

Higher susceptibility to amyloid fibril formation of the recombinant ovine prion protein modified by transglutaminase.  

PubMed

Prion proteins are known as the main agents of transmissible spongiform encephalopathies affecting humans as well as animals. A recombinant ovine prion protein was found to be in vitro able to act as an effective substrate for a microbial isoform of transglutaminase, an enzyme catalyzing the formation of isopeptide bonds inside the proteins. We proved that transglutaminase modifies the structure of the prion protein by leading to the formation of three intra-molecular crosslinks and that the crosslinked protein form is more competent in amyloid formation compared to the unmodified one. In addition, the crosslinked prion protein was shown also to be more resistant to proteinase K digestion. Our findings suggest a possible use of transglutaminase in stabilizing the prion protein three-dimensional structure in order to investigate the molecular basis of the conversion of the protein into its pathological form. PMID:22705206

Sorrentino, Angela; Giosafatto, Concetta Valeria L; Sirangelo, Ivana; De Simone, Carmela; Di Pierro, Prospero; Porta, Raffaele; Mariniello, Loredana

2012-10-01

109

Molecular Dynamics Simulations on the Oligomer-Formation Process of the GNNQQNY Peptide from Yeast Prion Protein Sup35  

E-print Network

Prion Protein Sup35 Zhuqing Zhang, Hao Chen, Hongjun Bai, and Luhua Lai Beijing National Laboratory prion-like protein Sup35 as a model system, for which a detailed atomic structure of the fibril formed

Luhua, Lai

110

A Variational Model for Oligomer-Formation Process of GNNQQNY Peptide from Yeast Prion Protein Sup35  

E-print Network

A Variational Model for Oligomer-Formation Process of GNNQQNY Peptide from Yeast Prion Protein Sup, the peptide GNNQQNY from yeast prion protein Sup35. By examining the free energy surface, we identified

Zhang, Yang

111

Sensitive detection of pathological prion protein by cyclic amplification of protein misfolding  

Microsoft Academic Search

Prions are the infectious agents responsible for transmissible spongiform encephalopathies. The principal component of prions is the glycoprotein PrPSc, which is a conformationally modified isoform of a normal cell-surface protein called PrPC (ref. 1). During the time between infection and the appearance of the clinical symptoms, minute amounts of PrPSc replicate by conversion of host PrPC, generating large amounts of

Gabriela P. Saborio; Bruno Permanne; Claudio Soto

2001-01-01

112

The tip of the iceberg: RNA-binding proteins with prion-like domains in neurodegenerative disease  

E-print Network

Ã?Ã? Ã? Ã?Ã?Ã? Ã? Ã?Ã? The tip of the iceberg: RNA-binding proteins with prion-like domains of the iceberg: RNA-binding proteins with prion-like domains in neurodegenerative disease, Brain Research (2012 MANUSCRIPT ACCEPTED MANUSCRIPT 1 The tip of the iceberg: RNA-binding proteins with prion-like domains

Shorter, James

113

NMR structure of the bovine prion protein Francisco Lo pez Garcia, Ralph Zahn, Roland Riek, and Kurt Wu thrich*  

E-print Network

NMR structure of the bovine prion protein Francisco Lo´ pez Garci´a, Ralph Zahn, Roland Riek structures of the recombinant 217-residue polypeptide chain of the mature bovine prion protein, bPrP(23 there are characteristic local differences relative to the confor- mations of the murine and Syrian hamster prion proteins

Riek, Roland

114

Dissection of Conformational Conversion Events during Prion Amyloid Fibril  

E-print Network

Dissection of Conformational Conversion Events during Prion Amyloid Fibril Formation Using Hydrogen A molecular understanding of prion diseases requires an understanding of the mechanism of amyloid fibril formation by the prion protein. In particular, it is necessary to define the sequence of the structural

115

Prion-like transmission of protein aggregates in neurodegenerative diseases  

PubMed Central

Neurodegenerative diseases are commonly associated with the accumulation of intracellular or extracellular protein aggregates. Recent studies suggest that these aggregates are capable of crossing cellular membranes and can directly contribute to the propagation of neurodegenerative disease pathogenesis. We propose that, once initiated, neuropathological changes might spread in a ‘prion-like’ manner and that disease progression is associated with the intercellular transfer of pathogenic proteins. The transfer of naked infectious particles between cells could therefore be a target for new disease-modifying therapies. PMID:20308987

Brundin, Patrik; Melki, Ronald; Kopito, Ron

2010-01-01

116

Two Mutant Prion Proteins Expressed in Cultured Cells Acquire Biochemical Properties Reminiscent of the Scrapie Isoform  

Microsoft Academic Search

Prion diseases are a group of fatal neurodegenerative disorders that are unique in being infectious, genetic, and sporadic in origin. Infectious cases are caused by prions, which are composed primarily of PrPSc, a posttranslationally modified isoform of the normal cellular prion protein PrPC. Inherited cases are linked to insertional or point mutations in the host gene encoding PrPC. To investigate

Sylvain Lehmann; David A. Harris

1996-01-01

117

Identification of scrapie prion protein-specific mRNA in scrapie-infected and uninfected brain  

Microsoft Academic Search

To date no nucleic acid has been found in the purified infectious agent which causes the spongiform encephalopathy known as scrapie. In an attempt to identify a unique scrapie virus-associated messenger RNA in tissues of infected animals, we have synthesized an oligonucleotide probe complementary to the mRNA sequence corresponding to the amino-acid sequence of the prion protein, PrP27-30 (ref. 1).

Bruce Chesebro; Richard Race; Kathy Wehrly; Jane Nishio; Marshall Bloom; David Lechner; Sven Bergstrom; Ken Robbins; Leonard Mayer; Jerry M. Keith; Claude Garon; Ashley Haase

1985-01-01

118

Classifying prion and prion-like phenomena.  

PubMed

The universe of prion and prion-like phenomena has expanded significantly in the past several years. Here, we overview the challenges in classifying this data informatically, given that terms such as "prion-like", "prion-related" or "prion-forming" do not have a stable meaning in the scientific literature. We examine the spectrum of proteins that have been described in the literature as forming prions, and discuss how "prion" can have a range of meaning, with a strict definition being for demonstration of infection with in vitro-derived recombinant prions. We suggest that although prion/prion-like phenomena can largely be apportioned into a small number of broad groups dependent on the type of transmissibility evidence for them, as new phenomena are discovered in the coming years, a detailed ontological approach might be necessary that allows for subtle definition of different "flavors" of prion / prion-like phenomena. PMID:24549098

Harbi, Djamel; Harrison, Paul M

2014-01-01

119

Manganese Upregulates Cellular Prion Protein and Contributes to Altered Stabilization and Proteolysis: Relevance to Role of Metals in Pathogenesis of Prion Disease  

PubMed Central

Prion diseases are fatal neurodegenerative diseases resulting from misfolding of normal cellular prion (PrPC) into an abnormal form of scrapie prion (PrPSc). The cellular mechanisms underlying the misfolding of PrPC are not well understood. Since cellular prion proteins harbor divalent metal-binding sites in the N-terminal region, we examined the effect of manganese on PrPC processing in in vitro models of prion disease. Exposure to manganese significantly increased PrPC levels both in cytosolic and in membrane-rich fractions in a time-dependent manner. Manganese-induced PrPC upregulation was independent of messenger RNA transcription or stability. Additionally, manganese treatment did not alter the PrPC degradation by either proteasomal or lysosomal pathways. Interestingly, pulse-chase analysis showed that the PrPC turnover rate was significantly altered with manganese treatment, indicating increased stability of PrPC with the metal exposure. Limited proteolysis studies with proteinase-K further supported that manganese increases the stability of PrPC. Incubation of mouse brain slice cultures with manganese also resulted in increased prion protein levels and higher intracellular manganese accumulation. Furthermore, exposure of manganese to an infectious prion cell model, mouse Rocky Mountain Laboratory–infected CAD5 cells, significantly increased prion protein levels. Collectively, our results demonstrate for the first time that divalent metal manganese can alter the stability of prion proteins and suggest that manganese-induced stabilization of prion protein may play a role in prion protein misfolding and prion disease pathogenesis. PMID:20176619

Choi, Christopher J.; Anantharam, Vellareddy; Martin, Dustin P.; Nicholson, Eric M.; Richt, Jurgen A.; Kanthasamy, Arthi; Kanthasamy, Anumantha G.

2010-01-01

120

Prion protein facilitates synaptic vesicle release by enhancing release probability  

PubMed Central

The cellular prion protein (PrPC) has been implicated in several neurodegenerative diseases as a result of protein misfolding. In humans, prion disease occurs typically with a sporadic origin where uncharacterized mechanisms induce spontaneous PrPC misfolding leading to neurotoxic PrP-scrapie formation (PrPSC). The consequences of misfolded PrPC signalling are well characterized but little is known about the physiological roles of PrPC and its involvement in disease. Here we investigated wild-type PrPC signalling in synaptic function as well as the effects of a disease-relevant mutation within PrPC (proline-to-leucine mutation at codon 101). Expression of wild-type PrPC at the Drosophila neuromuscular junction leads to enhanced synaptic responses as detected in larger miniature synaptic currents which are caused by enlarged presynaptic vesicles. The expression of the mutated PrPC leads to reduction of both parameters compared with wild-type PrPC. Wild-type PrPC enhances synaptic release probability and quantal content but reduces the size of the ready-releasable vesicle pool. Partially, these changes are not detectable following expression of the mutant PrPC. A behavioural test revealed that expression of either protein caused an increase in locomotor activities consistent with enhanced synaptic release and stronger muscle contractions. Both proteins were sensitive to proteinase digestion. These data uncover new functions of wild-type PrPC at the synapse with a disease-relevant mutation in PrPC leading to diminished functional phenotypes. Thus, our data present essential new information possibly related to prion pathogenesis in which a functional synaptic role of PrPC is compromised due to its advanced conversion into PrPSC thereby creating a lack-of-function scenario. PMID:24722203

Robinson, Susan W.; Nugent, Marie L.; Dinsdale, David; Steinert, Joern R.

2014-01-01

121

High CJD infectivity remains after prion protein is destroyed  

PubMed Central

The hypothesis that host prion protein (PrP) converts into an infectious prion form rests on the observation that infectivity progressively decreases in direct proportion to the decrease of PrP with proteinase K (PK) treatment. PrP that resists limited PK digestion (PrP-res, PrPsc) has been assumed to be the infectious form, with speculative types of misfolding encoding the many unique TSE agent strains. Recently, a PK sensitive form of PrP has been proposed as the prion. Thus we re-evaluated total PrP (sensitive and resistant) and used a cell-based assay for titration of infectious particles. A keratinase (NAP) known to effectively digest PrP was compared to PK. Total PrP in FU-CJD infected brain was reduced to ?0.3% in a 2hr PK digest, yet there was no reduction in titer. Remaining non-PrP proteins were easily visualized with colloidal gold in this highly infectious homogenate. In contrast to PK, NAP digestion left 0.8% residual PrP after 2hr, yet decreased titer by >2.5logs; few residual protein bands remained. FU-CJD infected cells with 10x the infectivity of brain by both animal and cell culture assays were also evaluated. NAP again significantly reduced cell infectivity (>3.5 logs). Extreme PK digestions were needed to reduce cell PrP to <0.2%, yet a very high titer of ?7.8 logs remained. Our FU-CJD brain results are in good accord with the only other report on maximal PrP digestion and titer. It is likely that one or more residual non-PrP proteins may protect agent nucleic acids in infectious particles. PMID:21793041

Miyazawa, Kohtaro; Emmerling, Kaitlin; Manuelidis, Laura

2011-01-01

122

Species barrier in prion diseases: a kinetic interpretation based on the conformational adaptation of the prion protein.  

PubMed Central

Prion diseases are thought to result from the conformational change of the normal cellular prion protein to a pathogenic protease-resistant isoform. However, brain extracts not containing the protease-resistant isoform of the prion protein can be infectious following interspecies transmission. The 'protein-only' hypothesis of pathogenesis is extended to provide possible explanations which could be interpreted in terms of a different infectious agent. It is proposed that normal cellular protein (PrPC) may be transformed into a form (PrP*) that is conformationally distinct from the host-specific abnormal isoform (PrPSc). In infection from a heterologous donor, the dimeric forms of heterologous PrPSc, which may catalyse the formation of host PrP* from PrPC, host PrP* and host PrPSc are all considered to be capable of catalysing, to some extent, the conversion of PrPC into PrPSc. However, depending on the species involved, PrP* may, or may not, be pathogenic, and may, or may not, be sensitive to proteolysis. It is shown, by numerical integration of the differential rate equations derived from this model, that a strain may be stabilized after two or three passages through a different species and that transmission might occur in the absence of detectable protease-resistant prion protein. The natural transmission of scrapie to cattle is discussed in relation to the model. PMID:9729459

Kellershohn, N; Laurent, M

1998-01-01

123

Analyses of protease resistance and aggregation state of abnormal prion protein across the spectrum of human prions.  

PubMed

Prion diseases are characterized by tissue accumulation of a misfolded, ?-sheet-enriched isoform (scrapie prion protein (PrP(Sc))) of the cellular prion protein (PrP(C)). At variance with PrP(C), PrP(Sc) shows a partial resistance to protease digestion and forms highly aggregated and detergent-insoluble polymers, two properties that have been consistently used to distinguish the two proteins. In recent years, however, the idea that PrP(Sc) itself comprises heterogeneous species has grown. Most importantly, a putative proteinase K (PK)-sensitive form of PrP(Sc) (sPrP(Sc)) is being increasingly investigated for its possible role in prion infectivity, neurotoxicity, and strain variability. The study of sPrP(Sc), however, remains technically challenging because of the need of separating it from PrP(C) without using proteases. In this study, we have systematically analyzed both PK resistance and the aggregation state of purified PrP(Sc) across the whole spectrum of the currently characterized human prion strains. The results show that PrP(Sc) isolates manifest significant strain-specific differences in their PK digestion profile that are only partially explained by differences in the size of aggregates, suggesting that other factors, likely acting on PrP(Sc) aggregate stability, determine its resistance to proteolysis. Fully protease-sensitive low molecular weight aggregates were detected in all isolates but in a limited proportion of the overall PrP(Sc) (i.e. <10%), arguing against a significant role of slowly sedimenting PK-sensitive PrP(Sc) in the biogenesis of prion strains. Finally, we highlight the limitations of current operational definitions of sPrP(Sc) and of the quantitative analytical measurements that are not based on the isolation of a fully PK-sensitive PrP(Sc) form. PMID:23897825

Saverioni, Daniela; Notari, Silvio; Capellari, Sabina; Poggiolini, Ilaria; Giese, Armin; Kretzschmar, Hans A; Parchi, Piero

2013-09-27

124

The Residue 129 Polymorphism in Human Prion Protein Does Not Confer Susceptibility to Creutzfeldt-Jakob Disease by Altering the  

E-print Network

The Residue 129 Polymorphism in Human Prion Protein Does Not Confer Susceptibility to Creutzfeldt¶ From the Medical Research Council Prion Unit, Department of Neurodegenerative Disease, Institute10 2TN, United Kingdom There are two common forms of prion protein (PrP) in humans, with either

Hosszu, Laszlo

125

Comparative genomic analysis of prion genes  

Microsoft Academic Search

BACKGROUND: The homologues of human disease genes are expected to contribute to better understanding of physiological and pathogenic processes. We made use of the present availability of vertebrate genomic sequences, and we have conducted the most comprehensive comparative genomic analysis of the prion protein gene PRNP and its homologues, shadow of prion protein gene SPRN and doppel gene PRND, and

Marko Premzl; Vera Gamulin

2007-01-01

126

Ubiquitin Ligase gp78 Targets Unglycosylated Prion Protein PrP for Ubiquitylation and Degradation  

PubMed Central

Prion protein PrP is a central player in several devastating neurodegenerative disorders, including mad cow disease and Creutzfeltd-Jacob disease. Conformational alteration of PrP into an aggregation-prone infectious form PrPSc can trigger pathogenic events. How levels of PrP are regulated is poorly understood. Human PrP is known to be degraded by the proteasome, but the specific proteolytic pathway responsible for PrP destruction remains elusive. Here, we demonstrate that the ubiquitin ligase gp78, known for its role in protein quality control, is critical for unglycosylated PrP ubiquitylation and degradation. Furthermore, C-terminal sequences of PrP protein are crucial for its ubiquitylation and degradation. Our study reveals the first ubiquitin ligase specifically involved in prion protein PrP degradation and PrP sequences crucial for its turnover. Our data may lead to a new avenue to control PrP level and pathogenesis. PMID:24714645

Cheng, Haili; Tsai, Yien Che; Weissman, Allan M.; Luo, Shiwen; Rao, Hai

2014-01-01

127

Prion protein inhibits microtubule assembly by inducing tubulin oligomerization  

SciTech Connect

A growing body of evidence points to an association of prion protein (PrP) with microtubular cytoskeleton. Recently, direct binding of PrP to tubulin has also been found. In this work, using standard light scattering measurements, sedimentation experiments, and electron microscopy, we show for First time the effect of a direct interaction between these proteins on tubulin polymerization. We demonstrate that full-length recombinant PrP induces a rapid increase in the turbidity of tubulin diluted below the critical concentration for microtubule assembly. This effect requires magnesium ions and is weakened by NaCl. Moreover, the PrP-induced light scattering structures of tubulin are cold-stable. In preparations of diluted tubulin incubated with PrP, electron microscopy revealed the presence of {approx}50 nm disc-shaped structures not reported so far. These unique tubulin oligomers may form large aggregates. The effect of PrP is more pronounced under the conditions promoting microtubule formation. In these tubulin samples, PrP induces formation of the above oligomers associated with short protofilaments and sheets of protofilaments into aggregates. Noticeably, this is accompanied by a significant reduction of the number and length of microtubules. Hence, we postulate that prion protein may act as an inhibitor of microtubule assembly by inducing formation of stable tubulin oligomers.

Nieznanski, Krzysztof [Nencki Institute of Experimental Biology, Department of Muscle Biochemistry, Warsaw (Poland)]. E-mail: k.nieznanski@nencki.gov.pl; Podlubnaya, Zoya A. [Institute of Theoretical and Experimental Biophysics, Laboratory of Structure and Function of Muscle Proteins, Pushchino (Russian Federation); Pushchino State University, Pushchino (Russian Federation); Nieznanska, Hanna [Nencki Institute of Experimental Biology, Department of Muscle Biochemistry, Warsaw (Poland)

2006-10-13

128

Prion proteins: physiological functions and role in neurological disorders.  

PubMed

Stanley Prusiner was the first to promote the concept of misfolded proteins as a cause for neurological disease. It has since been shown by him and other investigators that the scrapie isoform of prion protein (PrP(Sc)) functions as an infectious agent in numerous human and non-human disorders of the central nervous system (CNS). Interestingly, other organ systems appear to be less affected, and do not appear to lead to major co-morbidities. The physiological function of the endogenous cellular form of the prion protein (PrP(C)) is much less clear. It is intriguing that PrP(c) is expressed on most tissues in mammals, suggesting not only biological functions outside the CNS, but also a role other than the propagation of its misfolded isotype. In this review, we summarize accumulating in vitro and in vivo evidence regarding the physiological functions of PrP(C) in the nervous system, as well as in lymphoid organs. PMID:17707411

Hu, Wei; Kieseier, Bernd; Frohman, Elliot; Eagar, Todd N; Rosenberg, Roger N; Hartung, Hans-Peter; Stüve, Olaf

2008-01-15

129

Molecular Dynamics Studies on the NMR Structures of Rabbit Prion Protein Wild-type and Mutants  

E-print Network

Prion diseases are invariably fatal and highly infectious neurodegenerative diseases that affect a wide variety of mammalian species such as sheep, goats, mice, humans, chimpanzees, hamsters, cattle, elks, deers, minks, cats, chicken, pigs, turtles, etc. These neurodegenerative diseases are caused by the conversion from a soluble normal cellular protein into insoluble abnormally folded infectious prions and the conversion is believed to involve conformational change from a predominantly alpha-helical protein to one rich in beta-sheet structure. Such conformational changes may be amenable to study by molecular dynamics techniques. For rabbits, classical studies show they have a low susceptibility to be infected, but in 2012 it was reported that rabbit prion can be generated (though not directly) and the rabbit prion is infectious and transmissible (Proceedings of the National Academy of Sciences USA Volume 109 Issue 13 Pages from 5080 to 5085). This paper studies the molecular structure of rabbit prion protein...

Zhang, Jiapu

2014-01-01

130

Specificity of the J-protein Sis1 in the propagation of 3 yeast prions  

E-print Network

], are heritable elements formed by proteins capable of acquiring self-perpetuat- ing conformations Yeast prions are non-Mendelian inherited elements capable of forming self-perpetuating conformations (1 heritable traits. For propagation in the cell population, physical transmission of the prion template, often

Craig, Elizabeth A

131

Chaperone Proteins Select and Maintain [PIN+] Prion Conformations in Saccharomyces cerevisiae  

PubMed Central

Prions are proteins that can adopt different infectious conformations known as “strains” or “variants,” each with a distinct, epigenetically inheritable phenotype. Mechanisms by which prion variants are determined remain unclear. Here we use the Saccharomyces cerevisiae prion Rnq1p/[PIN+] as a model to investigate the effects of chaperone proteins upon prion variant determination. We show that deletion of specific chaperone genes alters [PIN+] variant phenotypes, including [PSI+] induction efficiency, Rnq1p aggregate morphology/size and variant dominance. Mating assays demonstrate that gene deletion-induced phenotypic changes are stably inherited in a non-Mendelian manner even after restoration of the deleted gene, confirming that they are due to a bona fide change in the [PIN+] variant. Together, our results demonstrate a role for chaperones in regulating the prion variant complement of a cell. PMID:23148221

Lancaster, David L.; Dobson, C. Melissa; Rachubinski, Richard A.

2013-01-01

132

Normal prion protein has an activity like that of superoxide dismutase.  

PubMed Central

We show here that mouse prion protein (PrP(C)) either as recombinant protein or immunoprecipitated from brain tissue has superoxide dismutase (SOD) activity. SOD activity was also associated with recombinant chicken PrP(C) confirming the evolutionary conserved phenotype suggested by sequence similarity. Acquisition of copper by PrP(C) during protein folding endowed SOD activity on the protein but the addition of copper following refolding did not. PrP(C) dependent SOD activity was abolished by deletion of the octapeptide-repeat region involved in copper binding. These results describe an enzymic function for PrP(C) consistent with its cellular distribution and suggest it has a direct role in cellular resistance to oxidative stress. PMID:10548526

Brown, D R; Wong, B S; Hafiz, F; Clive, C; Haswell, S J; Jones, I M

1999-01-01

133

An N-terminal Polybasic Domain and Cell Surface Localization Are Required for Mutant Prion Protein Toxicity*S  

E-print Network

An N-terminal Polybasic Domain and Cell Surface Localization Are Required for Mutant Prion Protein that alterations in the normal physiological activity of PrPC contribute to prion-induced neurotoxicityPSc . Prion diseases or transmissible spongiform encephalopa- thies comprise a group of fatal

Huettner, James E.

134

Accumulation of Pathological Prion Protein PrPSc in the Skin of Animals with Experimental and Natural Scrapie  

Microsoft Academic Search

Prion infectivity and its molecular marker, the pathological prion protein PrPSc, accumulate in the central nervous system and often also in lymphoid tissue of animals or humans affected by transmissible spongiform encephalopathies. Recently, PrPSc was found in tissues previously considered not to be invaded by prions (e.g., skeletal muscles). Here, we address the question of whether prions target the skin

Achim Thomzig; Walter Schulz-Schaeffer; Arne Wrede; Wilhelm Wemheuer; Bertram Brenig; Christine Kratzel; Karin Lemmer; Michael Beekes

2007-01-01

135

Mouse Prion Protein Polymorphism Phe-108/Val-189 Affects the Kinetics of Fibril Formation and the Response to Seeding  

PubMed Central

Prion diseases are fatal neurodegenerative disorders associated with the polymerization of the cellular form of prion protein (PrPC) into an amyloidogenic ?-sheet infectious form (PrPSc). The sequence of host PrP is the major determinant of host prion disease susceptibility. In mice, the presence of allele a (Prnpa, encoding the polymorphism Leu-108/Thr-189) or b (Prnpb, Phe-108/Val-189) is associated with short or long incubation times, respectively, following infection with PrPSc. The molecular bases linking PrP sequence, infection susceptibility, and convertibility of PrPC into PrPSc remain unclear. Here we show that recombinant PrPa and PrPb aggregate and respond to seeding differently in vitro. Our kinetic studies reveal differences during the nucleation phase of the aggregation process, where PrPb exhibits a longer lag phase that cannot be completely eliminated by seeding the reaction with preformed fibrils. Additionally, PrPb is more prone to propagate features of the seeds, as demonstrated by conformational stability and electron microscopy studies of the formed fibrils. We propose a model of polymerization to explain how the polymorphisms at positions 108 and 189 produce the phenotypes seen in vivo. This model also provides insight into phenomena such as species barrier and prion strain generation, two phenomena also influenced by the primary structure of PrP. PMID:23283973

Cortez, Leonardo M.; Kumar, Jitendra; Renault, Ludovic; Young, Howard S.; Sim, Valerie L.

2013-01-01

136

Glycan chains modulate prion protein binding to immobilized metal ions.  

PubMed

PrP(c) is the normal isoform of the prion protein which can be converted into PrP(Sc), the pathology-associated conformer in prion diseases. It contains two N-linked glycan chains attached to the C-proximal globular domain. While the biological functions of PrP(c) are still unknown, its ability to bind Cu(2+) is well documented. The main Cu(2+)-binding sites are located in the N-proximal, unstructured region of the molecule. Here we report that PrP(c) glycans influence the capacity of PrP(c) from sheep brain or cultured Rov cells to bind IMAC columns loaded with Cu(2+) or Co(2+). Using different anti-PrP antibodies and PrP(c) glycosylation mutants, we show that the full length non-glycosylated form of PrP(c) has a higher binding efficiency for column-bound Cu(2+) and Co(2+) than the corresponding glycosylated form. Our findings raise the possibility that the accessibility of the PrP(c) metal ion-binding sites might be controlled by the glycan chains. PMID:17293006

Moudjou, Mohammed; Bernard, Julie; Sabuncu, Elifsu; Langevin, Christelle; Laude, Hubert

2007-04-01

137

To develop with or without the prion protein  

PubMed Central

The deletion of the cellular form of the prion protein (PrPC) in mouse, goat, and cattle has no drastic phenotypic consequence. This stands in apparent contradiction with PrPC quasi-ubiquitous expression and conserved primary and tertiary structures in mammals, and its pivotal role in neurodegenerative diseases such as prion and Alzheimer's diseases. In zebrafish embryos, depletion of PrP ortholog leads to a severe loss-of-function phenotype. This raises the question of a potential role of PrPC in the development of all vertebrates. This view is further supported by the early expression of the PrPC encoding gene (Prnp) in many tissues of the mouse embryo, the transient disruption of a broad number of cellular pathways in early Prnp?/? mouse embryos, and a growing body of evidence for PrPC involvement in the regulation of cell proliferation and differentiation in various types of mammalian stem cells and progenitors. Finally, several studies in both zebrafish embryos and in mammalian cells and tissues in formation support a role for PrPC in cell adhesion, extra-cellular matrix interactions and cytoskeleton. In this review, we summarize and compare the different models used to decipher PrPC functions at early developmental stages during embryo- and organo-genesis and discuss their relevance. PMID:25364763

Halliez, Sophie; Passet, Bruno; Martin-Lanneree, Severine; Hernandez-Rapp, Julia; Laude, Hubert; Mouillet-Richard, Sophie; Vilotte, Jean-Luc; Beringue, Vincent

2014-01-01

138

Prions in Yeast  

PubMed Central

The concept of a prion as an infectious self-propagating protein isoform was initially proposed to explain certain mammalian diseases. It is now clear that yeast also has heritable elements transmitted via protein. Indeed, the “protein only” model of prion transmission was first proven using a yeast prion. Typically, known prions are ordered cross-? aggregates (amyloids). Recently, there has been an explosion in the number of recognized prions in yeast. Yeast continues to lead the way in understanding cellular control of prion propagation, prion structure, mechanisms of de novo prion formation, specificity of prion transmission, and the biological roles of prions. This review summarizes what has been learned from yeast prions. PMID:22879407

Liebman, Susan W.; Chernoff, Yury O.

2012-01-01

139

Prion protein biosynthesis and its emerging role in neurodegeneration  

PubMed Central

Various fatal neurodegenerative disorders are caused by altered metabolism of the prion protein (PrP). These diseases are typically transmissible by an unusual ‘protein-only’ mechanism in which a misfolded isomer, PrPSc, confers its aberrant conformation onto normal cellular PrP. An impressive range of studies has investigated nearly every aspect of this fascinating event; yet, our understanding of how PrPSc accumulation might lead to cellular dysfunction and neurodegeneration is trifling. Recent advances in our understanding of normal PrP biosynthesis and degradation might have unexpectedly shed new light on this complex problem. Indeed, our current understanding of normal PrP cell biology, coupled with a growing appreciation of its complex metabolism, is providing new hypotheses for PrP-mediated neurodegeneration. PMID:19447626

Chakrabarti, Oishee; Ashok, Aarthi; Hegde, Ramanujan S.

2011-01-01

140

NMR structure of the mouse prion protein domain PrP(121-231)  

Microsoft Academic Search

THE 'protein only' hypothesis1 states that a modified form of normal prion protein triggers infectious neurodegenerative diseases, such as bovine spongiform encephalopathy (BSE), or Creutzfeldt-Jakob disease (CJD) in humans2-4. Prion proteins are thought to exist in two different conformations5: the 'benign' PrPC form, and the infectious 'scrapie form', PrPSc. Knowledge of the three-dimensional structure of PrPC is essential for understanding

Roland Riek; Simone Hornemann; Gerhard Wider; Martin Billeter; Rudi Glockshuber; Kurt Wüthrich

1996-01-01

141

CYTOSOLIC PRION PROTEIN IS THE PREDOMINANT ANTI-BAX PRION PROTEIN FORM: EXCLUSION OF TRANSMEMBRANE AND SECRETED PRION PROTEIN FORMS IN THE ANTI-BAX FUNCTION  

PubMed Central

SUMMARY Prion protein (PrP) prevents Bax-mediated cell death by inhibiting the initial Bax conformational change that converts cytosolic Bax into a pro-apoptotic protein. PrP is mostly a glycophosphatidylinositol-anchored cell surface protein but it is also retrotranslocated into cytosolic PrP (CyPrP) or can become a type 1 or type 2 transmembrane protein. To determine the form and subcellular location of the PrP that has anti-Bax function, we co-expressed various Syrian hamster PrP (SHaPrP) mutants that favour specific PrP topologies and subcellular localization with N-terminally green fluorescent protein tagged pro-apoptotic Bax (EGFP-Bax) in MCF-7 cells and primary human neurons. Mutants that generate both CyPrP and secreted PrP (SecPrP) or only CyPrP have anti-Bax activity. Mutants that produce CtmPrP or NtmPrP lose the anti-Bax activity, despite their ability to also make SecPrP. Transmembrane generating mutants do not produce CyPrP and both normal and cognate mutant forms of CyPrP rescue against the loss of anti-Bax activity. SecPrP generating constructs also produce non-membrane attached SecPrP. However, this form of PrP has minimal anti-Bax activity. We conclude that CyPrP is the predominant form of PrP with anti-Bax function. These results imply that the retro-translocation of PrP encompasses a survival function and is not merely a pathway for the proteasomal degradation of misfolded protein. PMID:18590778

Lin, David T. S.; Jodoin, Julie; Baril, Michael; Goodyer, Cynthia G.; LeBlanc, Andrea C.

2008-01-01

142

Definable Equilibrium States in the Folding of Human Prion Protein Laszlo L. P. Hosszu,*,, Mark A. Wells,, Graham S. Jackson, Samantha Jones, Mark Batchelor,  

E-print Network

Definable Equilibrium States in the Folding of Human Prion Protein Laszlo L. P. Hosszu,*,,§ Mark A,§ Jonathan P. Waltho,§ and John Collinge MRC Prion Unit and National Prion Clinic, Institute of Neurology: The role of conformational intermediates in the conversion of prion protein from its normal cellular form

Hosszu, Laszlo

143

Prion diseases and the immune system.  

PubMed

Transmissible spongiform encephalopathies are caused by unusual infectious agents that are purported to contain a single type of macromolecule, a modified host glycoprotein. The term prion has been applied to this group of agents. Surprisingly, the immune system appears to behave as a Trojan's horse rather than a protective fortification during prion infections. Because prions seem to be essentially composed of a protein, PrP(Sc), identical in sequence to a host encoded protein, PrP(C), the specific immune system displays a natural tolerance. However, lymphoid organs are strongly implicated in the preclinical stages of the disease. Certain immunodeficient animals are resistant to prions after peripheral inoculation. In normal subjects, cells of the immune system support the replication of prions and/or allow neuroinvasion. A better understanding of these aspects of prion diseases could lead to immunomanipulation strategies aimed at preventing the spread of infectious agents to the central nervous system. PMID:10900153

Aucouturier, P; Carp, R I; Carnaud, C; Wisniewski, T

2000-08-01

144

Context-dependent perturbation of neural systems in transgenic mice expressing a cytosolic prion protein  

E-print Network

We analyzed the relationship between pathogenic protein expression and perturbations to brain anatomy and physiology in a genetic model of prion disease. In this model, the mouse line 1D4, neuropathology is promoted by ...

Lindquist, Susan

145

Conversion of a yeast prion protein to an infectious form in bacteria  

E-print Network

Prions are infectious, self-propagating protein aggregates that have been identified in evolutionarily divergent members of the eukaryotic domain of life. Nevertheless, it is not yet known whether prokaryotes can support ...

Lindquist, Susan

146

Transmissibility of Atypical Scrapie in Ovine Transgenic Mice: Major Effects of Host Prion Protein Expression and Donor Prion Genotype  

Microsoft Academic Search

Atypical scrapie or Nor98 has been identified as a transmissible spongiform encephalopathy (TSE) that is clearly distinguishable from classical scrapie and BSE, notably regarding the biochemical features of the protease-resistant prion protein PrPres and the genetic factors involved in susceptibility to the disease. In this study we transmitted the disease from a series of 12 French atypical scrapie isolates in

Jean-Noël Arsac; Dominique Bétemps; Eric Morignat; Cécile Féraudet; Anna Bencsik; Denise Aubert; Jacques Grassi; Thierry Baron

2009-01-01

147

Cell-surface prion protein interacts with glycosaminoglycans.  

PubMed Central

We used ELISA and flow cytometry to study the binding of prion protein PrP to glycosaminoglycans (GAGs). We found that recombinant human PrP (rPrP) binds GAGs including chondroitin sulphate A, chondroitin sulphate B, hyaluronic acid, and heparin. rPrP binding to GAGs occurs via the N-terminus, a region known to bind divalent cations. Additionally, rPrP binding to GAGs is enhanced in the presence of Cu2+ and Zn2+, but not Ca2+ and Mn2+. rPrP binds heparin strongest, and the binding is inhibited by certain heparin analogues, including heparin disaccharide and sulphate-containing monosaccharides, but not by acetylated heparin. Full-length normal cellular prion protein (PrPC), but not N-terminally truncated PrPC species, from human brain bind GAGs in a similar Cu2+/Zn2+-enhanced fashion. We found that GAGs specifically bind to a synthetic peptide corresponding to amino acid residues 23-35 in the N-terminus of rPrP. We further demonstrated that while both wild-type PrPC and an octapeptide-repeat-deleted mutant PrP produced by transfected cells bound heparin at the cell surface, the PrP N-terminal deletion mutant and non-transfectant control failed to bind heparin. Binding of heparin to wild-type PrPC on the cell surface results in a reduction of the level of cell-surface PrPC. These results provide strong evidence that PrPC is a surface receptor for GAGs. PMID:12186633

Pan, Tao; Wong, Boon-Seng; Liu, Tong; Li, Ruliang; Petersen, Robert B; Sy, Man-Sun

2002-01-01

148

Specificity of the J-protein Sis1 in the propagation of 3 yeast prions  

PubMed Central

Yeast prions, such as [PSI+], [RNQ+], and [URE3], are heritable elements formed by proteins capable of acquiring self-perpetuating conformations. Their propagation is dependent on fragmentation of the amyloid protein complexes formed to generate the additional seeds necessary for conversion of nascent soluble protein to the prion conformation. We report that, in addition to its known role in [RNQ+] propagation, Sis1, a J-protein cochaperone of Hsp70 Ssa, is also specifically required for propagation of [PSI+] and [URE3]. Whereas both [RNQ+] and [URE3] are cured rapidly upon SIS1 repression, [PSI+] loss is markedly slower. This disparity cannot be explained simply by differences in seed number, as [RNQ+] and [PSI+] are lost with similar kinetics upon inhibition of Hsp104, a remodeling protein required for propagation of all yeast prions. Rather, in the case of [PSI+], our results are consistent with the partial impairment, rather than the complete abolition, of fragmentation of prion complexes upon Sis1 depletion. We suggest that a common set of molecular chaperones, the J-protein Sis1, the Hsp70 Ssa, and the AAA+ ATPase Hsp104, act sequentially in the fragmentation of all yeast prions, but that the threshold of Sis1 activity required for each prion varies. PMID:18955697

Higurashi, Takashi; Hines, Justin K.; Sahi, Chandan; Aron, Rebecca; Craig, Elizabeth A.

2008-01-01

149

Lactoferrin from bovine colostrum regulates prolyl hydroxylase 2 activity and prevents prion protein-mediated neuronal cell damage via cellular prion protein.  

PubMed

Prion disorders are associated with the conversion of normal cellular prion protein (PrPc) to the abnormal scrapie isoform of prion protein (PrPsc). Recent studies have shown that expression of normal PrPc is regulated by hypoxia-inducible factor-1 alpha (HIF-1?), and that lactoferrin increases full-length PrPc on the cell surface. Lactoferrin is an 80-kDa iron-binding glycoprotein with various biological activities, including iron-chelating ability. HIF-1? and the associated ubiquitin-proteasome pathway are regulated by HIF prolyl-hydroxylases 2 (PHD2). We hypothesized that lactoferrin regulates PHD2 expression and enzymatic activity, and the PHD2 regulation promotes HIF-1? stability and prevention of neuronal cell death mediated by prion protein (PrP) residues (106-126). Lactoferrin prevented PrP (106-126)-induced neurotoxicity by the induction of PrPc expression via promoting HIF-1? stability in neuronal cells. Our results demonstrated that lactoferrin prevented PrP (106-126)-induced neurotoxicity via the up-regulation of HIF-1? stability determined by PHD2 expression and enzymatic activity. These findings suggest that possible therapies such as PHD2 inhibition, or promotion of lactoferrin secretion, may have clinical benefits in neurodegenerative diseases, including prion disease. PMID:24875174

Park, Y-G; Moon, J-H; Park, S-Y

2014-08-22

150

Crystallographic Studies of Prion Protein (PrP) Segments Suggest How Structural Changes Encoded by Polymorphism at Residue 129 Modulate Susceptibility to Human Prion Disease  

SciTech Connect

A single nucleotide polymorphism (SNP) in codon 129 of the human prion gene, leading to a change from methionine to valine at residue 129 of prion protein (PrP), has been shown to be a determinant in the susceptibility to prion disease. However, the molecular basis of this effect remains unexplained. In the current study, we determined crystal structures of prion segments having either Met or Val at residue 129. These 6-residue segments of PrP centered on residue 129 are 'steric zippers,' pairs of interacting {beta}-sheets. Both structures of these 'homozygous steric zippers' reveal direct intermolecular interactions between Met or Val in one sheet and the identical residue in the mating sheet. These two structures, plus a structure-based model of the heterozygous Met-Val steric zipper, suggest an explanation for the previously observed effects of this locus on prion disease susceptibility and progression.

Apostol, Marcin I.; Sawaya, Michael R.; Cascio, Duilio; Eisenberg, David (UCLA)

2010-09-23

151

Structural plasticity of the cellular prion protein and implications in health and disease  

PubMed Central

Two lines of transgenic mice expressing mouse/elk and mouse/horse prion protein (PrP) hybrids, which both form a well-structured ?2–?2 loop in the NMR structures at 20 °C termed rigid-loop cellular prion proteins (RL-PrPC), presented with accumulation of the aggregated scrapie form of PrP in brain tissue, and the mouse/elk hybrid has also been shown to develop a spontaneous transmissible spongiform encephalopathy. Independently, there is in vitro evidence for correlations between the amino acid sequence in the ?2–?2 loop and the propensity for conformational transitions to disease-related forms of PrP. To further contribute to the structural basis for these observations, this paper presents a detailed characterization of RL-PrPC conformations in solution. A dynamic local conformational polymorphism involving the ?2–?2 loop was found to be evolutionarily preserved among all mammalian species, including those species for which the WT PrP forms an RL-PrPC. The interconversion between two ensembles of PrPC conformers that contain, respectively, a 310-helix turn or a type I ?-turn structure of the ?2–?2 loop, exposes two different surface epitopes, which are analyzed for their possible roles in the still evasive function of PrPC in healthy organisms and/or at the onset of a transmissible spongiform encephalopathy. PMID:23650394

Christen, Barbara; Damberger, Fred F.; Perez, Daniel R.; Hornemann, Simone; Wuthrich, Kurt

2013-01-01

152

Structural plasticity of the cellular prion protein and implications in health and disease.  

PubMed

Two lines of transgenic mice expressing mouse/elk and mouse/horse prion protein (PrP) hybrids, which both form a well-structured ?2-?2 loop in the NMR structures at 20 °C termed rigid-loop cellular prion proteins (RL-PrP(C)), presented with accumulation of the aggregated scrapie form of PrP in brain tissue, and the mouse/elk hybrid has also been shown to develop a spontaneous transmissible spongiform encephalopathy. Independently, there is in vitro evidence for correlations between the amino acid sequence in the ?2-?2 loop and the propensity for conformational transitions to disease-related forms of PrP. To further contribute to the structural basis for these observations, this paper presents a detailed characterization of RL-PrP(C) conformations in solution. A dynamic local conformational polymorphism involving the ?2-?2 loop was found to be evolutionarily preserved among all mammalian species, including those species for which the WT PrP forms an RL-PrP(C). The interconversion between two ensembles of PrP(C) conformers that contain, respectively, a 310-helix turn or a type I ?-turn structure of the ?2-?2 loop, exposes two different surface epitopes, which are analyzed for their possible roles in the still evasive function of PrP(C) in healthy organisms and/or at the onset of a transmissible spongiform encephalopathy. PMID:23650394

Christen, Barbara; Damberger, Fred F; Pérez, Daniel R; Hornemann, Simone; Wüthrich, Kurt

2013-05-21

153

Chimeric elk/mouse prion proteins in transgenic mice  

PubMed Central

Chronic wasting disease (CWD) of deer and elk is a highly communicable neurodegenerative disorder caused by prions. Investigations of CWD are hampered by slow bioassays in transgenic (Tg) mice. Towards the development of Tg mice that will be more susceptible to CWD prions, we created a series of chimeric elk/mouse transgenes that encode the N terminus of elk PrP (ElkPrP) up to residue Y168 and the C terminus of mouse PrP (MoPrP) beyond residue 169 (mouse numbering), designated Elk3M(SNIVVK). Between codons 169 and 219, six residues distinguish ElkPrP from MoPrP: N169S, T173N, V183I, I202V, I214V and R219K. Using chimeric elk/mouse PrP constructs, we generated 12 Tg mouse lines and determined incubation times after intracerebral inoculation with the mouse-passaged RML scrapie or Elk1P CWD prions. Unexpectedly, one Tg mouse line expressing Elk3M(SNIVVK) exhibited incubation times of <70 days when inoculated with RML prions; a second line had incubation times of <90 days. In contrast, mice expressing full-length ElkPrP had incubation periods of >250 days for RML prions. Tg(Elk3M,SNIVVK) mice were less susceptible to CWD prions than Tg(ElkPrP) mice. Changing three C-terminal mouse residues (202, 214 and 219) to those of elk doubled the incubation time for mouse RML prions and rendered the mice resistant to Elk1P CWD prions. Mutating an additional two residues from mouse to elk at codons 169 and 173 increased the incubation times for mouse prions to >300 days, but made the mice susceptible to CWD prions. Our findings highlight the role of C-terminal residues in PrP that control the susceptibility and replication of prions. PMID:23100369

Tamguney, Gultekin; Giles, Kurt; Oehler, Abby; Johnson, Natrina L.; DeArmond, Stephen J.

2013-01-01

154

Prion Protein-Specific Antibodies-Development, Modes of Action and Therapeutics Application  

PubMed Central

Prion diseases or Transmissible Spongiform Encephalopathies (TSEs) are lethal neurodegenerative disorders involving the misfolding of the host encoded cellular prion protein, PrPC. This physiological form of the protein is expressed throughout the body, and it reaches the highest levels in the central nervous system where the pathology occurs. The conversion into the pathogenic isoform denoted as prion or PrPSc is the key event in prion disorders. Prominent candidates for the treatment of prion diseases are antibodies and their derivatives. Anti-PrPC antibodies are able to clear PrPSc from cell culture of infected cells. Furthermore, application of anti-PrPC antibodies suppresses prion replication in experimental animal models. Major drawbacks of immunotherapy are immune tolerance, the risks of neurotoxic side effects, limited ability of compounds to cross the blood-brain barrier and their unfavorable pharmacokinetic. The focus of this review is to recapitulate the current understanding of the molecular mechanisms for antibody mediated anti-prion activity. Although relevant for designing immunotherapeutic tools, the characterization of key antibody parameters shaping the molecular mechanism of the PrPC to PrPSc conversion remains elusive. Moreover, this review illustrates the various attempts towards the development of anti-PrP antibody compounds and discusses therapeutic candidates that modulate PrP expression. PMID:25275428

Rovis, Tihana Lenac; Legname, Giuseppe

2014-01-01

155

Molecular Dynamics Studies on the Structural Stability of Wild-type HORSE PRION PROTEIN  

E-print Network

Prion diseases {\\it (e.g. Creutzfeldt-Jakob disease (CJD), variant CJD (vCJD), Gerstmann-Str$\\ddot{\\text{a}}$ussler-Scheinker syndrome (GSS), Fatal Familial Insomnia (FFI) and Kuru in humans, scrapie in sheep, bovine spongiform encephalopathy (BSE or `mad-cow' disease) and chronic wasting disease (CWD) in cattles)} are invariably fatal and highly infectious neurodegenerative diseases affecting humans and animals. However, by now there have not been some effective therapeutic approaches or medications to treat all these prion diseases. Rabbits, dogs, and horses are the only mammalian species reported to be resistant to infection from prion diseases isolated from other species. Recently, the $\\beta$2--$\\alpha$2 loop has been reported to contribute to their protein structural stabilities. The author has found that rabbit prion protein has a strong salt bridge ASP177-ARG163 (like a taut bow string) keeping this loop linked. This paper confirms that this salt bridge also contributes to the structural stability of ...

Zhang, Jiapu

2011-01-01

156

Human Tonsil-Derived Follicular Dendritic-Like Cells are Refractory to Human Prion Infection in Vitro and Traffic Disease-Associated Prion Protein to Lysosomes  

PubMed Central

The molecular mechanisms involved in human cellular susceptibility to prion infection remain poorly defined. This is due, in part, to the absence of any well characterized and relevant cultured human cells susceptible to infection with human prions, such as those involved in Creutzfeldt-Jakob disease. In variant Creutzfeldt-Jakob disease, prion replication is thought to occur first in the lymphoreticular system and then spread into the brain. We have, therefore, examined the susceptibility of a human tonsil-derived follicular dendritic cell-like cell line (HK) to prion infection. HK cells were found to display a readily detectable, time-dependent increase in cell-associated abnormal prion protein (PrPTSE) when exposed to medium spiked with Creutzfeldt-Jakob disease brain homogenate, resulting in a coarse granular perinuclear PrPTSE staining pattern. Despite their high level of cellular prion protein expression, HK cells failed to support infection, as judged by longer term maintenance of PrPTSE accumulation. Colocalization studies revealed that exposure of HK cells to brain homogenate resulted in increased numbers of detectable lysosomes and that these structures immunostained intensely for PrPTSE after exposure to Creutzfeldt-Jakob disease brain homogenate. Our data suggest that human follicular dendritic-like cells and perhaps other human cell types are able to avoid prion infection by efficient lysosomal degradation of PrPTSE. PMID:24183781

Krejciova, Zuzana; De Sousa, Paul; Manson, Jean; Ironside, James W.; Head, Mark W.

2014-01-01

157

Prion Protein Misfolding Affects Calcium Homeostasis and Sensitizes Cells to Endoplasmic Reticulum Stress  

Microsoft Academic Search

Prion-related disorders (PrDs) are fatal neurodegenerative disorders characterized by progressive neuronal impairment as well as the accumulation of an abnormally folded and protease resistant form of the cellular prion protein, termed PrPRES. Altered endoplasmic reticulum (ER) homeostasis is associated with the occurrence of neurodegeneration in sporadic, infectious and familial forms of PrDs. The ER operates as a major intracellular calcium

Mauricio Torres; Karen Castillo; Ricardo Armisén; Andrés Stutzin; Claudio Soto; Claudio Hetz; Maria A. Deli

2010-01-01

158

Altered circadian activity rhythms and sleep in mice devoid of prion protein  

Microsoft Academic Search

THERE is a wealth of data supporting a central role for the prion protein (PrP) in the neurodegenerative prion diseases of both humans and other species1, yet the normal function of PrP, which is expressed at the cell surface of neurons and glial cells2,3, is unknown. It has been speculated that neuropathology may be due to loss of normal function

I. Tobler; S. E. Gaus; T. Deboer; P. Achermann; M. Fischer; T. Rülicke; M. Moser; B. Oesch; P. A. McBride; J. C. Manson

1996-01-01

159

Intraepithelial and Interstitial Deposition of Pathological Prion Protein in Kidneys of Scrapie-Affected Sheep  

Microsoft Academic Search

Prions have been documented in extra-neuronal and extra-lymphatic tissues of humans and various ruminants affected by Transmissible Spongiform Encephalopathy (TSE). The presence of prion infectivity detected in cervid and ovine blood tempted us to reason that kidney, the organ filtrating blood derived proteins, may accumulate disease associated PrPSc. We collected and screened kidneys of experimentally, naturally scrapie-affected and control sheep

Ciriaco Ligios; Giovanna Maria Cancedda; Ilan Margalith; Cinzia Santucciu; Laura Madau; Caterina Maestrale; Massimo Basagni; Mariangela Saba; Mathias Heikenwalder; Neil Mabbott

2007-01-01

160

Cloning and expression of prion protein encoding gene of flounder ( Paralichthys olivaceus)  

NASA Astrophysics Data System (ADS)

The prion protein (PrP) encoding gene of flounder ( Paralichthys olivaceus) was cloned. It was not interrupted by an intron. This gene has two promoters in its 5' upstream, indicating that its transcription may be intensive, and should have an important function. It was expressed in all 14 tissues tested, demonstrating that it is a house-keeping gene. Its expression in digestion and reproduction systems implies that the possible prions of fish may transfer horizontally.

Zhang, Zhiwen; Sun, Xiuqin; Zhang, Jinxing; Zan, Jindong

2008-02-01

161

Polymorphism distribution of prion protein codon 117, 129 and 171 in Taiwan.  

PubMed

Prion diseases compass transmissible spongiform neurodegenerative diseases from various causes, including the genetic and infectious ones. We investigated the prevalence of codon 117, 129 and 171 polymorphism in prion protein (PrP) in Taiwanese, mainly for the sake of the informative absence of this genetic distribution. Our subjects were 419 aged ones of Han ethic origin. We evaluated the PrP gene (PRNP) polymorphism by restriction fragment length polymorphism, after amplification of their genomic DNAs by polymerase chain reactions with specific primers, digested by restriction enzyme PvuII (for codon 117), NspI (for codon 129), and BbvI (for codon 171), respectively, and confirmed by nucleotide sequencing. All of the subjects were homozygotes at codon 117 (Ala/Ala, gca/gca) and 171 (Asn/Asn, aac/aac). There were no valine homozygotes (Val/Val) in our 419 subjects, and nine subjects (2.1%) showed methionine-valine heterozygosity (Mal/Val, atg/gtg). The methionine homozygotes (Met/Met) comprised the major population (97.9%), and the prevalence of distribution is different to that seen in Caucasians. The almost 100% conservation of the domain from codon 117 to 171 implies the warranty of PrP in cellular functions. The high prevalence of Met/Met alleles in Taiwan did not imply an increased risk of CJD, and the genetic susceptibility of CJD by codon 129 of PrP may be still elusive for the infectivity. PMID:17410475

Wang, Kaw-Chen; Wang, Vinchi; Sun, Ming-Chieh; Chiueh, Ti-I; Soong, Bing-Wen; Shan, Din-E

2007-01-01

162

N-terminal peptides from unprocessed prion proteins enter cells by macropinocytosis  

SciTech Connect

A peptide derived from the N-terminus of the unprocessed bovine prion protein (bPrPp), incorporating the hydrophobic signal sequence (residues 1-24) and a basic domain (KKRPKP, residues 25-30), internalizes into mammalian cells, even when coupled to a sizeable cargo, and therefore functions as a cell-penetrating peptide (CPP). Confocal microscopy and co-localization studies indicate that the internalization of bPrPp is mainly through macropinocytosis, a fluid-phase endocytosis process, initiated by binding to cell-surface proteoglycans. Electron microscopy studies show internalized bPrPp-DNA-gold complexes residing in endosomal vesicles. bPrPp induces expression of a complexed luciferase-encoding DNA plasmid, demonstrating the peptide's ability to transport the cargo across the endosomal membrane and into the cytosol and nucleus. The novel CPP activity of the unprocessed N-terminal domain of PrP could be important for the retrotranslocation of partly processed PrP and for PrP trafficking inside or between cells, with implications for the infectivity associated with prion diseases.

Magzoub, Mazin [Department of Biochemistry and Biophysics, Stockholm University (Sweden); Sandgren, Staffan [Department of Clinical Sciences, Section for Oncology, Lund University (Sweden); Lundberg, Pontus [Department of Neurochemistry, Stockholm University (Sweden); Oglecka, Kamila [Department of Biochemistry and Biophysics, Stockholm University (Sweden); Lilja, Johanna [Department of Clinical Sciences, Section for Oncology, Lund University (Sweden); Wittrup, Anders [Department of Clinical Sciences, Section for Oncology, Lund University (Sweden); Goeran Eriksson, L.E. [Department of Biochemistry and Biophysics, Stockholm University (Sweden); Langel, Ulo [Department of Neurochemistry, Stockholm University (Sweden); Belting, Mattias [Department of Clinical Sciences, Section for Oncology, Lund University (Sweden)]. E-mail: mattias.belting@med.lu.se; Graeslund, Astrid [Department of Biochemistry and Biophysics, Stockholm University (Sweden)]. E-mail: astrid@dbb.su.se

2006-09-22

163

Systemic Delivery of siRNA Down Regulates Brain Prion Protein and Ameliorates Neuropathology in Prion Disorder  

PubMed Central

One of the main challenges for neurodegenerative disorders that are principally incurable is the development of new therapeutic strategies, which raises important medical, scientific and societal issues. Creutzfeldt-Jakob diseases are rare neurodegenerative fatal disorders which today remain incurable. The objective of this study was to evaluate the efficacy of the down-regulation of the prion protein (PrP) expression using siRNA delivered by, a water-in-oil microemulsion, as a therapeutic candidate in a preclinical study. After 12 days rectal mucosa administration of Aonys/PrP-siRNA in mice, we observed a decrease of about 28% of the brain PrPC level. The effect of Aonys/PrP-siRNA was then evaluated on prion infected mice. Several mice presented a delay in the incubation and survival time compared to the control groups and a significant impact was observed on astrocyte reaction and neuronal survival in the PrP-siRNA treated groups. These results suggest that a new therapeutic scheme based an innovative delivery system of PrP-siRNA can be envisioned in prion disorders. PMID:24551164

Resina, Sarah; Brillaud, Elsa; Casanova, Danielle; Vincent, Charles; Hamela, Claire; Poupeau, Sophie; Laffont, Mathieu; Gabelle, Audrey; Delaby, Constance; Belondrade, Maxime; Arnaud, Jacques-Damien; Alvarez, Maria-Teresa; Maurel, Jean-Claude; Maurel, Patrick; Crozet, Carole

2014-01-01

164

Regulation of amyloid-? production by the prion protein  

PubMed Central

Alzheimer disease (AD) is characterized by the amyloidogenic processing of the amyloid precursor protein (APP), culminating in the accumulation of amyloid-? peptides in the brain. The enzymatic action of the ?-secretase, BACE1 is the rate-limiting step in this amyloidogenic processing of APP. BACE1 cleavage of wild-type APP (APPWT) is inhibited by the cellular prion protein (PrPC). Our recent study has revealed the molecular and cellular mechanisms behind this observation by showing that PrPC directly interacts with the pro-domain of BACE1 in the trans-Golgi network (TGN), decreasing the amount of BACE1 at the cell surface and in endosomes where it cleaves APPWT, while increasing BACE1 in the TGN where it preferentially cleaves APP with the Swedish mutation (APPSwe). PrPC deletion in transgenic mice expressing the Swedish and Indiana familial mutations (APPSwe,Ind) failed to affect amyloid-? accumulation, which is explained by the differential subcellular sites of action of BACE1 toward APPWT and APPSwe. This, together with our observation that PrPC is reduced in sporadic but not familial AD brain, suggests that PrPC plays a key protective role against sporadic AD. It also highlights the need for an APPWT transgenic mouse model to understand the molecular and cellular mechanisms underlying sporadic AD. PMID:22449984

Griffiths, Heledd H.; Whitehouse, Isobel J.; Hooper, Nigel M.

2012-01-01

165

Amyloid-? nanotubes are associated with prion protein-dependent synaptotoxicity  

PubMed Central

Growing evidence suggests water-soluble, non-fibrillar forms of amyloid-? protein (A?) have important roles in Alzheimer’s disease with toxicities mimicked by synthetic A?1–42. However, no defined toxic structures acting via specific receptors have been identified and roles of proposed receptors, such as prion protein (PrP), remain controversial. Here we quantify binding to PrP of A?1–42 after different durations of aggregation. We show PrP-binding and PrP-dependent inhibition of long-term potentiation (LTP) correlate with the presence of protofibrils. Globular oligomers bind less avidly to PrP and do not inhibit LTP, whereas fibrils inhibit LTP in a PrP-independent manner. That only certain transient A? assemblies cause PrP-dependent toxicity explains conflicting reports regarding the involvement of PrP in A?-induced impairments. We show that these protofibrils contain a defined nanotubular structure with a previously unidentified triple helical conformation. Blocking the formation of A? nanotubes or their interaction with PrP might have a role in treatment of Alzheimer’s disease. PMID:24022506

Nicoll, Andrew J.; Panico, Silvia; Freir, Darragh B.; Wright, Daniel; Terry, Cassandra; Risse, Emmanuel; Herron, Caroline E.; O'Malley, Tiernan; Wadsworth, Jonathan D. F.; Farrow, Mark A.; Walsh, Dominic M.; Saibil, Helen R.; Collinge, John

2013-01-01

166

NMR characterization of the full-length recombinant murine prion protein, mPrP(23–231)  

Microsoft Academic Search

The recombinant murine prion protein, mPrP(23–231), was expressed in E. coli with uniform 15N-labeling. NMR experiments showed that the previously determined globular three-dimensional structure of the C-terminal domain mPrP(121–231) is preserved in the intact protein, and that the N-terminal polypeptide segment 23–120 is flexibly disordered. This structural information is based on nearly complete sequence-specific assignments for the backbone amide nitrogens,

Roland Riek; Simone Hornemann; Gerhard Wider; Rudi Glockshuber; Kurt Wüthrich

1997-01-01

167

Molecular modelling indicates that the pathological conformations of prion proteins might be beta-helical.  

PubMed Central

Creutzfeldt-Jakob disease, kuru, scrapie and bovine spongiform encephalopathy are diseases of the mammalian central nervous system that involve the conversion of a cellular protein into an insoluble extracellular isoform. Spectroscopic studies have shown that the precursor protein contains mainly alpha-helical and random-coil conformations, whereas the prion isoform is largely in the beta conformation. The pathogenic prion is resistant to denaturation and protease digestion and can promote the conversion of the precursor protein to the pathogenic form. These properties have yet to be explained in terms of the structural conformations of the proteins. In the present study, molecular modelling showed that prion proteins could adopt the beta-helical conformation, which has been established for a number of fibrous proteins and has been suggested previously as the basis of amyloid fibrils. The beta-helical conformation provides explanations for the biophysical and biochemical stability of prions, their ability to form templates for the transmission of pathological conformation, and the existence of phenotypical strains of the prion diseases. PMID:10510313

Downing, D T; Lazo, N D

1999-01-01

168

Detection of protease-resistant cervid prion protein in water from a CWD-endemic area  

PubMed Central

Chronic wasting disease (CWD) is the only known transmissible spongiform encephalopathy affecting free-ranging wildlife. Although the exact mode of natural transmission remains unknown, substantial evidence suggests that prions can persist in the environment, implicating components thereof as potential prion reservoirs and transmission vehicles.1–4 CWD-positive animals may contribute to environmental prion load via decomposing carcasses and biological materials including saliva, blood, urine and feces.5–7 Sensitivity limitations of conventional assays hamper evaluation of environmental prion loads in soil and water. Here we show the ability of serial protein misfolding cyclic amplification (sPMCA) to amplify a 1.3 × 10?7 dilution of CWD-infected brain homogenate spiked into water samples, equivalent to approximately 5 × 107 protease resistant cervid prion protein (PrPCWD) monomers. We also detected PrPCWD in one of two environmental water samples from a CWD endemic area collected at a time of increased water runoff from melting winter snow pack, as well as in water samples obtained concurrently from the flocculation stage of water processing by the municipal water treatment facility. Bioassays indicated that the PrPCWD detected was below infectious levels. These data demonstrate detection of very low levels of PrPCWD in the environment by sPMCA and suggest persistence and accumulation of prions in the environment that may promote CWD transmission. PMID:19823039

Nichols, TA; Pulford, Bruce; Wyckoff, A Christy; Meyerett, Crystal; Michel, Brady; Gertig, Kevin; Hoover, Edward A; Jewell, Jean E; Telling, Glenn C

2009-01-01

169

Electrical Conductors and Devices from Prion-Like Proteins.  

National Technical Information Service (NTIS)

The present invention provides novel polypeptides comprising a prion-aggregation domain and a second domain; novel polynucleotides encoding such polypeptides; host cells transformed or transfected with such polynucleotides; novel fibrils with specific fun...

S. Lindquist, T. Scheibel

2005-01-01

170

The prion protein family: a view from the placenta  

PubMed Central

Based on its developmental pattern of expression, early studies suggested the implication of the mammalian Prion protein PrP, a glycosylphosphatidylinositol-anchored ubiquitously expressed and evolutionary conserved glycoprotein encoded by the Prnp gene, in early embryogenesis. However, gene invalidation in several species did not result in obvious developmental abnormalities and it was only recently that it was associated in mice with intra-uterine growth retardation and placental dysfunction. A proposed explanation for this lack of easily detectable developmental-related phenotype is the existence in the genome of one or more gene (s) able to compensate for the absence of PrP. Indeed, two other members of the Prnp gene family have been recently described, Doppel and Shadoo, and the consequences of their invalidation alongside that of PrP tested in mice. No embryonic defect was observed in mice depleted for Doppel and PrP. Interestingly, the co-invalidation of PrP and Shadoo in two independent studies led to apparently conflicting observations, with no apparent consequences in one report and the observation of a developmental defect of the ectoplacental cone that leads to early embryonic lethality in the other. This short review aims at summarizing these recent, apparently conflicting data highlighting the related biological questions and associated implications in terms of animal and human health. PMID:25364742

Makzhami, Samira; Passet, Bruno; Halliez, Sophie; Castille, Johan; Moazami-Goudarzi, Katayoun; Duchesne, Amandine; Vilotte, Marthe; Laude, Hubert; Mouillet-Richard, Sophie; Beringue, Vincent; Vaiman, Daniel; Vilotte, Jean-Luc

2014-01-01

171

Unique Structural Characteristics of the Rabbit Prion Protein*  

PubMed Central

Rabbits are one of the few mammalian species that appear to be resistant to transmissible spongiform encephalopathies due to the structural characteristics of the rabbit prion protein (RaPrPC) itself. Here, we determined the solution structures of the recombinant protein RaPrPC-(91–228) and its S173N variant and detected the backbone dynamics of their structured C-terminal domains-(121–228). In contrast to many other mammalian PrPCs, loop 165–172, which connects ?-sheet-2 and ?-helix-2, is well-defined in RaPrPC. For the first time, order parameters S2 are obtained for residues in this loop region, indicating that loop 165–172 of RaPrPC is highly ordered. Compared with the wild-type RaPrPC, less hydrogen bonds form in the S173N variant. The NMR dynamics analysis reveals a distinct increase in the structural flexibility of loop 165–172 and helix-3 after the S173N substitution, implying that the S173N substitution disturbs the long range interaction of loop 165–172 with helix-3, which further leads to a marked decrease in the global conformational stability. Significantly, RaPrPC possesses a unique charge distribution, carrying a continuous area of positive charges on the surface, which is distinguished from other PrPCs. The S173N substitution causes visible changes of the charge distribution around the recognition sites for the hypothetical protein X. Our results suggest that the ordered loop 165–172 and its interaction with helix-3, together with the unique distribution of surface electrostatic potential, significantly contribute to the unique structural characteristics of RaPrPC. PMID:20639199

Wen, Yi; Li, Jun; Yao, Wenming; Xiong, Minqian; Hong, Jing; Peng, Yu; Xiao, Gengfu; Lin, Donghai

2010-01-01

172

Cellular prion protein and NMDA receptor modulation: protecting against excitotoxicity  

PubMed Central

Although it is well established that misfolding of the cellular prion protein (PrPC) into the ?-sheet-rich, aggregated scrapie conformation (PrPSc) causes a variety of transmissible spongiform encephalopathies (TSEs), the physiological roles of PrPC are still incompletely understood. There is accumulating evidence describing the roles of PrPC in neurodegeneration and neuroinflammation. Recently, we identified a functional regulation of NMDA receptors by PrPC that involves formation of a physical protein complex between these proteins. Excessive NMDA receptor activity during conditions such as ischemia mediates enhanced Ca2+ entry into cells and contributes to excitotoxic neuronal death. In addition, NMDA receptors and/or PrPC play critical roles in neuroinflammation and glial cell toxicity. Inhibition of NMDA receptor activity protects against PrPSc-induced neuronal death. Moreover, in mice lacking PrPC, infarct size is increased after focal cerebral ischemia, and absence of PrPC increases susceptibility of neurons to NMDA receptor-dependent death. Recently, PrPC was found to be a receptor for oligomeric beta-amyloid (A?) peptides, suggesting a role for PrPC in Alzheimer's disease (AD). Our recent findings suggest that A? peptides enhance NMDA receptor current by perturbing the normal copper- and PrPC-dependent regulation of these receptors. Here, we review evidence highlighting a role for PrPC in preventing NMDA receptor-mediated excitotoxicity and inflammation. There is a need for more detailed molecular characterization of PrPC-mediated regulation of NMDA receptors, such as determining which NMDA receptor subunits mediate pathogenic effects upon loss of PrPC-mediated regulation and identifying PrPC binding site(s) on the receptor. This knowledge will allow development of novel therapeutic interventions for not only TSEs, but also for AD and other neurodegenerative disorders involving dysfunction of PrPC. PMID:25364752

Black, Stefanie A. G.; Stys, Peter K.; Zamponi, Gerald W.; Tsutsui, Shigeki

2014-01-01

173

Flexibility damps macromolecular crowding effects on protein folding dynamics: Application to the murine prion protein (121-231)  

NASA Astrophysics Data System (ADS)

A model of protein folding kinetics is applied to study the combined effects of protein flexibility and macromolecular crowding on protein folding rate and stability. It is found that the increase in stability and folding rate promoted by macromolecular crowding is damped for proteins with highly flexible native structures. The model is applied to the folding dynamics of the murine prion protein (121-231). It is found that the high flexibility of the native isoform of the murine prion protein (121-231) reduces the effects of macromolecular crowding on its folding dynamics. The relevance of these findings for the pathogenic mechanism are discussed.

Bergasa-Caceres, Fernando; Rabitz, Herschel A.

2014-01-01

174

The tip of the iceberg: RNA-binding proteins with prion-like domains in neurodegenerative disease  

E-print Network

Review The tip of the iceberg: RNA-binding proteins with prion-like domains in neurodegenerative- nected with ALS and FTLD-U. We strongly suspect that we have now merely reached the tip of the iceberg this article as: King, O.D., et al., The tip of the iceberg: RNA-binding proteins with prion-like domains

Shorter, James

175

Fatal Familial Insomnia and Familial Creutzfeldt-Jakob Disease: Different Prion Proteins Determined by a DNA Polymorphism  

Microsoft Academic Search

Fatal familial insomnia and a subtype of Creutzfeldt-Jakob disease, two clinically and pathologically distinct diseases, are linked to the same mutation at codon 178 (Asp-178 --> Asn) but segregate with different genotypes determined by this mutation and the methionine-valine polymorphism at codon 129 of the prion protein gene. The abnormal isoforms of the prion protein in these two diseases were

Lucia Monari; Shu G. Chen; Paul Brown; Piero Parchi; Robert B. Petersen; Jacqueline Mikol; Franscoise Gray; Pietro Cortelli; Pasquale Montagna; Bernardino Ghetti; Lev G. Goldfarb; D. Carleton Gajdusek; Elio Lugaresi; Pierluigi Gambetti; Lucila Autilio-Gambetti

1994-01-01

176

Copper-zinc cross-modulation in prion protein binding.  

PubMed

In this paper we report a systematic XAS study of a set of samples in which Cu(II) was progressively added to complexes in which Zn(II) was bound to the tetra-octarepeat portion of the prion protein. This work extends previous EPR and XAS analysis in which, in contrast, the effect of adding Zn(II) to Cu(II)-tetra-octarepeat complexes was investigated. Detailed structural analysis of the XAS spectra taken at both the Cu and Zn K-edge when the two metals are present at different relative concentrations revealed that Zn(II) and Cu(II) ions compete for binding to the tetra-octarepeat peptide by cross-regulating their relative binding modes. We show that the specific metal-peptide coordination mode depends not only, as expected, on the relative metal concentrations, but also on whether Zn(II) or Cu(II) was first bound to the peptide. In particular, it seems that the Zn(II) binding mode in the absence of Cu(II) is able to promote the formation of small peptide clusters in which triplets of tetra-octarepeats are bridged by pairs of Zn ions. When Cu(II) is added, it starts competing with Zn(II) for binding, disrupting the existing peptide cluster arrangement, despite the fact that Cu(II) is unable to completely displace Zn(II). These results may have a bearing on our understanding of peptide-aggregation processes and, with the delicate cross-regulation balancing we have revealed, seem to suggest the existence of an interesting, finely tuned interplay among metal ions affecting protein binding, capable of providing a mechanism for regulation of metal concentration in cells. PMID:25395329

Stellato, Francesco; Minicozzi, Velia; Millhauser, Glenn L; Pascucci, Marco; Proux, Olivier; Rossi, Giancarlo C; Spevacek, Ann; Morante, Silvia

2014-12-01

177

Monitoring prion protein expression in complex biological samples by SERS for diagnostic applications  

NASA Astrophysics Data System (ADS)

Surface-enhanced Raman spectroscopy (SERS) allows a new insight into the analysis of cell physiology. In this work, the difficulty of producing suitable substrates that, besides permitting the amplification of the Raman signal, do not interact with the biological material causing alteration, has been overcome by a combined method of hydrothermal green synthesis and thermal annealing. The SERS analysis of the cell membrane has been performed with special attention to the cellular prion protein PrPC. In addition, SERS has also been used to reveal the prion protein-Cu(II) interaction in four different cell models (B104, SH-SY5Y, GN11, HeLa), expressing PrPC at different levels. A significant implication of the current work consists of the intriguing possibility of revealing and quantifying prion protein expression in complex biological samples by a cheap SERS-based method, replacing the expensive and time-consuming immuno-assay systems commonly employed.

Manno, D.; Filippo, E.; Fiore, R.; Serra, A.; Urso, E.; Rizzello, A.; Maffia, M.

2010-04-01

178

Prevalence of the prion protein gene E211K variant in U.S. cattle  

PubMed Central

Background In 2006, an atypical U.S. case of bovine spongiform encephalopathy (BSE) was discovered in Alabama and later reported to be polymorphic for glutamate (E) and lysine (K) codons at position 211 in the bovine prion protein gene (Prnp) coding sequence. A bovine E211K mutation is important because it is analogous to the most common pathogenic mutation in humans (E200K) which causes hereditary Creutzfeldt – Jakob disease, an autosomal dominant form of prion disease. The present report describes a high-throughput matrix-associated laser desorption/ionization-time-of-flight mass spectrometry assay for scoring the Prnp E211K variant and its use to determine an upper limit for the K211 allele frequency in U.S. cattle. Results The K211 allele was not detected in 6062 cattle, including those from five commercial beef processing plants (3892 carcasses) and 2170 registered cattle from 42 breeds. Multiple nearby polymorphisms in Prnp coding sequence of 1456 diverse purebred cattle (42 breeds) did not interfere with scoring E211 or K211 alleles. Based on these results, the upper bounds for prevalence of the E211K variant was estimated to be extremely low, less than 1 in 2000 cattle (Bayesian analysis based on 95% quantile of the posterior distribution with a uniform prior). Conclusion No groups or breeds of U.S. cattle are presently known to harbor the Prnp K211 allele. Because a carrier was not detected, the number of additional atypical BSE cases with K211 will also be vanishingly low. PMID:18625065

Heaton, Michael P; Keele, John W; Harhay, Gregory P; Richt, Jürgen A; Koohmaraie, Mohammad; Wheeler, Tommy L; Shackelford, Steven D; Casas, Eduardo; King, D Andy; Sonstegard, Tad S; Van Tassell, Curtis P; Neibergs, Holly L; Chase, Chad C; Kalbfleisch, Theodore S; Smith, Timothy PL; Clawson, Michael L; Laegreid, William W

2008-01-01

179

Crystal structure of human prion protein bound to a therapeutic antibody.  

PubMed

Prion infection is characterized by the conversion of host cellular prion protein (PrP(C)) into disease-related conformers (PrP(Sc)) and can be arrested in vivo by passive immunization with anti-PrP monoclonal antibodies. Here, we show that the ability of an antibody to cure prion-infected cells correlates with its binding affinity for PrP(C) rather than PrP(Sc). We have visualized this interaction at the molecular level by determining the crystal structure of human PrP bound to the Fab fragment of monoclonal antibody ICSM 18, which has the highest affinity for PrP(C) and the highest therapeutic potency in vitro and in vivo. In this crystal structure, human PrP is observed in its native PrP(C) conformation. Interactions between neighboring PrP molecules in the crystal structure are mediated by close homotypic contacts between residues at position 129 that lead to the formation of a 4-strand intermolecular beta-sheet. The importance of this residue in mediating protein-protein contact could explain the genetic susceptibility and prion strain selection determined by polymorphic residue 129 in human prion disease, one of the strongest common susceptibility polymorphisms known in any human disease. PMID:19204296

Antonyuk, S V; Trevitt, C R; Strange, R W; Jackson, G S; Sangar, D; Batchelor, M; Cooper, S; Fraser, C; Jones, S; Georgiou, T; Khalili-Shirazi, A; Clarke, A R; Hasnain, S S; Collinge, J

2009-02-24

180

Stress-dependent Proteolytic Processing of the Actin Assembly Protein Lsb1 Modulates a Yeast Prion.  

PubMed

Yeast prions are self-propagating amyloid-like aggregates of Q/N-rich protein that confer heritable traits and provide a model of mammalian amyloidoses. [PSI(+)] is a prion isoform of the translation termination factor Sup35. Propagation of [PSI(+)] during cell division under normal conditions and during the recovery from damaging environmental stress depends on cellular chaperones and is influenced by ubiquitin proteolysis and the actin cytoskeleton. The paralogous yeast proteins Lsb1 and Lsb2 bind the actin assembly protein Las17 (a yeast homolog of human Wiskott-Aldrich syndrome protein) and participate in the endocytic pathway. Lsb2 was shown to modulate maintenance of [PSI(+)] during and after heat shock. Here, we demonstrate that Lsb1 also regulates maintenance of the Sup35 prion during and after heat shock. These data point to the involvement of Lsb proteins in the partitioning of protein aggregates in stressed cells. Lsb1 abundance and cycling between actin patches, endoplasmic reticulum, and cytosol is regulated by the Guided Entry of Tail-anchored proteins pathway and Rsp5-dependent ubiquitination. Heat shock-induced proteolytic processing of Lsb1 is crucial for prion maintenance during stress. Our findings identify Lsb1 as another component of a tightly regulated pathway controlling protein aggregation in changing environments. PMID:25143386

Ali, Moiez; Chernova, Tatiana A; Newnam, Gary P; Yin, Luming; Shanks, John; Karpova, Tatiana S; Lee, Andrew; Laur, Oskar; Subramanian, Sindhu; Kim, Dami; McNally, James G; Seyfried, Nicholas T; Chernoff, Yury O; Wilkinson, Keith D

2014-10-01

181

The Functional Role of Prion Protein (PrPC) on Autophagy  

PubMed Central

Cellular prion protein (PrPC) plays an important role in the cellular defense against oxidative stress. However, the exact protective mechanism of PrPC is unclear. Autophagy is essential for survival, differentiation, development, and homeostasis in several organisms. Although the role that autophagy plays in neurodegenerative disease has yet to be established, it is clear that autophagy-induced cell death is observed in neurodegenerative disorders that exhibit protein aggregations. Moreover, autophagy can promote cell survival and cell death under various conditions. In this review, we describe the involvement of autophagy in prion disease and the effects of PrPC.

Shin, Hae-Young; Oh, Jae-Min; Kim, Yong-Sun

2013-01-01

182

NMR solution structure of the human prion protein Ralph Zahn, Aizhuo Liu*, Thorsten Lu hrs, Roland Riek, Christine von Schroetter, Francisco Lo pez Garcia, Martin Billeter  

E-print Network

NMR solution structure of the human prion protein Ralph Zahn, Aizhuo Liu*, Thorsten Lu¨ hrs, Roland The NMR structures of the recombinant human prion protein, hPrP(23­230), and two C-terminal fragments, h compared with the previously reported structures of the murine and Syrian hamster prion pro- teins

Wider, Gerhard

183

Journal of Molecular Graphics and Modelling 20 (2001) 169182 Flexibility of the murine prion protein and its Asp178Asn mutant  

E-print Network

Journal of Molecular Graphics and Modelling 20 (2001) 169­182 Flexibility of the murine prion,segregatewithspecificpointmutationsoftheprionprotein.Ithasbeenproposedthatthepathologically relevant Asp178Asn (D178N) mutation might destabilize the structure of the prion protein because-terminal domain of the murine prion protein and the D178N mutant were performed to investigate this hypothesis

Caflisch, Amedeo

184

Protein Conformation and Diagnostic Tests: The Prion Protein  

Microsoft Academic Search

Background: Many clinical diagnostic tests depend on the accurate detection and quantification of proteins and peptides and their functions. Alterations of protein structure, and the resulting consequences on dynamics, can affect the outcome of laboratory tests. These changes can be a result of mutations, other in vivo factors, or even the experimental conditions of the diagnostic test. Approach: The relationship

Brian J. Bennion; Valerie Daggett

2002-01-01

185

Post-translational hydroxylation at the N-terminus of the prion protein reveals presence of PPII structure in vivo  

PubMed Central

The transmissible spongiform encephalopathies are characterized by conversion of a host protein, PrPC (cellular prion protein), to a protease-resistant isoform, PrPSc (prion protein scrapie isoform). The importance of the highly flexible, N-terminal region of PrP has recently become more widely appreciated, particularly the biological activities associated with its metal ion-binding domain and its potential to form a poly(l-proline) II (PPII) helix. Circular dichroism spectroscopy of an N-terminal peptide, PrP37–53, showed that the PPII helix is formed in aqueous buffer; as it also contains an Xaa–Pro–Gly consensus sequence, it may act as a substrate for the collagen-modifying enzyme prolyl 4-hydroxylase. Direct evidence for this modification was obtained by mass spectrometry and Edman sequencing in recombinant mouse PrP secreted from stably transfected Chinese hamster ovary cells. Almost complete conversion of proline to 4-hydroxyproline occurs specifically at residue Pro44 of this murine protein; the same hydroxylated residue was detected, at lower levels, in PrPSc from the brains of scrapie-infected mice. Cation binding and/or post-translational hydroxylation of this region of PrP may regulate its role in the physiology and pathobiology of the cell. PMID:11032800

Gill, Andrew C.; Ritchie, Mark A.; Hunt, Lawrence G.; Steane, Sarah E.; Davies, Kenneth G.; Bocking, Sharon P.; Rhie, Alexandre G.O.; Bennett, Alan D.; Hope, James

2000-01-01

186

Transport of the pathogenic prion protein through soils.  

PubMed

Transmissible spongiform encephalopathies (TSEs) are progressive neurodegenerative diseases and include bovine spongiform encephalopathy of cattle, chronic wasting disease (CWD) of deer and elk, scrapie in sheep and goats, and Creutzfeldt-Jakob disease in humans. An abnormally folded form of the prion protein (designated PrP(TSE)) is typically associated with TSE infectivity and may constitute the major, if not sole, component of the infectious agent. Transmission of CWD and scrapie is mediated in part by an environmental reservoir of infectivity. Soil appears to be a plausible candidate for this reservoir. The transport of TSE agent through soil is expected to influence the accessibility of the pathogen to animals after deposition and must be understood to assess the risks associated with burial of infected carcasses. We report the results of saturated column experiments designed to evaluate PrP(TSE) transport through five soils with relatively high sand or silt contents and low organic carbon content. Protease-treated TSE-infected brain homogenate was used as a model for PrP(TSE) present in decomposing infected tissue. Synthetic rainwater was used as the eluent. All five soils retained PrP(TSE); no detectable PrP(TSE) was eluted over more than 40 pore volumes of flow. Lower bound apparent attachment coefficients were estimated for each soil. Our results suggest that TSE agent released from decomposing tissues to soils with low organic carbon content would remain near the site of initial deposition. In the case of infected carcasses deposited on the land surface, this may result in local sources of infectivity to other animals. PMID:20830901

Jacobson, Kurt H; Lee, Seunghak; Somerville, Robert A; McKenzie, Debbie; Benson, Craig H; Pedersen, Joel A

2010-01-01

187

Compartment-Restricted Biotinylation Reveals Novel Features of Prion Protein Metabolism in Vivo  

PubMed Central

Proteins are often made in more than one form, with alternate versions sometimes residing in different cellular compartments than the primary species. The mammalian prion protein (PrP), a cell surface GPI-anchored protein, is a particularly noteworthy example for which minor cytosolic and transmembrane forms have been implicated in disease pathogenesis. To study these minor species, we used a selective labeling strategy in which spatially restricted expression of a biotinylating enzyme was combined with asymmetric engineering of the cognate acceptor sequence into PrP. Using this method, we could show that even wild-type PrP generates small amounts of the CtmPrP transmembrane form. Selective detection of CtmPrP allowed us to reveal its N-terminal processing, long half-life, residence in both intracellular and cell surface locations, and eventual degradation in the lysosome. Surprisingly, some human disease-causing mutants in PrP selectively stabilized CtmPrP, revealing a previously unanticipated mechanism of CtmPrP up-regulation that may contribute to disease. Thus, spatiotemporal tagging has uncovered novel aspects of normal and mutant PrP metabolism and should be readily applicable to the analysis of minor topologic isoforms of other proteins. PMID:20980618

Emerman, Amy B.; Zhang, Zai-Rong; Chakrabarti, Oishee

2010-01-01

188

[Amyloidgenesis in prion disease].  

PubMed

Prion diseases, also known as transmissible spongiform encephalopathies, are defined as fatal, neurodegenerative, and transmissible disorders that affect both humans and other mammalian species. The central pathogenic mechanism in prion diseases is the conformational conversion of the normal prion protein, PrPC into the pathogenic prion protein, PrPSc. PrPSc, which is rich in ?-sheet structure, is resistant to proteinase degradation and hence, accumulates within the brain. These processes closely relate to development of the pathology of prion diseases such as spongiform change, gliosis, and neuronal loss. Prion diseases are thought to be a type of localized amyloidosis of the brain because prion protein itself is amyloidogenic. We herein discuss prion diseases from the point of view of localized amyloidosis. PMID:24998829

Tsuboi, Yoshio

2014-07-01

189

Prion protein self-interactions: a gateway to novel therapeutic strategies?  

PubMed

Transmissible spongiform encephalopathies (TSEs) or prion diseases are fatal neurodegenerative disorders and include among others Creutzfeldt-Jakob disease in humans, bovine spongiform encephalopathy (BSE) in cattle, and scrapie in sheep. The central event in disease development in TSEs is the refolding of the normal host-encoded cellular prion protein (PrP) into abnormal and disease associated prion protein. The agent is thought to consist mainly or exclusively of these pathologically folded PrP molecules. The exact molecular mechanisms underlying this process and the role of normal PrP in the conversion to pathological isoforms of PrP are still poorly understood. The highly conserved PrP gene structure and organisation however, suggests that its function is important, even though PrP knockout mice appear to develop normally. Conversion of normal PrP is initiated by interaction with abnormal PrP (or "agent") resulting in refolding of normal PrP into new pathological PrP ("agent replication"). Normal PrP was shown to interact/bind with many different molecules including metal ions, nucleic acids, several (receptor) proteins, and the prion protein itself. The processes underlying agent replication (normal to abnormal PrP conversion) are most likely initiated by selective interaction between PrP molecules and potentially influenced by chaperone molecules. Thus far no vaccine, disease reversing therapeutic compounds or strategies (cure) exists, although there are some compounds capable of slowing the progression of prion disease. Studies towards interference to date have primarily focussed on inference with the interaction between normal and pathological isoforms of PrP in order to develop therapeutic strategies or find compounds capable of inhibiting prion propagation. Most described strategies are either directed at depletion of normal PrP and thus preventing pathological PrP formation and accumulation, or are based on preventing interaction between normal and abnormal PrP. Other therapeutic strategies focus on selective (self-)interaction of normal PrP molecules. Increased understanding of these interactions and the processes in which normal PrP plays a (active) role, could potentially lead to new modes of inhibiting prion protein conversion in which the physiological function(s) of normal PrP is retained. Ultimately this may lead to therapeutic strategies that are effective not only as a prophylactic but also in later stages of prion disease development. Here we review the data underlying these PrP-based approaches. PMID:20932496

Rigter, Alan; Langeveld, Jan P M; van Zijderveld, Fred G; Bossers, Alex

2010-11-16

190

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

PubMed

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. PMID:16957924

Díaz-San Segundo, Fayna; Salguero, Francisco J; de Avila, Ana; Espinosa, Juan C; Torres, Juan M; Brun, Alejandro

2006-11-01

191

Variably Protease-Sensitive Prionopathy: A New Sporadic Disease of the Prion Protein  

PubMed Central

Objective The objective of the study is to report 2 new genotypic forms of protease-sensitive prionopathy (PSPr), a novel prion disease described in 2008, in 11 subjects all homozygous for valine at codon 129 of the prion protein (PrP) gene (129VV). The 2 new PSPr forms affect individuals who are either homozygous for methionine (129MM) or heterozygous for methionine/valine (129MV). Methods Fifteen affected subjects with 129MM, 129MV, and 129VV underwent comparative evaluation at the National Prion Disease Pathology Surveillance Center for clinical, histopathologic, immunohistochemical, genotypical, and PrP characteristics. Results Disease duration (between 22 and 45 months) was significantly different in the 129VV and 129MV subjects. Most other phenotypic features along with the PrP electrophoretic profile were similar but distinguishable in the 3 129 genotypes. A major difference laid in the sensitivity to protease digestion of the disease-associated PrP, which was high in 129VV but much lower, or altogether lacking, in 129MV and 129MM. This difference prompted the substitution of the original designation with “variably protease-sensitive prionopathy” (VPSPr). None of the subjects had mutations in the PrP gene coding region. Interpretation Because all 3 129 genotypes are involved, and are associated with distinguishable phenotypes, VPSPr becomes the second sporadic prion protein disease with this feature after Creutzfeldt-Jakob disease, originally reported in 1920. However, the characteristics of the abnormal prion protein suggest that VPSPr is different from typical prion diseases, and perhaps more akin to subtypes of Gerstmann-Sträussler-Scheinker disease. PMID:20695009

Zou, Wen-Quan; Puoti, Gianfranco; Xiao, Xiangzhu; Yuan, Jue; Qing, Liuting; Cali, Ignazio; Shimoji, Miyuki; Langeveld, Jan P. M.; Castellani, Rudy; Notari, Silvio; Crain, Barbara; Schmidt, Robert E.; Geschwind, Michael; DeArmond, Stephen J.; Cairns, Nigel J.; Dickson, Dennis; Honig, Lawrence; Torres, Juan Maria; Mastrianni, James; Capellari, Sabina; Giaccone, Giorgio; Belay, Ermias D.; Schonberger, Lawrence B.; Cohen, Mark; Perry, George; Kong, Qingzhong; Parchi, Piero; Tagliavini, Fabrizio; Gambetti, Pierluigi

2011-01-01

192

Transmissibility of Atypical Scrapie in Ovine Transgenic Mice: Major Effects of Host Prion Protein Expression and Donor Prion Genotype  

PubMed Central

Atypical scrapie or Nor98 has been identified as a transmissible spongiform encephalopathy (TSE) that is clearly distinguishable from classical scrapie and BSE, notably regarding the biochemical features of the protease-resistant prion protein PrPres and the genetic factors involved in susceptibility to the disease. In this study we transmitted the disease from a series of 12 French atypical scrapie isolates in a transgenic mouse model (TgOvPrP4) overexpressing in the brain ?0.25, 1.5 or 6× the levels of the PrPARQ ovine prion protein under the control of the neuron-specific enolase promoter. We used an approach based on serum PrPc measurements that appeared to reflect the different PrPc expression levels in the central nervous system. We found that transmission of atypical scrapie, much more than in classical scrapie or BSE, was strongly influenced by the PrPc expression levels of TgOvPrP4 inoculated mice. Whereas TgOvPrP4 mice overexpressing ?6× the normal PrPc level died after a survival periods of 400 days, those with ?1.5× the normal PrPc level died at around 700 days. The transmission of atypical scrapie in TgOvPrP4 mouse line was also strongly influenced by the prnp genotypes of the animal source of atypical scrapie. Isolates carrying the AF141RQ or AHQ alleles, associated with increased disease susceptibility in the natural host, showed a higher transmissibility in TgOvPrP4 mice. The biochemical analysis of PrPres in TgOvPrP4 mouse brains showed a fully conserved pattern, compared to that in the natural host, with three distinct PrPres products. Our results throw light on the transmission features of atypical scrapie and suggest that the risk of transmission is intrinsically lower than that of classical scrapie or BSE, especially in relation to the expression level of the prion protein. PMID:19806224

Arsac, Jean-Noel; Betemps, Dominique; Morignat, Eric; Feraudet, Cecile; Bencsik, Anna; Aubert, Denise; Grassi, Jacques; Baron, Thierry

2009-01-01

193

Inactivation of Template-Directed Misfolding of Infectious Prion Protein by Ozone  

PubMed Central

Misfolded prions (PrPSc) are well known for their resistance to conventional decontamination processes. The potential risk of contamination of the water environment, as a result of disposal of specified risk materials (SRM), has raised public concerns. Ozone is commonly utilized in the water industry for inactivation of microbial contaminants and was tested in this study for its ability to inactivate prions (263K hamster scrapie = PrPSc). Treatment variables included initial ozone dose (7.6 to 25.7 mg/liter), contact time (5 s and 5 min), temperature (4°C and 20°C), and pH (pH 4.4, 6.0, and 8.0). Exposure of dilute suspensions of the infected 263K hamster brain homogenates (IBH) (0.01%) to ozone resulted in the in vitro destruction of the templating properties of PrPSc, as measured by the protein misfolding cyclic amplification (PMCA) assay. The highest levels of prion inactivation (?4 log10) were observed with ozone doses of 13.0 mg/liter, at pH 4.4 and 20°C, resulting in a CT (the product of residual ozone concentration and contact time) value as low as 0.59 mg · liter?1 min. A comparison of ozone CT requirements among various pathogens suggests that prions are more susceptible to ozone degradation than some model bacteria and protozoa and that ozone treatment may be an effective solution for inactivating prions in water and wastewater. PMID:22138993

Ding, Ning; Price, Luke M.; Braithwaite, Shannon L.; Balachandran, Aru; Belosevic, Miodrag

2012-01-01

194

Heterogeneity of the Abnormal Prion Protein (PrPSc) of the Chandler Scrapie Strain  

PubMed Central

The pathological prion protein, PrPSc, displays various sizes of aggregates. In this study, we investigated the conformation, aggregation stability and proteinase K (PK)-sensitivity of small and large PrPSc aggregates of mouse-adapted prion strains. We showed that small PrPSc aggregates, previously thought to be PK-sensitive, are resistant to PK digestion. Furthermore, we showed that small PrPSc aggregates of the Chandler scrapie strain have greater resistance to PK digestion and aggregation-denaturation than large PrPSc aggregates of this strain. We conclude that this strain consists of heterogeneous PrPSc.

Kasai, Kazuo; Iwamaru, Yoshifumi; Masujin, Kentaro; Imamura, Morikazu; Mohri, Shirou; Yokoyama, Takashi

2013-01-01

195

Low Density Subcellular Fractions Enhance Disease-specific Prion Protein Misfolding  

PubMed Central

The production of prion particles in vitro by amplification with or without exogenous seed typically results in infectivity titers less than those associated with PrPSc isolated ex vivo and highlights the potential role of co-factors that can catalyze disease-specific prion protein misfolding in vivo. We used a cell-free conversion assay previously shown to replicate many aspects of transmissible spongiform encephalopathy disease to investigate the cellular location of disease-specific co-factors using fractions derived from gradient centrifugation of a scrapie-susceptible cell line. Fractions from the low density region of the gradient doubled the efficiency of conversion of recombinant PrP. These fractions contain plasma membrane and cytoplasmic proteins, and conversion enhancement can be achieved using PrPSc derived from two different strains of mouse-passaged scrapie as seed. Equivalent fractions from a second scrapie-susceptible cell line also stimulate conversion. We also show that subcellular fractions enhancing disease-specific prion protein conversion prevent in vitro fibrillization of recombinant prion protein, suggesting the existence of separate, competing mechanisms of disease-specific and nonspecific misfolding in vivo. PMID:20106973

Graham, James F.; Agarwal, Sonya; Kurian, Dominic; Kirby, Louise; Pinheiro, Teresa J. T.; Gill, Andrew C.

2010-01-01

196

Prion induction involves an ancient system for the sequestration of aggregated proteins and heritable  

E-print Network

rise to new, often benefi- cial, phenotypes. Evidence suggests that prion induction involves a process long uninterrupted bundles of fibers, whereas Dot fibers were highly fragmented. Both forms were of nucleic acids. In fungi, they act as protein-based elements of heredity, stably propagating their altered

Lindquist, Susan

197

Prion Domain of Yeast Ure2 Protein Adopts a Completely Disordered Structure: A Solid-Support EPR Study  

PubMed Central

Amyloid fibril formation is associated with a range of neurodegenerative diseases in humans, including Alzheimer’s, Parkinson’s, and prion diseases. In yeast, amyloid underlies several non-Mendelian phenotypes referred to as yeast prions. Mechanism of amyloid formation is critical for a complete understanding of the yeast prion phenomenon and human amyloid-related diseases. Ure2 protein is the basis of yeast prion [URE3]. The Ure2p prion domain is largely disordered. Residual structures, if any, in the disordered region may play an important role in the aggregation process. Studies of Ure2p prion domain are complicated by its high aggregation propensity, which results in a mixture of monomer and aggregates in solution. Previously we have developed a solid-support electron paramagnetic resonance (EPR) approach to address this problem and have identified a structured state for the Alzheimer’s amyloid-? monomer. Here we use solid-support EPR to study the structure of Ure2p prion domain. EPR spectra of Ure2p prion domain with spin labels at every fifth residue from position 10 to position 75 show similar residue mobility profile for denaturing and native buffers after accounting for the effect of solution viscosity. These results suggest that Ure2p prion domain adopts a completely disordered structure in the native buffer. A completely disordered Ure2p prion domain implies that the amyloid formation of Ure2p, and likely other Q/N-rich yeast prion proteins, is primarily driven by inter-molecular interactions. PMID:23077577

Ngo, Sam; Chiang, Vicky; Ho, Elaine; Le, Linh; Guo, Zhefeng

2012-01-01

198

Detection of prion protein particles in blood plasma of scrapie infected sheep.  

PubMed

Prion diseases are transmissible neurodegenerative diseases affecting humans and animals. The agent of the disease is the prion consisting mainly, if not solely, of a misfolded and aggregated isoform of the host-encoded prion protein (PrP). Transmission of prions can occur naturally but also accidentally, e.g. by blood transfusion, which has raised serious concerns about blood product safety and emphasized the need for a reliable diagnostic test. In this report we present a method based on surface-FIDA (fluorescence intensity distribution analysis), that exploits the high state of molecular aggregation of PrP as an unequivocal diagnostic marker of the disease, and show that it can detect infection in blood. To prepare PrP aggregates from blood plasma we introduced a detergent and lipase treatment to separate PrP from blood lipophilic components. Prion protein aggregates were subsequently precipitated by phosphotungstic acid, immobilized on a glass surface by covalently bound capture antibodies, and finally labeled with fluorescent antibody probes. Individual PrP aggregates were visualized by laser scanning microscopy where signal intensity was proportional to aggregate size. After signal processing to remove the background from low fluorescence particles, fluorescence intensities of all remaining PrP particles were summed. We detected PrP aggregates in plasma samples from six out of ten scrapie-positive sheep with no false positives from uninfected sheep. Applying simultaneous intensity and size discrimination, ten out of ten samples from scrapie sheep could be differentiated from uninfected sheep. The implications for ante mortem diagnosis of prion diseases are discussed. PMID:22567169

Bannach, Oliver; Birkmann, Eva; Reinartz, Elke; Jaeger, Karl-Erich; Langeveld, Jan P M; Rohwer, Robert G; Gregori, Luisa; Terry, Linda A; Willbold, Dieter; Riesner, Detlev

2012-01-01

199

Detection of Prion Protein Particles in Blood Plasma of Scrapie Infected Sheep  

PubMed Central

Prion diseases are transmissible neurodegenerative diseases affecting humans and animals. The agent of the disease is the prion consisting mainly, if not solely, of a misfolded and aggregated isoform of the host-encoded prion protein (PrP). Transmission of prions can occur naturally but also accidentally, e.g. by blood transfusion, which has raised serious concerns about blood product safety and emphasized the need for a reliable diagnostic test. In this report we present a method based on surface-FIDA (fluorescence intensity distribution analysis), that exploits the high state of molecular aggregation of PrP as an unequivocal diagnostic marker of the disease, and show that it can detect infection in blood. To prepare PrP aggregates from blood plasma we introduced a detergent and lipase treatment to separate PrP from blood lipophilic components. Prion protein aggregates were subsequently precipitated by phosphotungstic acid, immobilized on a glass surface by covalently bound capture antibodies, and finally labeled with fluorescent antibody probes. Individual PrP aggregates were visualized by laser scanning microscopy where signal intensity was proportional to aggregate size. After signal processing to remove the background from low fluorescence particles, fluorescence intensities of all remaining PrP particles were summed. We detected PrP aggregates in plasma samples from six out of ten scrapie-positive sheep with no false positives from uninfected sheep. Applying simultaneous intensity and size discrimination, ten out of ten samples from scrapie sheep could be differentiated from uninfected sheep. The implications for ante mortem diagnosis of prion diseases are discussed. PMID:22567169

Reinartz, Elke; Jaeger, Karl-Erich; Langeveld, Jan P. M.; Rohwer, Robert G.; Gregori, Luisa; Terry, Linda A.; Willbold, Dieter; Riesner, Detlev

2012-01-01

200

New Insights into Metal Interactions with the Prion Protein  

PubMed Central

Copper coordination to the prion protein (PrP) has garnered considerable interest for almost 20 years, due in part to the possibility that this interaction may be part of the normal function of PrP. The most characterized form of copper binding to PrP has been Cu2+ interaction with the conserved tandem repeats in the N-terminal domain of PrP, termed the octarepeats, with many studies focusing on single and multiple repeats of PHGGGWGQ. Extended X-ray absorption fine structure (EXAFS) spectroscopy has been used in several previous instances to characterize the solution structure of Cu2+ binding into the peptide backbone in the HGGG portion of the octarepeats. All previous EXAFS studies, however, have benefitted from crystallographic structure information for [CuII (Ac-HGGGW-NH2)–2H], but have not conclusively demonstrated that the complex EXAFS spectrum represents the same coordination environment for Cu2+ bound to the peptide backbone. Density functional structure calculations as well as full multiple scattering EXAFS curve fitting analysis are brought to bear on the predominant coordination mode for Cu2+ with the Ac-PHGGGWGQ-NH2 peptide at physiological pH, under high Cu2+ occupancy conditions. In addition to the structure calculations, which provide a thermodynamic link to structural information, methods are also presented for extensive deconvolution of the EXAFS spectrum. We demonstrate how the EXAFS data can be analyzed to extract the maximum structural information and arrive at a structural model that is significantly improved over previous EXAFS characterizations. The EXAFS spectrum for the chemically reduced form of copper binding to the Ac-PHGGGWGQ-NH2 peptide is presented, which is best modeled as a linear 2-coordinate species with a single His imidazole ligand and a water molecule. The extent of in situ photoreduction of the copper center during standard data collection is also presented and EXAFS curve fitting of the photoreduced species reveals an intermediate structure that is similar to the Cu2+ form with reduced coordination number. PMID:24102071

McDonald, Alex; Pushie, M. Jake; Millhauser, Glenn L.; George, Graham N.

2013-01-01

201

Cellular prion protein regulates ?-secretase cleavage of the Alzheimer's amyloid precursor protein  

PubMed Central

Proteolytic processing of the amyloid precursor protein (APP) by ?-secretase, ?-site APP cleaving enzyme (BACE1), is the initial step in the production of the amyloid ? (A?) peptide, which is involved in the pathogenesis of Alzheimer's disease. The normal cellular function of the prion protein (PrPC), the causative agent of the transmissible spongiform encephalopathies such as Creutzfeldt–Jakob disease in humans, remains enigmatic. Because both APP and PrPC are subject to proteolytic processing by the same zinc metalloproteases, we tested the involvement of PrPC in the proteolytic processing of APP. Cellular overexpression of PrPC inhibited the ?-secretase cleavage of APP and reduced A? formation. Conversely, depletion of PrPC in mouse N2a cells by siRNA led to an increase in A? peptides secreted into the medium. In the brains of PrP knockout mice and in the brains from two strains of scrapie-infected mice, A? levels were significantly increased. Two mutants of PrP, PG14 and A116V, that are associated with familial human prion diseases failed to inhibit the ?-secretase cleavage of APP. Using constructs of PrP, we show that this regulatory effect of PrPC on the ?-secretase cleavage of APP required the localization of PrPC to cholesterol-rich lipid rafts and was mediated by the N-terminal polybasic region of PrPC via interaction with glycosaminoglycans. In conclusion, this is a mechanism by which the cellular production of the neurotoxic A? is regulated by PrPC and may have implications for both Alzheimer's and prion diseases. PMID:17573534

Parkin, Edward T.; Watt, Nicole T.; Hussain, Ishrut; Eckman, Elizabeth A.; Eckman, Christopher B.; Manson, Jean C.; Baybutt, Herbert N.; Turner, Anthony J.; Hooper, Nigel M.

2007-01-01

202

Differential Contribution of Superoxide Dismutase Activity by Prion Protein in Vivo  

Microsoft Academic Search

Normal prion protein (PrPC) is a copper binding protein and may play a role in cellular resistance to oxidative stress. Recently, copper-bound recombinant PrPC has been shown to exhibit superoxide dismutase (SOD)-like activity. However, as PrPC affinity for copper is low in comparison to other cupro-proteins, the question remains as to whether PrPC could contribute SOD activity in vivo. To

Boon-Seng Wong; Tao Pan; Tong Liu; Ruliang Li; Pierluigi Gambetti; Man-Sun Sy

2000-01-01

203

A bipolar functionality of Q/N-rich proteins: Lsm4 amyloid causes clearance of yeast prions  

PubMed Central

Prions are epigenetic modifiers that cause partially loss-of-function phenotypes of the proteins in Saccharomyces cerevisiae. The molecular chaperone network that supports prion propagation in the cell has seen a great progress in the last decade. However, the cellular machinery to activate or deactivate the prion states remains an enigma, largely due to insufficient knowledge of prion-regulating factors. Here, we report that overexpression of a [PSI+]-inducible Q/N-rich protein, Lsm4, eliminates the three major prions [PSI+], [URE3], and [RNQ+]. Subcloning analysis revealed that the Q/N-rich region of Lsm4 is responsible for the prion loss. Lsm4 formed an amyloid in vivo, which seemed to play a crucial role in the prion elimination. Fluorescence correlation spectroscopy analysis revealed that in the course of the Lsm4-driven [PSI+] elimination, the [PSI+] aggregates undergo a size increase, which ultimately results in the formation of conspicuous foci in otherwise [psi?]-like mother cells. We also found that the antiprion activity is a general property of [PSI+]-inducible factors. These data provoked a novel “unified” model that explains both prion induction and elimination by a single scheme. PMID:23512891

Oishi, Keita; Kurahashi, Hiroshi; Pack, Chan-Gi; Sako, Yasushi; Nakamura, Yoshikazu

2013-01-01

204

Prion propagation in mice expressing human and chimeric PrP transgenes implicates the interaction of cellular PrP with another protein  

Microsoft Academic Search

Transgenic (Tg) mice expressing human (Hu) and chimeric prion protein (PrP) genes were inoculated with brain extracts from humans with inherited or sporadic prion disease to investigate the mechanism by which PrPc is transformed into PrPSc. Although Tg(HuPrP) mice expressed high levels of HuPrPc, they were resistant to human prions. They became susceptible to human prions upon ablation of the

Glenn C. Telling; Michael Scott; James Mastrianni; Ruth Gabizon; Marilyn Torchia; Fred E. Cohen; Stephen J. DeArmond

1995-01-01

205

Linkage of prion protein and scrapie incubation time genes  

Microsoft Academic Search

A single gene (Prn-i) that affects scrapie incubation period in mice has been identified. I\\/LnJ mice have a very long incubation period after inoculation of scrapie prions (200-385 days) and NZW\\/LacJ mice have a short one (113 +\\/- 2.8 days). (NZW X I\\/Ln)F1 hybrid mice had incubation times of 223 +\\/- 2.8 days indicating longer incubation times were dominant. Incubation

G A Carlson; D T Kingsbury; P A Goodman; S Coleman; S T Marshall; S DeArmond; D Westaway; S B Prusiner

1986-01-01

206

Assessing Proteinase K Resistance of Fish Prion Proteins in a Scrapie-Infected Mouse Neuroblastoma Cell Line  

PubMed Central

The key event in prion pathogenesis is the structural conversion of the normal cellular protein, PrPC, into an aberrant and partially proteinase K resistant isoform, PrPSc. Since the minimum requirement for a prion disease phenotype is the expression of endogenous PrP in the host, species carrying orthologue prion genes, such as fish, could in theory support prion pathogenesis. Our previous work has demonstrated the development of abnormal protein deposition in sea bream brain, following oral challenge of the fish with natural prion infectious material. In this study, we used a prion-infected mouse neuroblastoma cell line for the expression of three different mature fish PrP proteins and the evaluation of the resistance of the exogenously expressed proteins to proteinase K treatment (PK), as an indicator of a possible prion conversion. No evidence of resistance to PK was detected for any of the studied recombinant proteins. Although not indicative of an absolute inability of the fish PrPs to structurally convert to pathogenic isoforms, the absence of PK-resistance may be due to supramolecular and conformational differences between the mammalian and piscine PrPs. PMID:25402173

Salta, Evgenia; Kanata, Eirini; Ouzounis, Christos A.; Gilch, Sabine; Schätzl, Hermann; Sklaviadis, Theodoros

2014-01-01

207

Assessing proteinase k resistance of fish prion proteins in a scrapie-infected mouse neuroblastoma cell line.  

PubMed

The key event in prion pathogenesis is the structural conversion of the normal cellular protein, PrPC, into an aberrant and partially proteinase K resistant isoform, PrPSc. Since the minimum requirement for a prion disease phenotype is the expression of endogenous PrP in the host, species carrying orthologue prion genes, such as fish, could in theory support prion pathogenesis. Our previous work has demonstrated the development of abnormal protein deposition in sea bream brain, following oral challenge of the fish with natural prion infectious material. In this study, we used a prion-infected mouse neuroblastoma cell line for the expression of three different mature fish PrP proteins and the evaluation of the resistance of the exogenously expressed proteins to proteinase K treatment (PK), as an indicator of a possible prion conversion. No evidence of resistance to PK was detected for any of the studied recombinant proteins. Although not indicative of an absolute inability of the fish PrPs to structurally convert to pathogenic isoforms, the absence of PK-resistance may be due to supramolecular and conformational differences between the mammalian and piscine PrPs. PMID:25402173

Salta, Evgenia; Kanata, Eirini; Ouzounis, Christos A; Gilch, Sabine; Schätzl, Hermann; Sklaviadis, Theodoros

2014-01-01

208

Development of techniques in magnetic resonance and structural studies of the prion protein  

SciTech Connect

Magnetic resonance is the most powerful analytical tool used by chemists today. Its applications range from determining structures of large biomolecules to imaging of human brains. Nevertheless, magnetic resonance remains a relatively young field, in which many techniques are currently being developed that have broad applications. In this dissertation, two new techniques are presented, one that enables the determination of torsion angles in solid-state peptides and proteins, and another that involves imaging of heterogenous materials at ultra-low magnetic fields. In addition, structural studies of the prion protein via solid-state NMR are described. More specifically, work is presented in which the dependence of chemical shifts on local molecular structure is used to predict chemical shift tensors in solid-state peptides with theoretical ab initio surfaces. These predictions are then used to determine the backbone dihedral angles in peptides. This method utilizes the theoretical chemicalshift tensors and experimentally determined chemical-shift anisotropies (CSAs) to predict the backbone and side chain torsion angles in alanine, leucine, and valine residues. Additionally, structural studies of prion protein fragments are described in which conformationally-dependent chemical-shift measurements were made to gain insight into the structural differences between the various conformational states of the prion protein. These studies are of biological and pathological interest since conformational changes in the prion protein are believed to cause prion diseases. Finally, an ultra-low field magnetic resonance imaging technique is described that enables imaging and characterization of heterogeneous and porous media. The notion of imaging gases at ultra-low fields would appear to be very difficult due to the prohibitively low polarization and spin densities as well as the low sensitivities of conventional Faraday coil detectors. However, Chapter 5 describes how gas imaging at ultra-low fields is realized by incorporating the high sensitivities of a dc superconducting quantum interference device (SQUID) with the high polarizations attainable through optica11y pumping {sup 129}Xe gas.

Bitter, Hans-Marcus L.

2000-07-01

209

Role of Lipid Rafts and GM1 in the Segregation and Processing of Prion Protein  

PubMed Central

The prion protein (PrPC) is highly expressed within the nervous system. Similar to other GPI-anchored proteins, PrPC is found in lipid rafts, membrane domains enriched in cholesterol and sphingolipids. PrPC raft association, together with raft lipid composition, appears essential for the conversion of PrPC into the scrapie isoform PrPSc, and the development of prion disease. Controversial findings were reported on the nature of PrPC-containing rafts, as well as on the distribution of PrPC between rafts and non-raft membranes. We investigated PrPC/ganglioside relationships and their influence on PrPC localization in a neuronal cellular model, cerebellar granule cells. Our findings argue that in these cells at least two PrPC conformations coexist: in lipid rafts PrPC is present in the native folding (?-helical), stabilized by chemico-physical condition, while it is mainly present in other membrane compartments in a PrPSc-like conformation. We verified, by means of antibody reactivity and circular dichroism spectroscopy, that changes in lipid raft-ganglioside content alters PrPC conformation and interaction with lipid bilayers, without modifying PrPC distribution or cleavage. Our data provide new insights into the cellular mechanism of prion conversion and suggest that GM1-prion protein interaction at the cell surface could play a significant role in the mechanism predisposing to pathology. PMID:24859148

Botto, Laura; Cunati, Diana; Coco, Silvia; Sesana, Silvia; Bulbarelli, Alessandra; Biasini, Emiliano; Colombo, Laura; Negro, Alessandro; Chiesa, Roberto; Masserini, Massimo; Palestini, Paola

2014-01-01

210

N-terminal domain of prion protein directs its oligomeric association.  

PubMed

The self-association of prion protein (PrP) is a critical step in the pathology of prion diseases. It is increasingly recognized that small non-fibrillar ?-sheet-rich oligomers of PrP may be of crucial importance in the prion disease process. Here, we characterize the structure of a well defined ?-sheet-rich oligomer, containing ?12 PrP molecules, and often enclosing a central cavity, formed using full-length recombinant PrP. The N-terminal region of prion protein (residues 23-90) is required for the formation of this distinct oligomer; a truncated form comprising residues 91-231 forms a broad distribution of aggregated species. No infectivity or toxicity was found using cell and animal model systems. This study demonstrates that examination of the full repertoire of conformers and assembly states that can be accessed by PrP under specific experimental conditions should ideally be done using the full-length protein. PMID:25074940

Trevitt, Clare R; Hosszu, Laszlo L P; Batchelor, Mark; Panico, Silvia; Terry, Cassandra; Nicoll, Andrew J; Risse, Emmanuel; Taylor, William A; Sandberg, Malin K; Al-Doujaily, Huda; Linehan, Jacqueline M; Saibil, Helen R; Scott, David J; Collinge, John; Waltho, Jonathan P; Clarke, Anthony R

2014-09-12

211

Translation of the Prion Protein mRNA Is Robust in Astrocytes but Does Not Amplify during Reactive Astrocytosis in the Mouse Brain  

PubMed Central

Prion diseases induce neurodegeneration in specific brain areas for undetermined reasons. A thorough understanding of the localization of the disease-causing molecule, the prion protein (PrP), could inform on this issue but previous studies have generated conflicting conclusions. One of the more intriguing disagreements is whether PrP is synthesized by astrocytes. We developed a knock-in reporter mouse line in which the coding sequence of the PrP expressing gene (Prnp), was replaced with that for green fluorescent protein (GFP). Native GFP fluorescence intensity varied between and within brain regions. GFP was present in astrocytes but did not increase during reactive gliosis induced by scrapie prion infection. Therefore, reactive gliosis associated with prion diseases does not cause an acceleration of local PrP production. In addition to aiding in Prnp gene activity studies, this reporter mouse line will likely prove useful for analysis of chimeric animals produced by stem cell and tissue transplantation experiments. PMID:24752288

Jackson, Walker S.; Krost, Clemens; Borkowski, Andrew W.; Kaczmarczyk, Lech

2014-01-01

212

Cellular Prion Protein Expression Is Not Regulated by the Alzheimer's Amyloid Precursor Protein Intracellular Domain  

PubMed Central

There is increasing evidence of molecular and cellular links between Alzheimer's disease (AD) and prion diseases. The cellular prion protein, PrPC, modulates the post-translational processing of the AD amyloid precursor protein (APP), through its inhibition of the ?-secretase BACE1, and oligomers of amyloid-? bind to PrPC which may mediate amyloid-? neurotoxicity. In addition, the APP intracellular domain (AICD), which acts as a transcriptional regulator, has been reported to control the expression of PrPC. Through the use of transgenic mice, cell culture models and manipulation of APP expression and processing, this study aimed to clarify the role of AICD in regulating PrPC. Over-expression of the three major isoforms of human APP (APP695, APP751 and APP770) in cultured neuronal and non-neuronal cells had no effect on the level of endogenous PrPC. Furthermore, analysis of brain tissue from transgenic mice over-expressing either wild type or familial AD associated mutant human APP revealed unaltered PrPC levels. Knockdown of endogenous APP expression in cells by siRNA or inhibition of ?-secretase activity also had no effect on PrPC levels. Overall, we did not detect any significant difference in the expression of PrPC in any of the cell or animal-based paradigms considered, indicating that the control of cellular PrPC levels by AICD is not as straightforward as previously suggested. PMID:22363722

Lewis, Victoria; Whitehouse, Isobel J.; Baybutt, Herbert; Manson, Jean C.; Collins, Steven J.; Hooper, Nigel M.

2012-01-01

213

Normal levels of the antiprion proteins Btn2 and Cur1 cure most newly formed [URE3] prion variants.  

PubMed

[URE3] is an amyloid prion of the Saccharomyces cerevisiae Ure2p, a regulator of nitrogen catabolism. Overproduction of Btn2p, involved in late endosome to Golgi protein transport, or its paralog Cur1p, cures [URE3]. Btn2p, in curing, is colocalized with Ure2p in a single locus, suggesting sequestration of Ure2p amyloid filaments. We find that most [URE3] variants generated in a btn2 cur1 double mutant are cured by restoring normal levels of Btn2p and Cur1p, with both proteins needed for efficient curing. The [URE3] variants cured by normal levels of Btn2p and Cur1p all have low seed number, again suggesting a seed sequestration mechanism. Hsp42 overproduction also cures [URE3], and Hsp42p aids Btn2 overproduction curing. Cur1p is needed for Hsp42 overproduction curing of [URE3], but neither Btn2p nor Cur1p is needed for overproduction curing by the other. Although hsp42? strains stably propagate [URE3-1], hsp26? destabilizes this prion. Thus, Btn2p and Cur1p are antiprion system components at their normal levels, acting with Hsp42. Btn2p is related in sequence to human Hook proteins, involved in aggresome formation and other transport activities. PMID:24938787

Wickner, Reed B; Bezsonov, Evgeny; Bateman, David A

2014-07-01

214

IDENTIFICATION AND REMOVAL OF PROTEINS THAT CO-PURIFY WITH INFECTIOUS PRION PROTEIN IMPROVES THE ANALYSIS OF ITS SECONDARY STRUCTURE  

PubMed Central

Prion diseases are neurodegenerative disorders associated with the accumulation of an abnormal isoform of the mammalian prion protein (PrP). Fourier transform infrared spectroscopy (FTIR) has previously been used to show that the conformation of aggregated, infectious PrP (PrPSc) varies between prion strains and these unique conformations may determine strain-specific disease phenotypes. However, the relative amounts of ?-helix, ?-sheet and other secondary structures have not always been consistent between studies suggesting that other proteins might be confounding the analysis of PrPSc secondary structure. We have used FTIR and tandem mass spectrometry to analyze enriched PrPSc from mouse and hamster prion strains both before and after the removal of protein contaminants that commonly co-purify with PrPSc. Our data show that non-PrP proteins do contribute to absorbances that have been associated with ?-helical, loop, turn, and ?-sheet structures attributed to PrPSc. The major contaminant, the ?-helical protein ferritin, absorbs strongly at 1652cm?1 in the FTIR spectrum associated with PrPSc. However, even the removal of greater than 99% of the ferritin from PrPSc did not completely abolish absorbance at 1652cm?1. Our results show that contaminating proteins alter the FTIR spectrum attributed to PrPSc and suggest that the ?-helical, loop/turn, and ?-sheet secondary structure that remains following their removal are derived from PrPSc itself. PMID:21805638

Moore, Roger A.; Timmes, Andrew; Wilmarth, Phillip A.; Safronetz, David; Priola, Suzette A.

2013-01-01

215

Caffeine prevents human prion protein-mediated neurotoxicity through the induction of autophagy.  

PubMed

The human prion protein (PrP) fragment PrP(106?126) possesses the majority of the pathogenic properties associated with the infectious scrapie isoform of PrP, known as PrPSc. The accumulation of PrPSc in the brain of humans and animals affects the central nervous system. Recent epidemiological studies have suggested that caffeine, one of the major components of coffee, exerts protective effects against the development of neurodegeneration. However, the protective effects of caffeine against prion disease have not been reported to date. In this study, we therefore investigated the effects of caffeine on PrP-mediated neurotoxicity. The protein expression of the autophagosomal marker, LC3-II, was increased by caffeine in a dose-dependent manner, and the autophagy induced by caffeine protected the neuronal cells against PrP(106?126)?induced cell death. On the contrary, the downregulation of LC3-II using the autophagy inhibitors, 3-methyladenine (3-??) and wortmannin, prevented the caffeine-mediated neuroprotective effects. To the best of our knowledge, the present study provides the first evidence that treatment with caffeine protects human neuronal cells against prion?mediated neurotoxicity and these neuroprotective effects are mediated by caffeine-induced autophagy signals. Our data suggest that treatment with caffeine may be a novel therapeutic strategy for prion peptide?induced apoptosis. PMID:24938171

Moon, Ji-Hong; Lee, Ju-Hee; Park, Jin-Young; Kim, Sung-Wook; Lee, You-Jin; Kang, Seog-Jin; Seol, Jae-Won; Ahn, Dong-Choon; Park, Sang-Youel

2014-08-01

216

Identification of Misfolded Proteins in Body Fluids for the Diagnosis of Prion Diseases  

PubMed Central

Transmissible spongiform encephalopathy (TSE) or prion diseases are fatal rare neurodegenerative disorders affecting man and animals and caused by a transmissible infectious agent. TSE diseases are characterized by spongiform brain lesions with neuronal loss and the abnormal deposition in the CNS, and to less extent in other tissues, of an insoluble and protease resistant form of the cellular prion protein (PrPC), named PrPTSE. In man, TSE diseases affect usually people over 60 years of age with no evident disease-associated risk factors. In some cases, however, TSE diseases are unequivocally linked to infectious episodes related to the use of prion-contaminated medicines, medical devices, or meat products as in the variant Creutzfeldt-Jakob disease (CJD). Clinical signs occur months or years after infection, and during this silent period PrPTSE, the only reliable marker of infection, is not easily measurable in blood or other accessible tissues or body fluids causing public health concerns. To overcome the limit of PrPTSE detection, several highly sensitive assays have been developed, but attempts to apply these techniques to blood of infected hosts have been unsuccessful or not yet validated. An update on the latest advances for the detection of misfolded prion protein in body fluids is provided. PMID:24027585

Pocchiari, Maurizio

2013-01-01

217

Lentivector-mediated RNAi efficiently suppresses prion protein and prolongs survival of scrapie-infected mice  

PubMed Central

Prion diseases are fatal neurodegenerative diseases characterized by the accumulation of PrPSc, the infectious and protease-resistant form of the cellular prion protein (PrPC). We generated lentivectors expressing PrPC-specific short hairpin RNAs (shRNAs) that efficiently silenced expression of the prion protein gene (Prnp) in primary neuronal cells. Treatment of scrapie-infected neuronal cells with these lentivectors resulted in an efficient and stable suppression of PrPSc accumulation. After intracranial injection, lentiviral shRNA reduced PrPC expression in transgenic mice carrying multiple copies of Prnp. To test the therapeutic potential of lentiviral shRNA, we used what we believe to be a novel approach in which the clinical situation was mimicked. We generated chimeric mice derived from lentivector-transduced embryonic stem cells. Depending on the degree of chimerism, these animals carried the lentiviral shRNAs in a certain percentage of brain cells and expressed reduced levels of PrPC. Importantly, in highly chimeric mice, survival after scrapie infection was significantly extended. Taken together, these data suggest that lentivector-mediated RNA interference could be an approach for the treatment of prion disease. PMID:17143329

Pfeifer, Alexander; Eigenbrod, Sabina; Al-Khadra, Saba; Hofmann, Andreas; Mitteregger, Gerda; Moser, Markus; Bertsch, Uwe; Kretzschmar, Hans

2006-01-01

218

Ultrasensitive detection of pathological prion protein aggregates by dual-color scanning for intensely fluorescent targets  

PubMed Central

A definite diagnosis of prion diseases such as Creutzfeldt–Jakob disease (CJD) relies on the detection of pathological prion protein (PrPSc). However, no test for PrPSc in cerebrospinal fluid (CSF) has been available thus far. Based on a setup for confocal dual-color fluorescence correlation spectroscopy, a technique suitable for single molecule detection, we developed a highly sensitive detection method for PrPSc. Pathological prion protein aggregates were labeled by specific antibody probes tagged with fluorescent dyes, resulting in intensely fluorescent targets, which were measured by dual-color fluorescence intensity distribution analysis in a confocal scanning setup. In a diagnostic model system, PrPSc aggregates were detected down to a concentration of 2 pM PrPSc, corresponding to an aggregate concentration of approximately 2 fM, which was more than one order of magnitude more sensitive than Western blot analysis. A PrPSc-specific signal could also be detected in a number of CSF samples from patients with CJD but not in control samples, providing the basis for a rapid and specific test for CJD and other prion diseases. Furthermore, this method could be adapted to the sensitive detection of other disease-associated amyloid aggregates such as in Alzheimer's disease. PMID:10805803

Bieschke, J.; Giese, A.; Schulz-Schaeffer, W.; Zerr, I.; Poser, S.; Eigen, M.; Kretzschmar, H.

2000-01-01

219

Fatal familial insomnia and familial Creutzfeldt-Jakob disease: different prion proteins determined by a DNA polymorphism.  

PubMed Central

Fatal familial insomnia and a subtype of Creutzfeldt-Jakob disease, two clinically and pathologically distinct diseases, are linked to the same mutation at codon 178 (Asp-178-->Asn) but segregate with different genotypes determined by this mutation and the methionine-valine polymorphism at codon 129 of the prion protein gene. The abnormal isoforms of the prion protein in these two diseases were found to differ both in the relative abundance of glycosylated forms and in the size of the protease-resistant fragments. The size difference was consistent with a different protease cleavage site, suggesting a different conformation of the protease-resistant prion protein present in the two diseases. These differences are likely to be responsible for the type and location of the lesions that characterize these two diseases. Therefore, the combination of the mutation at codon 178 and the polymorphism at codon 129 determines the disease phenotype by producing two altered conformations of the prion protein. Images PMID:7908444

Monari, L; Chen, S G; Brown, P; Parchi, P; Petersen, R B; Mikol, J; Gray, F; Cortelli, P; Montagna, P; Ghetti, B

1994-01-01

220

Synthesis and structural characterization of a mimetic membrane-anchored prion protein  

Microsoft Academic Search

During pathogenesis of transmissible spongiform encephalopathies (TSEs) an abnormal form (PrPSc) of the host encoded prion protein (PrPC) accu- mulates in insoluble fibrils and plaques. The two forms of PrP appear to have identical covalent structures, but differ in secondary and tertiary structure. Both PrPC and PrPSc have glycosylphospatidylinositol (GPI) anchors through which the protein is tethered to cell membranes.

Matthew R. Hicks; Andrew C. Gill; Imanpreet K. Bath; Atvinder K. Rullay; Ian D. Sylvester; David H. Crout; Teresa J. T. Pinheiro

2006-01-01

221

Distinct prion proteins in short and long scrapie incubation period mice  

Microsoft Academic Search

The Prn-i gene, controlling scrapie incubation period, is linked to or congruent with the murine prion protein (PrP) gene, Prn-p. In prototypic mouse strains with long (l\\/Ln) and short (NZW) incubation periods, Prn-p transcription is initiated at similar multiple sites. The predicted NZW and l\\/Ln PrP proteins differ at codons 108 and 189. Codon 189, highly conserved in mammals, lies

D Westaway; P A Goodman; C A Mirenda; M P McKinley; G A Carlson; S B Prusiner

1987-01-01

222

Implications of prion polymorphisms  

PubMed Central

The sequence of a host’s prion protein (PrP) can affect that host’s susceptibility to prion disease and is the primary basis for the species barrier to transmission. Yet within many species, polymorphisms of the prion protein gene (Prnp) exist, each of which can further affect susceptibility or influence incubation period, pathology and phenotype. As strains are defined by these features (incubation period, pathology, phenotype), polymorphisms may also lead to the preferential propagation or generation of certain strains. In our recent study of the mouse Prnpa and Prnpb polymorphisms (which produced the proteins PrPa and PrPb, respectively), we found differences in aggregation tendency, strain adaptability and conformational variability. Comparing our in vitro data with that of in vivo studies, we found that differing incubation periods between Prnpa and Prnpb mice can primarily be explained on the basis of faster or more efficient aggregation of PrPa. In addition, and more importantly, we found that the faithful propagation of strains in Prnpb mice can be explained by the ability of PrPb to adopt a wider range of conformations. This adaptability allows PrPb to successfully propagate the structural features of a seed. In contrast, Prnpa mice revert PrPb strains into PrPa -type strains, and overall they have a narrower distribution of incubation periods. This can be explained by PrPa having fewer preferred conformations. We propose that Prnp polymorphisms are one route by which certain prion strains may preferentially propagate. This has significant implications for prion disease, chronic wasting disease (CWD) in particular, as it is spreading through North America. Deer which are susceptible to CWD also carry polymorphisms which influence their susceptibility. If these polymorphisms also preferentially allow strain diversification and propagation, this may accelerate the crossing of species barriers and propagation of the disease up the food chain. PMID:23807178

Cortez, Leonardo M; Sim, Valerie L

2013-01-01

223

In vitro prion protein conversion suggests risk of bighorn sheep (Ovis canadensis) to transmissible spongiform encephalopathies  

PubMed Central

Background Transmissible spongiform encephalopathies (TSEs) affect both domestic sheep (scrapie) and captive and free-ranging cervids (chronic wasting disease; CWD). The geographical range of bighorn sheep (Ovis canadensis; BHS) overlaps with states or provinces that have contained scrapie-positive sheep or goats and areas with present epizootics of CWD in cervids. No TSEs have been documented in BHS, but the susceptibility of this species to TSEs remains unknown. Results We acquired a library of BHS tissues and found no evidence of preexisting TSEs in these animals. The prion protein gene (Prnp) in all BHS in our library was identical to scrapie-susceptible domestic sheep (A136R154Q171 genotype). Using an in vitro prion protein conversion assay, which has been previously used to assess TSE species barriers and, in our study appears to recollect known species barriers in mice, we assessed the potential transmissibility of TSEs to BHS. As expected based upon Prnp genotype, we observed BHS prion protein conversion by classical scrapie agent and evidence for a species barrier between transmissible mink encephalopathy (TME) and BHS. Interestingly, our data suggest that the species barrier of BHS to white-tailed deer or wapiti CWD agents is likely low. We also used protein misfolding cyclic amplification to confirm that CWD, but not TME, can template prion protein misfolding in A136R154Q171 genotype sheep. Conclusions Our results indicate the in vitro conversion assay used in our study does mimic the species barrier of mice to the TSE agents that we tested. Based on Prnp genotype and results from conversion assays, BHS are likely to be susceptible to infection by classical scrapie. Despite mismatches in amino acids thought to modulate prion protein conversion, our data indicate that A136R154Q171 genotype sheep prion protein is misfolded by CWD agent, suggesting that these animals could be susceptible to CWD. Further investigation of TSE transmissibility to BHS, including animal studies, is warranted. The lack of reported TSEs in BHS may be attributable to other host factors or a lack of TSE surveillance in this species. PMID:23938169

2013-01-01

224

Flexibility of the murine prion protein and its Asp178Asn mutant investigated by molecular dynamics simulations  

Microsoft Academic Search

Inherited forms of transmissible spongiform encephalopathy, e.g. familial Creutzfeldt–Jakob disease, Gerstmann–Sträussler–Scheinker syndrome and fatal familial insomnia, segregate with specific point mutations of the prion protein. It has been proposed that the pathologically relevant Asp178Asn (D178N) mutation might destabilize the structure of the prion protein because of the loss of the Arg164–Asp178 salt bridge. Molecular dynamics simulations of the structured C-terminal

Jörg Gsponer; Philippe Ferrara; Amedeo Caflisch

2001-01-01

225

Origins and Evolution of the HET-s Prion-Forming Protein: Searching for Other Amyloid-Forming Solenoids  

Microsoft Academic Search

The HET-s prion-forming domain from the filamentous fungus Podospora anserina is gaining considerable interest since it yielded the first well-defined atomic structure of a functional amyloid fibril. This structure has been identified as a left-handed beta solenoid with a triangular hydrophobic core. To delineate the origins of the HET-s prion-forming protein and to discover other amyloid-forming proteins, we searched for

Deena M. A. Gendoo; Paul M. Harrison

2011-01-01

226

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

PubMed

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. PMID:20518029

Moore, Roger A; Timmes, Andrew; Wilmarth, Phillip A; Priola, Suzette A

2010-08-01

227

Unique structural properties associated with mouse prion ?105-125 protein  

PubMed Central

Murine prion protein deleted for residues 105–125 is intrinsically neurotoxic and mediates a TSE-like phenotype in transgenic mice. Equivalent and overlapping deletions were expressed in E.coli, purified and analyzed. Among mutants spanning the region 95–135, a construct lacking solely residues 105–125 had distinct properties when compared with the full-length prion protein 23–231 or other deletions. This distinction was also apparent followed expression in eukaryotic cells. Unlike the full-length protein, all deletion mutants failed to bind to synthetic membranes in vitro. These data suggest a novel structure for the 105–125 deleted variant that may relate to its biological properties. PMID:23764837

Patel, Avnish; Vasiljevic, Snezana; Jones, Ian M.

2013-01-01

228

Prion-like properties of Tau protein: the importance of extracellular Tau as a therapeutic target.  

PubMed

Work over the past 4 years indicates that multiple proteins associated with neurodegenerative diseases, especially Tau and ?-synuclein, can propagate aggregates between cells in a prion-like manner. This means that once an aggregate is formed it can escape the cell of origin, contact a connected cell, enter the cell, and induce further aggregation via templated conformational change. The prion model predicts a key role for extracellular protein aggregates in mediating progression of disease. This suggests new therapeutic approaches based on blocking neuronal uptake of protein aggregates and promoting their clearance. This will likely include therapeutic antibodies or small molecules, both of which can be developed and optimized in vitro prior to preclinical studies. PMID:24860099

Holmes, Brandon B; Diamond, Marc I

2014-07-18

229

Disease-associated Mutations in the Prion Protein Impair Laminin-induced Process Outgrowth and Survival*  

PubMed Central

Prions, the agents of transmissible spongiform encephalopathies, require the expression of prion protein (PrPC) to propagate disease. PrPC is converted into an abnormal insoluble form, PrPSc, that gains neurotoxic activity. Conversely, clinical manifestations of prion disease may occur either before or in the absence of PrPSc deposits, but the loss of normal PrPC function contribution for the etiology of these diseases is still debatable. Prion disease-associated mutations in PrPC represent one of the best models to understand the impact of PrPC loss-of-function. PrPC associates with various molecules and, in particular, the interaction of PrPC with laminin (Ln) modulates neuronal plasticity and memory formation. To assess the functional alterations associated with PrPC mutations, wild-type and mutated PrPC proteins were expressed in a neural cell line derived from a PrPC-null mouse. Treatment with the laminin ?1 chain peptide (Ln ?1), which mimics the Ln binding site for PrPC, increased intracellular calcium in cells expressing wild-type PrPC, whereas a significantly lower response was observed in cells expressing mutated PrPC molecules. The Ln ?1 did not promote process outgrowth or protect against staurosporine-induced cell death in cells expressing mutated PrPC molecules in contrast to cells expressing wild-type PrPC. The co-expression of wild-type PrPC with mutated PrPC molecules was able to rescue the Ln protective effects, indicating the lack of negative dominance of PrPC mutated molecules. These results indicate that PrPC mutations impair process outgrowth and survival mediated by Ln ?1 peptide in neural cells, which may contribute to the pathogenesis of genetic prion diseases. PMID:23132868

Machado, Cleiton F.; Beraldo, Flavio H.; Santos, Tiago G.; Bourgeon, Dominique; Landemberger, Michele C.; Roffe, Martin; Martins, Vilma R.

2012-01-01

230

A survey and a molecular dynamics study on the (central) hydrophobic region of prion proteins  

E-print Network

Prion diseases are invariably fatal neurodegenerative diseases that affect humans and animals. Unlike most other amyloid forming neurodegenerative diseases, these can be highly infectious. Prion diseases occur in a variety of species. They include the fatal human neurodegenerative diseases Creutzfeldt-Jakob Disease (CJD), Fatal Familial Insomnia (FFI), Gerstmann-Straussler-Scheinker syndrome (GSS), Kuru, the bovine spongiform encephalopathy (BSE or 'mad-cow' disease) in cattle, the chronic wasting disease (CWD) in deer and elk, and scrapie in sheep and goats, etc. Transmission across the species barrier to humans, especially in the case of BSE in Europe, CWD in North America, and variant CJDs (vCJDs) in young people of UK, is a major public health concern. Fortunately, scientists reported that the (central) hydrophobic region of prion proteins (PrP) controls the formation of diseased prions. This article gives a detailed survey on PrP hydrophobic region and does molecular dynamics studies of human PrP(110-136...

Zhang, Jiapu

2014-01-01

231

Reduced Translocation of Nascent Prion Protein During ER Stress Contributes to Neurodegeneration  

PubMed Central

Summary During acute stress in the endoplasmic reticulum (ER), mammalian prion protein (PrP) is temporarily prevented from translocation into the ER and instead routed directly for cytosolic degradation. This ‘pre-emptive’ quality control (pQC) system benefits cells by minimizing PrP aggregation in the secretory pathway during ER stress. However, the potential toxicity of cytosolic PrP raised the possibility that persistent pQC of PrP contributes to neurodegeneration in prion diseases. Here, we find evidence of ER stress and decreased translocation of nascent PrP during prion infection. Transgenic mice expressing a PrP variant with reduced translocation at levels expected during ER stress was sufficient to cause several mild age-dependent clinical and histological manifestations of PrP-mediated neurodegeneration. Thus, an ordinarily adaptive quality control pathway can be contextually detrimental over long time periods. We propose that one mechanism of prion-mediated neurodegeneration involves an indirect ER stress-dependent effect on nascent PrP biosynthesis and metabolism. PMID:18804434

Rane, Neena S.; Kang, Sang-Wook; Chakrabarti, Oishee; Feigenbaum, Lionel; Hegde, Ramanujan S.

2008-01-01

232

An N-terminal Fragment of the Prion Protein Binds to Amyloid-? Oligomers and Inhibits Their Neurotoxicity in Vivo*  

PubMed Central

A hallmark of Alzheimer disease (AD) is the accumulation of the amyloid-? (A?) peptide in the brain. Considerable evidence suggests that soluble A? oligomers are responsible for the synaptic dysfunction and cognitive deficit observed in AD. However, the mechanism by which these oligomers exert their neurotoxic effect remains unknown. Recently, it was reported that A? oligomers bind to the cellular prion protein with high affinity. Here, we show that N1, the main physiological cleavage fragment of the cellular prion protein, is necessary and sufficient for binding early oligomeric intermediates during A? polymerization into amyloid fibrils. The ability of N1 to bind A? oligomers is influenced by positively charged residues in two sites (positions 23–31 and 95–105) and is dependent on the length of the sequence between them. Importantly, we also show that N1 strongly suppresses A? oligomer toxicity in cultured murine hippocampal neurons, in a Caenorhabditis elegans-based assay, and in vivo in a mouse model of A?-induced memory dysfunction. These data suggest that N1, or small peptides derived from it, could be potent inhibitors of A? oligomer toxicity and represent an entirely new class of therapeutic agents for AD. PMID:23362282

Fluharty, Brian R.; Biasini, Emiliano; Stravalaci, Matteo; Sclip, Alessandra; Diomede, Luisa; Balducci, Claudia; La Vitola, Pietro; Messa, Massimo; Colombo, Laura; Forloni, Gianluigi; Borsello, Tiziana; Gobbi, Marco; Harris, David A.

2013-01-01

233

Chemical Induction of Misfolded Prion Protein Conformers in Cell Culture*  

PubMed Central

Prion-infected cells accumulate a heterogeneous population of aberrantly folded PrP conformers, including the disease-causing isoform (PrPSc). We found that specific chemicals can modulate the levels of various PrP conformers in cultured cells. Positively charged polyamidoamines (dendrimers) eliminated protease-resistant (r) PrPSc from prion-infected cells and induced the formation of insoluble, protease-sensitive PrP aggregates (designated PrPA). Larger, positively charged polyamidoamines more efficaciously induced the formation of PrPA and cleared rPrPSc, whereas negatively charged polyamidoamines neither induced PrPA nor cleared rPrPSc. Although the biochemical properties of PrPA were shown to be similar to protease-sensitive (s) PrPSc, bioassays of PrPA indicated that it is not infectious. Our studies argue that PrPA represents an aggregated PrP species that is off-pathway relative to the formation of rPrPSc. It remains to be established whether the formation of PrPA inhibits the formation of rPrPSc by sequestering PrPC in the form of benign, insoluble aggregates. PMID:19955177

Ghaemmaghami, Sina; Ullman, Julie; Ahn, Misol; St. Martin, Susan; Prusiner, Stanley B.

2010-01-01

234

Protein Sequence Databases  

Microsoft Academic Search

\\u000a With the availability of almost 150 completed genome sequences from both eukaryotic and prokaryotic organisms, efforts are\\u000a now being focused on the identification and functional analysis of the proteins encoded by these genomes. The rapidly emerging\\u000a field of proteomics, the large-scale analysis of these proteins, has started to generate huge amounts of data as a result\\u000a of the new information

Michele Magrane; Maria Jesus Martin; Claire O’Donovan; Rolf Apweiler

235

Inactivation of template-directed misfolding of infectious prion protein by ozone.  

PubMed

Misfolded prions (PrP(Sc)) are well known for their resistance to conventional decontamination processes. The potential risk of contamination of the water environment, as a result of disposal of specified risk materials (SRM), has raised public concerns. Ozone is commonly utilized in the water industry for inactivation of microbial contaminants and was tested in this study for its ability to inactivate prions (263K hamster scrapie = PrP(Sc)). Treatment variables included initial ozone dose (7.6 to 25.7 mg/liter), contact time (5 s and 5 min), temperature (4°C and 20°C), and pH (pH 4.4, 6.0, and 8.0). Exposure of dilute suspensions of the infected 263K hamster brain homogenates (IBH) (0.01%) to ozone resulted in the in vitro destruction of the templating properties of PrP(Sc), as measured by the protein misfolding cyclic amplification (PMCA) assay. The highest levels of prion inactivation (?4 log(10)) were observed with ozone doses of 13.0 mg/liter, at pH 4.4 and 20°C, resulting in a CT (the product of residual ozone concentration and contact time) value as low as 0.59 mg · liter(-1) min. A comparison of ozone CT requirements among various pathogens suggests that prions are more susceptible to ozone degradation than some model bacteria and protozoa and that ozone treatment may be an effective solution for inactivating prions in water and wastewater. PMID:22138993

Ding, Ning; Neumann, Norman F; Price, Luke M; Braithwaite, Shannon L; Balachandran, Aru; Belosevic, Miodrag; El-Din, Mohamed Gamal

2012-02-01

236

?-Helical to ?-Helical Conformation Change in the C-Terminal of the Mammalian Prion Protein  

NASA Astrophysics Data System (ADS)

We employ all-atom structure-based models with mixed basis contact maps to explore whether there are any significant geometric or energetic constraints limiting conjectured conformational transitions between the alpha-helical (?H) and the left handed beta helical (LHBH) conformations for the C-terminal (residues 166-226) of the mammalian prion protein. The LHBH structure has been proposed to describe infectious oligomers and one class of in vitro grown fibrils, as well as possibly self- templating the conversion of normal cellular prion protein to the infectious form. Our results confirm that the kinetics of the conformation change are not strongely limited by large scale geometry modification and there exists an overall preference for the LHBH conformation.

Singh, Jesse; Whitford, Paul; Hayre, Natha; Cox, Daniel; Onuchic, José.

2011-03-01

237

Prion induction involves an ancient system for the sequestration of aggregated proteins and heritable changes in prion fragmentation  

E-print Network

When the translation termination factor Sup35 adopts the prion state, [PSI+], the read-through of stop codons increases, uncovering hidden genetic variation and giving rise to new, often beneficial, phenotypes. Evidence ...

Tyedmers, Jens

238

Novel N-terminal domain mutation in prion protein detected in 2 patients diagnosed with frontotemporal lobar degeneration syndrome.  

PubMed

Prion protein gene mutations have been associated with clinical pictures mimicking neurodegenerative diseases different from inherited prion diseases (IPD). We report a novel missense P39L mutation in the N-terminal domain of prion protein in 2 patients affected by frontotemporal lobar degeneration syndrome, negative for mutations in genes causative of dementia. Neither the first carrier, a 67-year-old male in which the onset was a progressive non-fluent aphasia, nor the second carrier, a 78-year-old male affected by frontotemporal dementia and parkinsonism, showed any clinical or instrumental findings suggestive of IPD. Genetic screening of healthy controls and in silico analysis provide support for the potential pathogenicity of this variant. Patient phenotypes, unclassifiable as prion disease, may depend on the location of the mutation in the N-terminal domain, outside the amyloid core of pathologic prion protein, although further functional studies are required to determine whether and how this mutation exerts its pathogenic effect. However, genetic screening of prion protein gene becomes relevant in familial degenerative dementia, particularly in geographical areas with high IPD prevalence. PMID:25022973

Bernardi, Livia; Cupidi, Chiara; Frangipane, Francesca; Anfossi, Maria; Gallo, Maura; Conidi, Maria Elena; Vasso, Franca; Colao, Rosanna; Puccio, Gianfranco; Curcio, Sabrina A M; Mirabelli, Maria; Clodomiro, Alessandra; Di Lorenzo, Raffaele; Smirne, Nicoletta; Maletta, Raffaele; Bruni, Amalia C

2014-11-01

239

Subcellular Colocalization of the Cellular and Scrapie Prion Proteins in Caveolae-Like Membranous Domains  

Microsoft Academic Search

Results of transgenetic studies argue that the scrapie isoform of the prion protein (PrPSc) interacts with the substrate cellular PrP (PrPC) during conversion into nascent PrPSc. While PrPSc appears to accumulate primarily in lysosomes, caveolae-like domains (CLDs) have been suggested to be the site where PrPC is converted into PrPSc. We report herein that CLDs isolated from scrapie-infected neuroblastoma (ScN2a)

Martin Vey; Susanne Pilkuhn; Holger Wille; Randal Nixon; Stephen J. Dearmond; Eric J. Smart; Richard G. W. Anderson; Albert Taraboulos

1996-01-01

240

Contributions of neuronal prion protein on sleep recovery and stress response following sleep deprivation  

Microsoft Academic Search

In order to gain insights on the function of the cellular prion protein (PrPC) sleep and the levels of the stress hormones corticosterone (CORT) and the adrenocorticotropic hormone (ACTH) before and after sleep deprivation (SD) were compared in two wild type (WT) mice strains and the following three PrPC transgenic lines: mice null for PrPC (mPrP0\\/0) and mice with specific

Manuel Sánchez-Alavez; Bruno Conti; Gianluca Moroncini; José R. Criado

2007-01-01

241

Scrapie-associated Prion Protein in the Gastro-intestinal Tract of Sheep with Natural Scrapie  

Microsoft Academic Search

The scrapie-associated prion protein (PrPSc), which is closely associated with scrapie infectivity, accumulates in the brain and lymphoid tissues of sheep with natural scrapie. The most probable portal of entry of the scrapie agent in sheep is the alimentary tract; little attention, however, has been paid to the gastro-intestinal tract in scrapie research. In this study, we examined the presence

L. J. M. van Keulen; B. E. C. Schreuder; M. E. W. Vromans; J. P. M. Langeveld; M. A. Smits

1999-01-01

242

Genetic variability of the coding region for the prion protein gene ( PRNP ) in gayal ( Bos frontalis )  

Microsoft Academic Search

The gayal (Bos frontalis) is a rare semi-wild bovid species in which bovine spongiform encephalopathy (BSE) has not been reported. Polymorphisms of\\u000a the prion protein gene (PRNP) have been correlated significantly with resistance to BSE. In this study, the coding region of PRNP was cloned and characterized in samples from 125 gayal. A total of ten single nucleotide polymorphisms (SNPs),

Dongmei XiQing; Qing Liu; Jianhong Guo; Hongman Yu; Yuai Yang; Yiduo He; Huaming Mao; Xiao Gou; Weidong Deng

243

Nonenzymatic glycation at the N terminus of pathogenic prion protein in transmissible spongiform encephalopathies.  

PubMed

Transmissible spongiform encephalopathies (TSEs) are transmissible neurodegenerative diseases characterized by the accumulation of an abnormally folded prion protein, termed PrPSc, and the development of pathological features of astrogliosis, vacuolation, neuronal cell loss, and in some cases amyloid plaques. Although considerable structural characterization of prion protein has been reported, neither the method of conversion of cellular prion protein, PrPC, into the pathogenic isoform nor the post-translational modification processes involved is known. We report that in animal and human TSEs, one or more lysines at residues 23, 24, and 27 of PrPSc are covalently modified with advanced glycosylation end products (AGEs), which may be carboxymethyl-lysine (CML), one of the structural varieties of AGEs. The arginine residue at position 37 may also be modified with AGE, but not the arginine residue at position 25. This result suggests that nonenzymatic glycation is one of the post-translational modifications of PrP(Sc). Furthermore, immunostaining studies indicate that, at least in clinically affected hamsters, astrocytes are the first site of this glycation process. PMID:15084583

Choi, Yeong-Gon; Kim, Jae-Il; Jeon, Yong-Chul; Park, Seok-Joo; Choi, Eun-Kyoung; Rubenstein, Richard; Kascsak, Richard J; Carp, Richard I; Kim, Yong-Sun

2004-07-16

244

Absolute Quantification of Prion Protein (90-231) Using Stable Isotope-Labeled Chymotryptic Peptide Standards in a LC-MRM AQUA Workflow  

NASA Astrophysics Data System (ADS)

Substantial evidence indicates that the disease-associated conformer of the prion protein (PrPTSE) constitutes the etiologic agent in prion diseases. These diseases affect multiple mammalian species. PrPTSE has the ability to convert the conformation of the normal prion protein (PrPC) into a ?-sheet rich form resistant to proteinase K digestion. Common immunological techniques lack the sensitivity to detect PrPTSE at subfemtomole levels, whereas animal bioassays, cell culture, and in vitro conversion assays offer higher sensitivity but lack the high-throughput the immunological assays offer. Mass spectrometry is an attractive alternative to the above assays as it offers high-throughput, direct measurement of a protein's signature peptide, often with subfemtomole sensitivities. Although a liquid chromatography-multiple reaction monitoring (LC-MRM) method has been reported for PrPTSE, the chemical composition and lack of amino acid sequence conservation of the signature peptide may compromise its accuracy and make it difficult to apply to multiple species. Here, we demonstrate that an alternative protease (chymotrypsin) can produce signature peptides suitable for a LC-MRM absolute quantification (AQUA) experiment. The new method offers several advantages, including: (1) a chymotryptic signature peptide lacking chemically active residues (Cys, Met) that can confound assay accuracy; (2) low attomole limits of detection and quantitation (LOD and LOQ); and (3) a signature peptide retaining the same amino acid sequence across most mammals naturally susceptible to prion infection as well as important laboratory models. To the authors' knowledge, this is the first report on the use of a non-tryptic peptide in a LC-MRM AQUA workflow.

Sturm, Robert; Sheynkman, Gloria; Booth, Clarissa; Smith, Lloyd M.; Pedersen, Joel A.; Li, Lingjun

2012-09-01

245

Prion formation by a yeast GLFG nucleoporin  

PubMed Central

The self-assembly of proteins into higher order structures is both central to normal biology and a dominant force in disease. Certain glutamine/asparagine (Q/N)-rich proteins in the budding yeast Saccharomyces cerevisiae assemble into self-replicating amyloid-like protein polymers, or prions, that act as genetic elements in an entirely protein-based system of inheritance. The nuclear pore complex (NPC) contains multiple Q/N-rich proteins whose self-assembly has also been proposed to underlie structural and functional properties of the NPC. Here we show that an essential sequence feature of these proteins—repeating GLFG motifs—strongly promotes their self-assembly into amyloids with characteristics of prions. Furthermore, we demonstrate that Nup100 can form bona fide prions, thus establishing a previously undiscovered ability of yeast GLFG nucleoporins to adopt this conformational state in vivo. PMID:22561191

Halfmann, Randal; Wright, Jessica R.; Alberti, Simon; Lindquist, Susan; Rexach, Michael

2012-01-01

246

A new paradigm for enzymatic control of ?-cleavage and ?-cleavage of the prion protein.  

PubMed

The cellular form of the prion protein (PrP(C)) is found in both full-length and several different cleaved forms in vivo. Although the precise functions of the PrP(C) proteolytic products are not known, cleavage between the unstructured N-terminal domain and the structured C-terminal domain at Lys-109?His-110 (mouse sequence), termed ?-cleavage, has been shown to produce the anti-apoptotic N1 and the scrapie-resistant C1 peptide fragments. ?-Cleavage, residing adjacent to the octarepeat domain and N-terminal to the ?-cleavage site, is thought to arise from the action of reactive oxygen species produced from redox cycling of coordinated copper. We sought to elucidate the role of key members of the ADAM (a disintegrin and metalloproteinase) enzyme family, as well as Cu(2+) redox cycling, in recombinant mouse PrP (MoPrP) cleavage through LC/MS analysis. Our findings show that although Cu(2+) redox-generated reactive oxygen species do produce fragmentation corresponding to ?-cleavage, ADAM8 also cleaves MoPrP in the octarepeat domain in a Cu(2+)- and Zn(2+)-dependent manner. Additional cleavage by ADAM8 was observed at the previously proposed location of ?-cleavage, Lys-109?His-110 (MoPrP sequencing); however, upon addition of Cu(2+), the location of ?-cleavage shifted by several amino acids toward the C terminus. ADAM10 and ADAM17 have also been implicated in ?-cleavage at Lys-109?His-110; however, we observed that they instead cleaved MoPrP at a novel location, Ala-119?Val-120, with additional cleavage by ADAM10 at Gly-227?Arg-228 near the C terminus. Together, our results show that MoPrP cleavage is far more complex than previously thought and suggest a mechanism by which PrP(C) fragmentation responds to Cu(2+) and Zn(2+). PMID:24247244

McDonald, Alex J; Dibble, Jessie P; Evans, Eric G B; Millhauser, Glenn L

2014-01-10

247

Functional role of Tia1/Pub1 and Sup35 prion domains: directing protein synthesis machinery to the tubulin cytoskeleton.  

PubMed

Tia1/Pub1 is a stress granule component carrying a Q/N-rich prion domain. We provide direct evidence that Tia1 forms a prion in yeast. Moreover, Tia1/Pub1 acts cooperatively with release factor Sup35/eRF3 to establish a two-protein self-propagating state. This two-protein prion driven by the Q/N-rich prion domains of Sup35 and Tia1/Pub1 can be visualized as distinctive line structures along tubulin cytoskeleton. Furthermore, we find that tubulin-associated complex containing Pub1 and Sup35 oligomers normally exists in yeast, and its assembly depends on prion domains of Pub1 and Sup35. This Sup35/Pub1 complex, which also contains TUB1 mRNA and components of translation machinery, is important for the integrity of the tubulin cytoskeleton: PUB1 disruption and Sup35 depletion from the complex lead to cytoskeletal defects. We propose that the complex is implicated in protein synthesis at the site of microtubule assembly. Thus our study identifies the role for prion domains in the assembly of multiprotein complexes. PMID:24981173

Li, Xiang; Rayman, Joseph B; Kandel, Eric R; Derkatch, Irina L

2014-07-17

248

Evolution of vertebrate genes related to prion and Shadoo proteins--clues from comparative genomic analysis.  

PubMed

Recent findings of new genes in fish related to the prion protein (PrP) gene PRNP, including our recent report of SPRN coding for Shadoo (Sho) protein found also in mammals, raise issues of their function and evolution. Here we report additional novel fish genes found in public databases, including a duplicated SPRN gene, SPRNB, in Fugu, Tetraodon, carp, and zebrafish encoding the Sho2 protein, and we use comparative genomic analysis to analyze the evolutionary relationships and to infer evolutionary trajectories of the complete data set. Phylogenetic footprinting performed on aligned human, mouse, and Fugu SPRN genes to define candidate regulatory promoter regions, detected 16 conserved motifs, three of which are known transcription factor-binding sites for a receptor and transcription factors specific to or associated with expression in brain. This result and other homology-based (VISTA global genomic alignment; protein sequence alignment and phylogenetics) and context-dependent (genomic context; relative gene order and orientation) criteria indicate fish and mammalian SPRN genes are orthologous and suggest a strongly conserved basic function in brain. Whereas tetrapod PRNPs share context with the analogous stPrP-2-coding gene in fish, their sequences are diverged, suggesting that the tetrapod and fish genes are likely to have significantly different functions. Phylogenetic analysis predicts the SPRN/SPRNB duplication occurred before divergence of fish from tetrapods, whereas that of stPrP-1 and stPrP-2 occurred in fish. Whereas Sho appears to have a conserved function in vertebrate brain, PrP seems to have an adaptive role fine-tuned in a lineage-specific fashion. An evolutionary model consistent with our findings and literature knowledge is proposed that has an ancestral prevertebrate SPRN-like gene leading to all vertebrate PrP-related and Sho-related genes. This provides a new framework for exploring the evolution of this unusual family of proteins and for searching for members in other fish branches and intermediate vertebrate groups. PMID:15342797

Premzl, Marko; Gready, Jill E; Jermiin, Lars S; Simonic, Tatjana; Marshall Graves, Jennifer A

2004-12-01

249

Comparative genomic analysis of prion genes  

PubMed Central

Background The homologues of human disease genes are expected to contribute to better understanding of physiological and pathogenic processes. We made use of the present availability of vertebrate genomic sequences, and we have conducted the most comprehensive comparative genomic analysis of the prion protein gene PRNP and its homologues, shadow of prion protein gene SPRN and doppel gene PRND, and prion testis-specific gene PRNT so far. Results While the SPRN and PRNP homologues are present in all vertebrates, PRND is known in tetrapods, and PRNT is present in primates. PRNT could be viewed as a TE-associated gene. Using human as the base sequence for genomic sequence comparisons (VISTA), we annotated numerous potential cis-elements. The conserved regions in SPRNs harbour the potential Sp1 sites in promoters (mammals, birds), C-rich intron splicing enhancers and PTB intron splicing silencers in introns (mammals, birds), and hsa-miR-34a sites in 3'-UTRs (eutherians). We showed the conserved PRNP upstream regions, which may be potential enhancers or silencers (primates, dog). In the PRNP 3'-UTRs, there are conserved cytoplasmic polyadenylation element sites (mammals, birds). The PRND core promoters include highly conserved CCAAT, CArG and TATA boxes (mammals). We deduced 42 new protein primary structures, and performed the first phylogenetic analysis of all vertebrate prion genes. Using the protein alignment which included 122 sequences, we constructed the neighbour-joining tree which showed four major clusters, including shadoos, shadoo2s and prion protein-likes (cluster 1), fish prion proteins (cluster 2), tetrapode prion proteins (cluster 3) and doppels (cluster 4). We showed that the entire prion protein conformationally plastic region is well conserved between eutherian prion proteins and shadoos (18–25% identity and 28–34% similarity), and there could be a potential structural compatibility between shadoos and the left-handed parallel beta-helical fold. Conclusion It is likely that the conserved genomic elements identified in this analysis represent bona fide cis-elements. However, this idea needs to be confirmed by functional assays in transgenic systems. PMID:17199895

Premzl, Marko; Gamulin, Vera

2007-01-01

250

Integrity of helix 2-helix 3 domain of the PrP protein is not mandatory for prion replication.  

PubMed

The process of prion conversion is not yet well understood at the molecular level. The regions critical for the conformational change of PrP remain mostly debated and the extent of sequence change acceptable for prion conversion is poorly documented. To achieve progress on these issues, we applied a reverse genetic approach using the Rov cell system. This allowed us to test the susceptibility of a number of insertion mutants to conversion into prion in the absence of wild-type PrP molecules. We were able to propagate several prions with 8 to 16 extra amino acids, including a polyglycine stretch and His or FLAG tags, inserted in the middle of the protease-resistant fragment. These results demonstrate the possibility to increase the length of the loop between helices H2 and H3 up to 4-fold, without preventing prion replication. They also indicate that this loop probably remains unstructured in PrP(Sc). We also showed that bona fide prions can be produced following insertion of octapeptides in the two C-terminal turns of H2. These insertions do not interfere with the overall fold of the H2-H3 domain indicating that the highly conserved sequence of the terminal part of H2 is not critical for the conversion. Altogether these data showed that the amplitude of modifications acceptable for prion conversion in the core of the globular domain of PrP is much greater than one might have assumed. These observations should help to refine structural models of PrP(Sc) and elucidate the conformational changes underlying prions generation. PMID:22511770

Salamat, Khalid; Moudjou, Mohammed; Chapuis, Jérôme; Herzog, Laetitia; Jaumain, Emilie; Béringue, Vincent; Rezaei, Human; Pastore, Annalisa; Laude, Hubert; Dron, Michel

2012-06-01

251

Integrity of Helix 2-Helix 3 Domain of the PrP Protein Is Not Mandatory for Prion Replication*  

PubMed Central

The process of prion conversion is not yet well understood at the molecular level. The regions critical for the conformational change of PrP remain mostly debated and the extent of sequence change acceptable for prion conversion is poorly documented. To achieve progress on these issues, we applied a reverse genetic approach using the Rov cell system. This allowed us to test the susceptibility of a number of insertion mutants to conversion into prion in the absence of wild-type PrP molecules. We were able to propagate several prions with 8 to 16 extra amino acids, including a polyglycine stretch and His or FLAG tags, inserted in the middle of the protease-resistant fragment. These results demonstrate the possibility to increase the length of the loop between helices H2 and H3 up to 4-fold, without preventing prion replication. They also indicate that this loop probably remains unstructured in PrPSc. We also showed that bona fide prions can be produced following insertion of octapeptides in the two C-terminal turns of H2. These insertions do not interfere with the overall fold of the H2-H3 domain indicating that the highly conserved sequence of the terminal part of H2 is not critical for the conversion. Altogether these data showed that the amplitude of modifications acceptable for prion conversion in the core of the globular domain of PrP is much greater than one might have assumed. These observations should help to refine structural models of PrPSc and elucidate the conformational changes underlying prions generation. PMID:22511770

Salamat, Khalid; Moudjou, Mohammed; Chapuis, Jerome; Herzog, Laetitia; Jaumain, Emilie; Beringue, Vincent; Rezaei, Human; Pastore, Annalisa; Laude, Hubert; Dron, Michel

2012-01-01

252

Consequences of manganese replacement of copper for prion protein function and proteinase resistance  

PubMed Central

The prion protein (PrP) binds copper and has antioxidant activity enhancing the survival of neurones in culture. The ability of the PrP to bind other cations was tested and it was found that only manganese could substitute for copper. Although initially manganese-loaded PrP exhibited similar structure and activity to copper-loaded PrP, after aging, manganese-loaded PrP became proteinase resistant and lost function. It was also found that manganese could be incorporated into PrP expressed by astrocytes and that this PrP was partially proteinase resistant. These results show that it is possible to generate proteinase-resistant PrP from cells and suggest a possible mechanism for the formation of the scrapie isoform of the PrP as generated in sporadic prion disease. PMID:10716918

Brown, David R.; Hafiz, Farida; Glasssmith, Leslie L.; Wong, Boon-Seng; Jones, Ian M.; Clive, Christine; Haswell, Stephen J.

2000-01-01

253

Failure of Prion Protein Oxidative Folding Guides the Formation of Toxic Transmembrane Forms*  

PubMed Central

The mechanism by which pathogenic mutations in the globular domain of the cellular prion protein (PrPC) increase the likelihood of misfolding and predispose to diseases is not yet known. Differences in the evidences provided by structural and metabolic studies of these mutants suggest that in vivo folding could be playing an essential role in their pathogenesis. To address this role, here we use the single or combined M206S and M213S artificial mutants causing labile folds and express them in cells. We find that these mutants are highly toxic, fold as transmembrane PrP, and lack the intramolecular disulfide bond. When the mutations are placed in a chain with impeded transmembrane PrP formation, toxicity is rescued. These results suggest that oxidative folding impairment, as on aging, can be fundamental for the genesis of intracellular neurotoxic intermediates key in prion neurodegenerations. PMID:22955286

Lisa, Silvia; Domingo, Beatriz; Martinez, Javier; Gilch, Sabine; Llopis, Juan F.; Schatzl, Hermann M.; Gasset, Maria

2012-01-01

254

Electrochemical aptasensor of cellular prion protein based on modified polypyrrole with redox dendrimers.  

PubMed

This work consists of the development of an electrochemical aptasensor based on polyprrole modified with redox dendrimers, able to detect human cellular prions PrP(C) with high sensitivity. The gold surface was modified by conductive polypyrrole film coupled to polyamidoamine dendrimers of fourth generation (PAMAM G4) and ferrocenyl group as redox marker. The aptamers were immobilized on the surface via biotin/streptavidin chemistry. Electrochemical signal was detected by ferrocenyl group incorporated between dendrimers and aptamers layers. We demonstrated that the interaction between aptamer and prion protein led to variation in electrochemical signal of the ferrocenyl group. The kinetics parameters (diffusion coefficient D and heterogeneous constant transfer ket) calculated from electrochemical signals demonstrate that the variation in redox signal results from the lower diffusion process of ions during redox reaction after prion interaction due to bulk effect of larger protein. The association of redox dendrimers with conducting polypyrrole leads to high sensitivity of PrP(C) determination with detection limit of 0.8 pM, which is three orders of magnitude lower, compared to flat ferrocene-functionalized polypyrrole. Detection of PrP(C) in spiked blood plasma has been achieved and demonstrated a recovery up to 90%. PMID:24480126

Miodek, A; Castillo, G; Hianik, T; Korri-Youssoufi, H

2014-06-15

255

Mapping the Prion Protein Distribution in Marsupials: Insights from Comparing Opossum with Mouse CNS  

PubMed Central

The cellular form of the prion protein (PrPC) is a sialoglycoprotein widely expressed in the central nervous system (CNS) of mammalian species during neurodevelopment and in adulthood. The location of the protein in the CNS may play a role in the susceptibility of a species to fatal prion diseases, which are also known as the transmissible spongiform encephalopathies (TSEs). To date, little is known about PrPC distribution in marsupial mammals, for which no naturally occurring prion diseases have been reported. To extend our understanding of varying PrPC expression profiles in different mammals we carried out a detailed expression analysis of PrPC distribution along the neurodevelopment of the metatherian South American short-tailed opossum (Monodelphis domestica). We detected lower levels of PrPC in white matter fiber bundles of opossum CNS compared to mouse CNS. This result is consistent with a possible role for PrPC in the distinct neurodevelopment and neurocircuitry found in marsupials compared to other mammalian species. PMID:23209725

Poggiolini, Ilaria; Legname, Giuseppe

2012-01-01

256

A simple quantitative model of macromolecular crowding effects on protein folding: Application to the murine prion protein(121-231)  

NASA Astrophysics Data System (ADS)

A model of protein folding kinetics is applied to study the effects of macromolecular crowding on protein folding rate and stability. Macromolecular crowding is found to promote a decrease of the entropic cost of folding of proteins that produces an increase of both the stability and the folding rate. The acceleration of the folding rate due to macromolecular crowding is shown to be a topology-dependent effect. The model is applied to the folding dynamics of the murine prion protein (121-231). The differential effect of macromolecular crowding as a function of protein topology suffices to make non-native configurations relatively more accessible.

Bergasa-Caceres, Fernando; Rabitz, Herschel A.

2013-06-01

257

Motomasa Tanaka Recent progress in prion biology  

E-print Network

Motomasa Tanaka RIKEN BSI Recent progress in prion biology Abstract Proteins often misfold is the prion strain phenomenon, in which prion particles apparently composed of the same protein lead to phenotypically distinct heritable states. In this lecture, I will review recent progress in prion biology

Fukai, Tomoki

258

Mouse-Hamster Chimeric Prion Protein (PrP) Devoid of N-Terminal Residues 23-88 Restores Susceptibility to 22L Prions, but Not to RML Prions in PrP-Knockout Mice  

PubMed Central

Prion infection induces conformational conversion of the normal prion protein PrPC, into the pathogenic isoform PrPSc, in prion diseases. It has been shown that PrP-knockout (Prnp0/0) mice transgenically reconstituted with a mouse-hamster chimeric PrP lacking N-terminal residues 23-88, or Tg(MHM2?23-88)/Prnp0/0 mice, neither developed the disease nor accumulated MHM2Sc?23-88 in their brains after inoculation with RML prions. In contrast, RML-inoculated Tg(MHM2?23-88)/Prnp0/+ mice developed the disease with abundant accumulation of MHM2Sc?23-88 in their brains. These results indicate that MHM2?23-88 itself might either lose or greatly reduce the converting capacity to MHM2Sc?23-88, and that the co-expressing wild-type PrPC can stimulate the conversion of MHM2?23-88 to MHM2Sc?23-88 in trans. In the present study, we confirmed that Tg(MHM2?23-88)/Prnp0/0 mice remained resistant to RML prions for up to 730 days after inoculation. However, we found that Tg(MHM2?23-88)/Prnp0/0 mice were susceptible to 22L prions, developing the disease with prolonged incubation times and accumulating MHM2Sc?23-88 in their brains. We also found accelerated conversion of MHM2?23-88 into MHM2Sc?23-88 in the brains of RML- and 22L-inoculated Tg(MHM2?23-88)/Prnp0/+ mice. However, wild-type PrPSc accumulated less in the brains of these inoculated Tg(MHM2?23-88)/Prnp0/+ mice, compared with RML- and 22L-inoculated Prnp0/+ mice. These results show that MHM2?23-88 itself can convert into MHM2Sc?23-88 without the help of the trans-acting PrPC, and that, irrespective of prion strains inoculated, the co-expressing wild-type PrPC stimulates the conversion of MHM2?23-88 into MHM2Sc?23-88, but to the contrary, the co-expressing MHM2?23-88 disturbs the conversion of wild-type PrPC into PrPSc. PMID:25330286

Yano, Masashi; Yamaguchi, Yoshitaka; Imamura, Morikazu; Muramatsu, Naomi; Das, Nandita Rani; Chida, Junji; Hara, Hideyuki; Sakaguchi, Suehiro

2014-01-01

259

Spontaneous Generation of Infectious Prion Disease in Transgenic Mice  

PubMed Central

We generated transgenic mice expressing bovine cellular prion protein (PrPC) with a leucine substitution at codon 113 (113L). This protein is homologous to human protein with mutation 102L, and its genetic link with Gerstmann–Sträussler–Scheinker syndrome has been established. This mutation in bovine PrPC causes a fully penetrant, lethal, spongiform encephalopathy. This genetic disease was transmitted by intracerebral inoculation of brain homogenate from ill mice expressing mutant bovine PrP to mice expressing wild-type bovine PrP, which indicated de novo generation of infectious prions. Our findings demonstrate that a single amino acid change in the PrPC sequence can induce spontaneous generation of an infectious prion disease that differs from all others identified in hosts expressing the same PrPC sequence. These observations support the view that a variety of infectious prion strains might spontaneously emerge in hosts displaying random genetic PrPC mutations. PMID:24274622

Castilla, Joaquin; Pintado, Belen; Gutierrez-Adan, Alfonso; Andreoletti, Olivier; Aguilar-Calvo, Patricia; Arroba, Ana-Isabel; Parra-Arrondo, Beatriz; Ferrer, Isidro; Manzanares, Jorge; Espinosa, Juan-Carlos

2013-01-01

260

Stability of the ?-structure in prion protein: A molecular dynamics study based on polarized force field  

NASA Astrophysics Data System (ADS)

Conformational changes of the antiparallel ?-sheet in normal cellular prion protein (PrPC) of rat, bovine, and human are investigated by molecular dynamics simulations in both neutral and acidic environment. Using a recently developed simulation method based on an on-the-fly polarized protein-specific charge (PPC) update scheme during the simulation process, we evaluate and compare the cross-species performances of the ?-sheet during the early stage transition from the PrPC to its mutant configuration. Through this study, we observe the growth of the ?-sheet structure in all species studied with the extent of elongation in ?-sheet being different across the three species.

Xu, Zhijun; Lazim, Raudah; Mei, Ye; Zhang, Dawei

2012-06-01

261

Two Misfolding Routes for the Prion Protein around pH 4.5  

PubMed Central

Using molecular dynamics simulations, we show that the prion protein (PrP) exhibits a dual behavior, with two possible transition routes, upon protonation of H187 around pH 4.5, which mimics specific conditions encountered in endosomes. Our results suggest a picture in which the protonated imidazole ring of H187 experiences an electrostatic repulsion with the nearby guanidinium group of R136, to which the system responds by pushing either H187 or R136 sidechains away from their native cavities. The regions to which H187 and R136 are linked, namely the C-terminal part of H2 and the loop connecting S1 to H1, respectively, are affected in a different manner depending on which pathway is taken. Specific in vivo or in vitro conditions, such as the presence of molecular chaperones or a particular experimental setup, may favor one transition pathway over the other, which can result in very different monomers. This has some possible connections with the observation of various fibril morphologies and the outcome of prion strains. In addition, the finding that the interaction of H187 with R136 is a weak point in mammalian PrP is supported by the absence of the residue pair in non-mammalian species that are known to be resistant to prion diseases. PMID:23696721

Garrec, Julian; Tavernelli, Ivano; Rothlisberger, Ursula

2013-01-01

262

Combined Pharmacological Induction of Hsp70 Suppresses Prion Protein Neurotoxicity in Drosophila  

PubMed Central

Prion diseases are rare and aggressive neurodegenerative disorders caused by the accumulation of misfolded, toxic conformations of the prion protein (PrP). Therapeutic strategies directed at reducing the levels of PrP offer the best chance of delaying or halting disease progression. The challenge, though, is to define pharmacologic targets that result in reduced PrP levels. We previously reported that expression of wild type hamster PrP in flies induces progressive locomotor dysfunction and accumulation of pathogenic PrP conformations, while co-expression of human Hsp70 delayed these changes. To validate the therapeutic potential of Hsp70, we treated flies with drugs known to induce Hsp70 expression, including the Hsp90 inhibitor 17-DMAG and the glucocorticoid dexamethasone. Although the individual treatment with these compounds produced no significant benefits, their combination significantly increased the level of inducible Hsp70, decreased the level of total PrP, reduced the accumulation of pathogenic PrP conformers, and improved locomotor activity. Thus, the combined action of two pharmacological activators of Hsp70 with distinct targets results in sustained high levels of inducible Hsp70 with improved behavioral output. These findings can have important therapeutic applications for the devastating prion diseases and other related proteinopathies. PMID:24523910

Zhang, Yan; Casas-Tinto, Sergio; Rincon-Limas, Diego E.; Fernandez-Funez, Pedro

2014-01-01

263

Prion Protein and Shadoo Are Involved in Overlapping Embryonic Pathways and Trophoblastic Development  

PubMed Central

The potential requirement of either the Prion or Shadoo protein for early mouse embryogenesis was recently suggested. However, the current data did not allow to precise the developmental process that was affected in the absence of both proteins and that led to the observed early lethal phenotype. In the present study, using various Prnp transgenic mouse lines and lentiviral vectors expressing shRNAs that target the Shadoo-encoding mRNA, we further demonstrate the specific requirement of at least one of these two PrP-related proteins at early developmental stages. Histological analysis reveals developmental defect of the ectoplacental cone and important hemorrhage surrounding the Prnp-knockout-Sprn-knockdown E7.5 embryos. By restricting the RNA interference to the trophoblastic cell lineages, the observed lethal phenotype could be attributed to the sole role of these proteins in this trophectoderm-derived compartment. RNAseq analysis performed on early embryos of various Prnp and Sprn genotypes indicated that the simultaneous down-regulation of these two proteins affects cell-adhesion and inflammatory pathways as well as the expression of ectoplacental-specific genes. Overall, our data provide biological clues in favor of a crucial and complementary embryonic role of the prion protein family in Eutherians and emphasizes the need to further evaluate its implication in normal and pathological human placenta biology. PMID:22860039

Makhzami, Samira; Vilotte, Marthe; Jaffrezic, Florence; Halliez, Sophie; Bouet, Stephan; Marthey, Sylvain; Khalife, Manal; Kanellopoulos-Langevin, Colette; Beringue, Vincent; Le Provost, Fabienne; Laude, Hubert; Vilotte, Jean-Luc

2012-01-01

264

Kinetics of Amyloid Aggregation: A Study of the GNNQQNY Prion Sequence  

PubMed Central

The small amyloid-forming GNNQQNY fragment of the prion sequence has been the subject of extensive experimental and numerical studies over the last few years. Using unbiased molecular dynamics with the OPEP coarse-grained potential, we focus here on the onset of aggregation in a 20-mer system. With a total of 16.9 of simulations at 280 K and 300 K, we show that the GNNQQNY aggregation follows the classical nucleation theory (CNT) in that the number of monomers in the aggregate is a very reliable descriptor of aggregation. We find that the critical nucleus size in this finite-size system is between 4 and 5 monomers at 280 K and 5 and 6 at 300 K, in overall agreement with experiment. The kinetics of growth cannot be fully accounted for by the CNT, however. For example, we observe considerable rearrangements after the nucleus is formed, as the system attempts to optimize its organization. We also clearly identify two large families of structures that are selected at the onset of aggregation demonstrating the presence of well-defined polymorphism, a signature of amyloid growth, already in the 20-mer aggregate. PMID:23209391

Nasica-Labouze, Jessica; Mousseau, Normand

2012-01-01

265

Correlation of structure and infectivity of the HET-s prion  

PubMed Central

Prions are believed to be infectious self-propagating polymers of otherwise soluble host-encoded proteins1,2. This concept is now strongly supported by the recent findings that amyloid fibrils of recombinant prion proteins from yeast3-5, Podospora anserina6, and mammals7 can induce prion phenotypes in the corresponding hosts. However, the structural basis of prion infectivity remains largely elusive because acquisition of atomic resolution structural properties of amyloid fibrils represents a largely unsolved technical challenge. HET-s, the prion protein of P. anserina, contains a C-terminal prion domain comprising residues 218-289. Amyloid fibrils of HET-s(218-289) are necessary and sufficient for the induction and propagation of prion infectivity6. Here, we have used fluorescence studies, quenched hydrogen exchange NMR and solid state NMR to determine the sequence specific positions of secondary structure elements of amyloid fibrils of HET-s(218-289). This revealed four ?-strands constituted by two pseudo repeat sequences, each forming a ?-strand-turn-?-strand motif. We show that this conformation is the functional and infectious entity of the HET-s prion by using a structure-based mutagenesis approach. These results correlate for the first time distinct structural elements with prion infectivity. PMID:15944710

Ritter, Christiane; Maddelein, Marie-Lise; Siemer, Ansgar B.; Lührs, Thorsten; Ernst, Matthias; Meier, Beat H.; Saupe, Sven; Riek, Roland

2006-01-01

266

Conformational properties of prion strains can be transmitted to recombinant prion protein fibrils in real-time quaking-induced conversion.  

PubMed

The phenomenon of prion strains with distinct biological characteristics has been hypothesized to be involved in the structural diversity of abnormal prion protein (PrP(Sc)). However, the molecular basis of the transmission of strain properties remains poorly understood. Real-time quaking-induced conversion (RT-QUIC) is a cell-free system that uses Escherichia coli-derived recombinant PrP (rPrP) for the sensitive detection of PrP(Sc). To investigate whether the properties of various prion strains can be transmitted to amyloid fibrils consisting of rPrP (rPrP fibrils) using RT-QUIC, we examined the secondary structure, conformational stability, and infectivity of rPrP fibrils seeded with PrP(Sc) derived from either the Chandler or the 22L strain. In the first round of the reaction, there were differences in the secondary structures, especially in bands attributed to ?-sheets, as determined by infrared spectroscopy, and conformational stability between Chandler-seeded (1st-rPrP-fib(Ch)) and 22L-seeded (1st-rPrP-fib(22L)) rPrP fibrils. Of note, specific identifying characteristics of the two rPrP fibril types seen in the ?-sheets resembled those of the original PrP(Sc). Furthermore, the conformational stability of 1st-rPrP-fib(Ch) was significantly higher than that of 1st-rPrP-fib(22L), as with Chandler and 22L PrP(Sc). The survival periods of mice inoculated with 1st-rPrP-fib(Ch) or 1st-rPrP-fib(22L) were significantly shorter than those of mice inoculated with mixtures from the mock 1st-round RT-QUIC procedure. In contrast, these biochemical characteristics were no longer evident in subsequent rounds, suggesting that nonspecific uninfected rPrP fibrils became predominant probably because of their high growth rate. Together, these findings show that at least some strain-specific conformational properties can be transmitted to rPrP fibrils and unknown cofactors or environmental conditions may be required for further conservation. Importance: The phenomenon of prion strains with distinct biological characteristics is assumed to result from the conformational variations in the abnormal prion protein (PrP(Sc)). However, important questions remain about the mechanistic relationship between the conformational differences and the strain diversity, including how strain-specific conformations are transmitted. In this study, we investigated whether the properties of diverse prion strains can be transmitted to amyloid fibrils consisting of E. coli-derived recombinant PrP (rPrP) generated by real-time quaking-induced conversion (RT-QUIC), a recently developed in vitro PrP(Sc) formation method. We demonstrate that at least some of the strain-specific conformational properties can be transmitted to rPrP fibrils in the first round of RT-QUIC by examining the secondary structure, conformational stability, and infectivity of rPrP fibrils seeded with PrP(Sc) derived from either the Chandler or the 22L prion strain. We believe that these findings will advance our understanding of the conformational basis underlying prion strain diversity. PMID:25078700

Sano, Kazunori; Atarashi, Ryuichiro; Ishibashi, Daisuke; Nakagaki, Takehiro; Satoh, Katsuya; Nishida, Noriyuki

2014-10-01

267

Molecular dynamics studies on the NMR structures of rabbit prion protein wild-type and mutants: surface electrostatic charge distributions  

E-print Network

Prion is a misfolded protein found in mammals that causes infectious diseases of the nervous system in humans and animals. Prion diseases are invariably fatal and highly infectious neurodegenerative diseases that affect a wide variety of mammalian species such as sheep and goats, cattle, deer, elk and humans etc. Recent studies have shown that rabbits have a low susceptibility to be infected by prion diseases with respect to other animals including humans. The present study employs molecular dynamics (MD) means to unravel the mechanism of rabbit prion proteins (RaPrPC) based on the recently available rabbit NMR structures (of the wild-type and its two mutants of two surface residues). The electrostatic charge distributions on the protein surface are the focus when analysing the MD trajectories. It is found that we can conclude that surface electrostatic charge distributions indeed contribute to the structural stability of wild-type RaPrPC; this may be useful for the medicinal treatment of prion diseases.

Zhang, Jiapu

2014-01-01

268

Prions and the Oral Cavity  

Microsoft Academic Search

Prion diseases have recently emerged as a significant challenge to health-care workers, including those involved in dentistry. Abnormal prion proteins are resistant to complete inactivation by conventional sterilization techniques. In the last decade, a new form of prion disease emerged in the UK, termed “variant CJD”, thought to be acquired by consumption of bovine spongiform encephalopathy-contaminated food products. At present,

A. J. Smith; J. Bagg; J. W. Ironside; R. G. Will; C. Scully

2003-01-01

269

Degradation of the disease-associated prion protein by a serine protease from lichens.  

PubMed

The disease-associated prion protein (PrP(TSE)), the probable etiological agent of the transmissible spongiform encephalopathies (TSEs), is resistant to degradation and can persist in the environment. Lichens, mutualistic symbioses containing fungi, algae, bacteria and occasionally cyanobacteria, are ubiquitous in the environment and have evolved unique biological activities allowing their survival in challenging ecological niches. We investigated PrP(TSE) inactivation by lichens and found acetone extracts of three lichen species (Parmelia sulcata, Cladonia rangiferina and Lobaria pulmonaria) have the ability to degrade prion protein (PrP) from TSE-infected hamsters, mice and deer. Immunoblots measuring PrP levels and protein misfolding cyclic amplification indicated at least two logs of reductions in PrP(TSE). Degradative activity was not found in closely related lichen species or in algae or a cyanobacterium that inhabit lichens. Degradation was blocked by Pefabloc SC, a serine protease inhibitor, but not inhibitors of other proteases or enzymes. Additionally, we found that PrP levels in PrP(TSE)-enriched preps or infected brain homogenates are also reduced following exposure to freshly-collected P. sulcata or an aqueous extract of the lichen. Our findings indicate that these lichen extracts efficiently degrade PrP(TSE) and suggest that some lichens could have potential to inactivate TSE infectivity on the landscape or be a source for agents to degrade prions. Further work to clone and characterize the protease, assess its effect on TSE infectivity and determine which organism or organisms present in lichens produce or influence the protease activity is warranted. PMID:21589935

Johnson, Christopher J; Bennett, James P; Biro, Steven M; Duque-Velasquez, Juan Camilo; Rodriguez, Cynthia M; Bessen, Richard A; Rocke, Tonie E

2011-01-01

270

Degradation of the disease-associated prion protein by a serine protease from lichens.  

USGS Publications Warehouse

The disease-associated prion protein (PrPTSE), the probable etiological agent of the transmissible spongiform encephalopathies (TSEs), is resistant to degradation and can persist in the environment. Lichens, mutualistic symbioses containing fungi, algae, bacteria and occasionally cyanobacteria, are ubiquitous in the environment and have evolved unique biological activities allowing their survival in challenging ecological niches. We investigated PrPTSE inactivation by lichens and found acetone extracts of three lichen species (Parmelia sulcata, Cladonia rangiferina and Lobaria pulmonaria) have the ability to degrade prion protein (PrP) from TSE-infected hamsters, mice and deer. Immunoblots measuring PrP levels and protein misfolding cyclic amplification indicated at least two logs of reductions in PrPTSE. Degradative activity was not found in closely related lichen species or in algae or a cyanobacterium that inhabit lichens. Degradation was blocked by Pefabloc SC, a serine protease inhibitor, but not inhibitors of other proteases or enzymes. Additionally, we found that PrP levels in PrPTSE-enriched preps or infected brain homogenates are also reduced following exposure to freshly-collected P. sulcata or an aqueous extract of the lichen. Our findings indicate that these lichen extracts efficiently degrade PrPTSE and suggest that some lichens could have potential to inactivate TSE infectivity on the landscape or be a source for agents to degrade prions. Further work to clone and characterize the protease, assess its effect on TSE infectivity and determine which organism or organisms present in lichens produce or influence the protease activity is warranted.

Johnson, C. J.; Bennett, J. P.; Biro, S. M.; Duque-Velasquez, J. C.; Rodriguez, C. M.; Bessen, R. A.; Rocke, T. E.

2011-01-01

271

Degradation of the disease-associated prion protein by a serine protease from lichens  

USGS Publications Warehouse

The disease-associated prion protein (PrP(TSE)), the probable etiological agent of the transmissible spongiform encephalopathies (TSEs), is resistant to degradation and can persist in the environment. Lichens, mutualistic symbioses containing fungi, algae, bacteria and occasionally cyanobacteria, are ubiquitous in the environment and have evolved unique biological activities allowing their survival in challenging ecological niches. We investigated PrP(TSE) inactivation by lichens and found acetone extracts of three lichen species (Parmelia sulcata, Cladonia rangiferina and Lobaria pulmonaria) have the ability to degrade prion protein (PrP) from TSE-infected hamsters, mice and deer. Immunoblots measuring PrP levels and protein misfolding cyclic amplification indicated at least two logs of reductions in PrP(TSE). Degradative activity was not found in closely related lichen species or in algae or a cyanobacterium that inhabit lichens. Degradation was blocked by Pefabloc SC, a serine protease inhibitor, but not inhibitors of other proteases or enzymes. Additionally, we found that PrP levels in PrP(TSE)-enriched preps or infected brain homogenates are also reduced following exposure to freshly-collected P. sulcata or an aqueous extract of the lichen. Our findings indicate that these lichen extracts efficiently degrade PrP(TSE) and suggest that some lichens could have potential to inactivate TSE infectivity on the landscape or be a source for agents to degrade prions. Further work to clone and characterize the protease, assess its effect on TSE infectivity and determine which organism or organisms present in lichens produce or influence the protease activity is warranted.

Johnson, C.J.; Bennett, J.P.; Biro, S.M.; Duque-Velasquez, J.C.; Rodriguez, C.M.; Bessen, R.A.; Rocke, T.E.

2011-01-01

272

Degradation of the disease-associated prion protein by a serine protease from lichens  

USGS Publications Warehouse

The disease-associated prion protein (PrPTSE), the probable etiological agent of the transmissible spongiform encephalopathies (TSEs), is resistant to degradation and can persist in the environment. Lichens, mutualistic symbioses containing fungi, algae, bacteria and occasionally cyanobacteria, are ubiquitous in the environment and have evolved unique biological activities allowing their survival in challenging ecological niches. We investigated PrPTSE inactivation by lichens and found acetone extracts of three lichen species (Parmelia sulcata, Cladonia rangiferina and Lobaria pulmonaria) have the ability to degrade prion protein (PrP) from TSE-infected hamsters, mice and deer. Immunoblots measuring PrP levels and protein misfolding cyclic amplification indicated at least two logs of reductions in PrPTSE. Degradative activity was not found in closely related lichen species or in algae or a cyanobacterium that inhabit lichens. Degradation was blocked by Pefabloc SC, a serine protease inhibitor, but not inhibitors of other proteases or enzymes. Additionally, we found that PrP levels in PrPTSE-enriched preps or infected brain homogenates are also reduced following exposure to freshly-collected P. sulcata or an aqueous extract of the lichen. Our findings indicate that these lichen extracts efficiently degrade PrPTSE and suggest that some lichens could have potential to inactivate TSE infectivity on the landscape or be a source for agents to degrade prions. Further work to clone and characterize the protease, assess its effect on TSE infectivity and determine which organism or organisms present in lichens produce or influence the protease activity is warranted.

Johnson, C. J.; Bennett, J. P.; Biro, S. M.; Duque-Velasquez, J. C.; Rodriguez, C. M.; Bessen, R. A.; Rocke, T. E.

2011-01-01

273

Effect of Charged Residues in the N-domain of Sup35 Protein on Prion [PSI+] Stability and Propagation*  

PubMed Central

Recent studies have shown that Sup35p prion fibrils probably have a parallel in-register ?-structure. However, the part(s) of the N-domain critical for fibril formation and maintenance of the [PSI+] phenotype remains unclear. Here we designed a set of five SUP35 mutant alleles (sup35KK) with lysine substitutions in each of five N-domain repeats, and investigated their effect on infectivity and ability of corresponding proteins to aggregate and coaggregate with wild type Sup35p in the [PSI+] strain. Alleles sup35-M1 (Y46K/Q47K) and sup35-M2 (Q61K/Q62K) led to prion loss, whereas sup35-M3 (Q70K/Q71K), sup35-M4 (Q80K/Q81K), and sup35-M5 (Q89K/Q90K) were able to maintain the [PSI+] prion. This suggests that the critical part of the parallel in-register ?-structure for the studied [PSI+] prion variant lies in the first 63–69 residues. Our study also reveals an unexpected interplay between the wild type Sup35p and proteins expressed from the sup35KK alleles during prionization. Both Sup35-M1p and Sup35-M2p coaggregated with Sup35p, but only sup35-M2 led to prion loss in a dominant manner. We suggest that in the fibrils, Sup35p can bind to Sup35-M1p in the same conformation, whereas Sup35-M2p only allowed the Sup35p conformation that leads to the non-heritable fold. Mutations sup35-M4 and sup35-M5 influence the structure of the prion forming region to a lesser extent, and can lead to the formation of new prion variants. PMID:23965990

Bondarev, Stanislav A.; Shchepachev, Vadim V.; Kajava, Andrey V.; Zhouravleva, Galina A.

2013-01-01

274

Probing the N-terminal ?-sheet conversion in the crystal structure of the human prion protein bound to a nanobody.  

PubMed

Prions are fatal neurodegenerative transmissible agents causing several incurable illnesses in humans and animals. Prion diseases are caused by the structural conversion of the cellular prion protein, PrP(C), into its misfolded oligomeric form, known as prion or PrP(Sc). The canonical human PrP(C) (HuPrP) fold features an unstructured N-terminal part (residues 23-124) and a well-defined C-terminal globular domain (residues 125-231). Compelling evidence indicates that an evolutionary N-terminal conserved motif AGAAAAGA (residues 113-120) plays an important role in the conversion to PrP(Sc). The intrinsic flexibility of the N-terminal has hampered efforts to obtain detailed atomic information on the structural features of this palindromic region. In this study, we crystallized the full-length HuPrP in complex with a nanobody (Nb484) that inhibits prion propagation. In the complex, the prion protein is unstructured from residue 23 to 116. The palindromic motif adopts a stable and fully extended configuration to form a three-stranded antiparallel ?-sheet with the ?1 and ?2 strands, demonstrating that the full-length HuPrP(C) can adopt a more elaborate ?0-?1-?1-?2-?2-?3 structural organization than the canonical ?1-?1-?2-?2-?3 prion-like fold. From this structure, it appears that the palindromic motif mediates ?-enrichment in the PrP(C) monomer as one of the early events in the conversion of PrP(C) into PrP(Sc). PMID:24400836

Abskharon, Romany N N; Giachin, Gabriele; Wohlkonig, Alexandre; Soror, Sameh H; Pardon, Els; Legname, Giuseppe; Steyaert, Jan

2014-01-22

275

Biochem. J. (2006) 399, 435444 (Printed in Great Britain) doi:10.1042/BJ20060458 435 A reassessment of copper(II) binding in the full-length prion protein  

E-print Network

A reassessment of copper(II) binding in the full-length prion protein Mark A. WELLS*, Graham S. JACKSON, Samantha, University of Sheffield, Sheffield S10 2TN, U.K., and MRC Prion Unit, Department of Neurodegenerative Disease previously that the unfolded N-terminal domain of the prion protein can bind up to six Cu2+ ions in vitro

Hosszu, Laszlo

276

Functional Depletion of Mahogunin by Cytosolically Exposed Prion Protein Contributes to Neurodegeneration  

Microsoft Academic Search

SUMMARY The pathways leading from aberrant Prion protein (PrP) metabolism to neurodegeneration are poorly understood. Some familial PrP mutants generate increased CtmPrP, a transmembrane isoform associ- atedwithdisease.Inotherdiseasesituations,apoten- tially toxic cytosolic form (termed cyPrP) might be produced. However, the mechanisms by which Ctm PrPorcyPrPcauseselectiveneuronaldysfunction are unknown. Here, we show that both CtmPrP and cyPrPcaninteractwithanddisruptthefunctionofMa- hogunin (Mgrn), a cytosolic ubiquitin ligase whose

Oishee Chakrabarti; Ramanujan S. Hegde

2009-01-01

277

Distinct Proteinase K-Resistant Prion Protein Fragment in Goats with No Signs of Disease in a Classical Scrapie Outbreak?†  

PubMed Central

Considerable efforts have been directed toward the identification of small-ruminant prion diseases, i.e., classical and atypical scrapie as well as bovine spongiform encephalopathy (BSE). Here we report the in-depth molecular analysis of the proteinase K-resistant prion protein core fragment (PrPres) in a highly scrapie-affected goat flock in Greece. The PrPres profile by Western immunoblotting in most animals was that of classical scrapie in sheep. However, in a series of clinically healthy goats we identified a unique C- and N-terminally truncated PrPres fragment, which is akin but not identical to that observed for atypical scrapie. These findings reveal novel aspects of the nature and diversity of the molecular PrPres phenotypes in goats and suggest that these animals display a previously unrecognized prion protein disorder. PMID:21450953

Bouzalas, Ilias G.; Lortscher, Florian; Dovas, Chrysostomos I.; Oevermann, Anna; Langeveld, Jan P. M.; Papanastassopoulou, Maria; Papadopoulos, Orestis; Zurbriggen, Andreas; Seuberlich, Torsten

2011-01-01

278

Distinct proteinase K-resistant prion protein fragment in goats with no signs of disease in a classical scrapie outbreak.  

PubMed

Considerable efforts have been directed toward the identification of small-ruminant prion diseases, i.e., classical and atypical scrapie as well as bovine spongiform encephalopathy (BSE). Here we report the in-depth molecular analysis of the proteinase K-resistant prion protein core fragment (PrP(res)) in a highly scrapie-affected goat flock in Greece. The PrP(res) profile by Western immunoblotting in most animals was that of classical scrapie in sheep. However, in a series of clinically healthy goats we identified a unique C- and N-terminally truncated PrP(res) fragment, which is akin but not identical to that observed for atypical scrapie. These findings reveal novel aspects of the nature and diversity of the molecular PrP(res) phenotypes in goats and suggest that these animals display a previously unrecognized prion protein disorder. PMID:21450953

Bouzalas, Ilias G; Lörtscher, Florian; Dovas, Chrysostomos I; Oevermann, Anna; Langeveld, Jan P M; Papanastassopoulou, Maria; Papadopoulos, Orestis; Zurbriggen, Andreas; Seuberlich, Torsten

2011-06-01

279

A Review on the Salt Bridge Between ASP177 and ARG163 of Wild-Type Rabbit Prion Protein  

E-print Network

Prion diseases are invariably fatal and highly infectious neurodegenerative diseases that affect a wide variety of mammalian species such as sheep and goats, cattle, deer, elks, humans and mice etc., but rabbits have a low susceptibility to be infected by prion diseases with respect to other species. The stability of rabbit prion protein is due to its highly ordered beta2-alpha2 loop [PLoS One 5 (10) e13273 (2010); Journal of Biological Chemistry 285 (41) 31682-31693 (2010)] and a helix-capping motif within this loop [PLoS One 8 (5) e63047 (2013)]. The beta2-alpha2 loop has been a focus in prion studies. For this loop we found a salt bridge linkage ASP177-ARG163 (O-N) [Journal of Theoretical Biology 342 (7 February 2014) 70-82 (2014)]. Some scientists said on the 2FJ3.pdb NMR file of the rabbit prion protein, the distance of ASP177-ARG163 (O-N) gives the salt bridge of about 10 angstroms which is nearly null in terms of energy thus think our result is wrong. This opinion is clearly wrong simply due to the 3O7...

Zhang, Jiapu

2014-01-01

280

Identification and structural analysis of C-terminally truncated collapsin response mediator protein-2 in a murine model of prion diseases  

PubMed Central

Background Prion diseases are fatal neurodegenerative disorders that accompany an accumulation of the disease-associated form(s) of prion protein (PrPSc) in the central nervous system. The neuropathological changes in the brain begin with focal deposits of PrPSc, followed by pathomorphological abnormalities of axon terminal degeneration, synaptic loss, atrophy of dendritic trees, and eventual neuronal cell death in the lesions. However, the underlying molecular basis for these neuropathogenic abnormalities is not fully understood. Results In a proteomic analysis of soluble proteins in the brains of mice challenged intracerebrally with scrapie prion (Obihiro I strain), we found that the amount of the full-length form of collapsin response mediator protein-2 (CRMP-2; 61 kDa) decreased in the late stages of the disease, while the amount of its truncated form (56 kDa) increased to comparable levels observed for the full-length form. Detailed analysis by liquid chromatography-electrospray ionization-tandem mass spectrometry showed that the 56-kDa form (named CRMP-2-?C) lacked the sequence from serine518 to the C-terminus, including the C-terminal phosphorylation sites important for the regulation of axonal growth and axon-dendrite specification in developing neurons. The invariable size of the mRNA transcript in Northern blot analysis suggested that the truncation was due to post-translational proteolysis. By overexpression of CRMP-2-?C in primary cultured neurons, we observed the augmentation of the development of neurite branch tips to the same levels as for CRMP-2T514A/T555A, a non-phosphorylated mimic of the full-length protein. This suggests that the increased level of CRMP-2-?C in the brain modulates the integrity of neurons, and may be involved in the pathogenesis of the neuronal abnormalities observed in the late stages of the disease. Conclusions We identified the presence of CRMP-2-?C in the brain of a murine model of prion disease. Of note, C-terminal truncations of CRMP-2 have been recently observed in models for neurodegenerative disorders such as ischemia, traumatic brain injury, and Wallerian degeneration. While the structural identity of CRMP-2-?C in those models remains unknown, the present study should provide clues to the molecular pathology of degenerating neurons in prion diseases in connection with other neurodegenerative disorders. PMID:20961402

2010-01-01

281

The [PSI+] Prion Exists as a Dynamic Cloud of Variants  

PubMed Central

[PSI+] is an amyloid-based prion of Sup35p, a subunit of the translation termination factor. Prion “strains” or “variants” are amyloids with different conformations of a single protein sequence, conferring different phenotypes, but each relatively faithfully propagated. Wild Saccharomyces cerevisiae isolates have SUP35 alleles that fall into three groups, called reference, ?19, and E9, with limited transmissibility of [PSI+] between cells expressing these different polymorphs. Here we show that prion transmission pattern between different Sup35 polymorphs is prion variant-dependent. Passage of one prion variant from one Sup35 polymorph to another need not change the prion variant. Surprisingly, simple mitotic growth of a [PSI+] strain results in a spectrum of variant transmission properties among the progeny clones. Even cells that have grown for >150 generations continue to vary in transmission properties, suggesting that simple variant segregation is insufficient to explain the results. Rather, there appears to be continuous generation of a cloud of prion variants, with one or another becoming stochastically dominant, only to be succeeded by a different mixture. We find that among the rare wild isolates containing [PSI+], all indistinguishably “weak” [PSI+], are several different variants based on their transmission efficiencies to other Sup35 alleles. Most show some limitation of transmission, indicating that the evolved wild Sup35 alleles are effective in limiting the spread of [PSI+]. Notably, a “strong [PSI+]” can have any of several different transmission efficiency patterns, showing that “strong” versus “weak” is insufficient to indicate prion variant uniformity. PMID:23382698

Bateman, David A.; Wickner, Reed B.

2013-01-01

282

Single-particle tracking of quantum dot-conjugated prion proteins inside yeast cells  

SciTech Connect

Research highlights: {yields} We develop a method to track a quantum dot-conjugated protein in yeast cells. {yields} We incorporate the conjugated quantum dot proteins into yeast spheroplasts. {yields} We track the motions by conventional or 3D tracking microscopy. -- Abstract: Yeast is a model eukaryote with a variety of biological resources. Here we developed a method to track a quantum dot (QD)-conjugated protein in the budding yeast Saccharomyces cerevisiae. We chemically conjugated QDs with the yeast prion Sup35, incorporated them into yeast spheroplasts, and tracked the motions by conventional two-dimensional or three-dimensional tracking microscopy. The method paves the way toward the individual tracking of proteins of interest inside living yeast cells.

Tsuji, Toshikazu; Kawai-Noma, Shigeko [Department of Biomolecular Engineering, Graduate School of Biosciences and Biotechnology, Tokyo Institute of Technology, B56, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501 (Japan)] [Department of Biomolecular Engineering, Graduate School of Biosciences and Biotechnology, Tokyo Institute of Technology, B56, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501 (Japan); Pack, Chan-Gi [Cellular Informatics Laboratory, RIKEN Advanced Science Institute, Wako-shi, Saitama 351-0198 (Japan)] [Cellular Informatics Laboratory, RIKEN Advanced Science Institute, Wako-shi, Saitama 351-0198 (Japan); Terajima, Hideki [Department of Biomolecular Engineering, Graduate School of Biosciences and Biotechnology, Tokyo Institute of Technology, B56, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501 (Japan)] [Department of Biomolecular Engineering, Graduate School of Biosciences and Biotechnology, Tokyo Institute of Technology, B56, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501 (Japan); Yajima, Junichiro; Nishizaka, Takayuki [Department of Physics, Gakushuin University, 1-5-1 Mejiro, Toshima-ku, Tokyo 171-8588 (Japan)] [Department of Physics, Gakushuin University, 1-5-1 Mejiro, Toshima-ku, Tokyo 171-8588 (Japan); Kinjo, Masataka [Laboratory of Molecular Cell Dynamics, Graduate School of Life Sciences, Hokkaido University, Sapporo 001-0021 (Japan)] [Laboratory of Molecular Cell Dynamics, Graduate School of Life Sciences, Hokkaido University, Sapporo 001-0021 (Japan); Taguchi, Hideki, E-mail: taguchi@bio.titech.ac.jp [Department of Biomolecular Engineering, Graduate School of Biosciences and Biotechnology, Tokyo Institute of Technology, B56, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501 (Japan)] [Department of Biomolecular Engineering, Graduate School of Biosciences and Biotechnology, Tokyo Institute of Technology, B56, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501 (Japan)

2011-02-25

283

Prion Protein-Specific Antibodies that Detect Multiple TSE Agents with High Sensitivity  

PubMed Central

This paper describes the generation, characterisation and potential applications of a panel of novel anti-prion protein monoclonal antibodies (mAbs). The mAbs were generated by immunising PRNP null mice, using a variety of regimes, with a truncated form of recombinant ovine prion protein spanning residues 94–233. Epitopes of specific antibodies were mapped using solid-phase Pepscan analysis and clustered to four distinct regions within the PrP molecule. We have demonstrated the utility of these antibodies by use of Western blotting and immunohistochemistry in tissues from a range of different species affected by transmissible spongiform encephalopathy (TSE). In comparative tests against extensively-used and widely-published, commercially available antibodies, similar or improved results can be obtained using these new mAbs, specifically in terms of sensitivity of detection. Since many of these antibodies recognise native PrPC, they could also be applied to a broad range of immunoassays such as flow cytometry, DELFIA analysis or immunoprecipitation. We are using these reagents to increase our understanding of TSE pathogenesis and for use in potential diagnostic screening assays. PMID:24608105

McCutcheon, Sandra; Langeveld, Jan P. M.; Tan, Boon Chin; Gill, Andrew C.; de Wolf, Christopher; Martin, Stuart; Gonzalez, Lorenzo; Alibhai, James; Blanco, A. Richard Alejo; Campbell, Lauren; Hunter, Nora; Houston, E. Fiona

2014-01-01

284

Prion protein-specific antibodies that detect multiple TSE agents with high sensitivity.  

PubMed

This paper describes the generation, characterisation and potential applications of a panel of novel anti-prion protein monoclonal antibodies (mAbs). The mAbs were generated by immunising PRNP null mice, using a variety of regimes, with a truncated form of recombinant ovine prion protein spanning residues 94-233. Epitopes of specific antibodies were mapped using solid-phase Pepscan analysis and clustered to four distinct regions within the PrP molecule. We have demonstrated the utility of these antibodies by use of Western blotting and immunohistochemistry in tissues from a range of different species affected by transmissible spongiform encephalopathy (TSE). In comparative tests against extensively-used and widely-published, commercially available antibodies, similar or improved results can be obtained using these new mAbs, specifically in terms of sensitivity of detection. Since many of these antibodies recognise native PrPC, they could also be applied to a broad range of immunoassays such as flow cytometry, DELFIA analysis or immunoprecipitation. We are using these reagents to increase our understanding of TSE pathogenesis and for use in potential diagnostic screening assays. PMID:24608105

McCutcheon, Sandra; Langeveld, Jan P M; Tan, Boon Chin; Gill, Andrew C; de Wolf, Christopher; Martin, Stuart; Gonzalez, Lorenzo; Alibhai, James; Blanco, A Richard Alejo; Campbell, Lauren; Hunter, Nora; Houston, E Fiona

2014-01-01

285

High incidence of single nucleotide polymorphisms in the prion protein gene of native Brazilian Caracu cattle.  

PubMed

Different alleles of the human and ovine prion protein gene correlate with a varying susceptibility to transmissible spongiform encephalopathies. However, the pathogenic implications of specific polymorphisms in the bovine prion protein gene (PRNP) are only poorly understood. Previous studies on the bovine PRNP gene investigated common European and North American cattle breeds. As a consequence of decades of intensive breeding for specific traits, these modern breeds represent only a small fraction of the bovine gene pool. In this study, we analysed PRNP polymorphisms in the native Brazilian Caracu breed, which developed in geographical isolation since the 16th century. A total of 10 single nucleotide polymorphisms (SNPs) were discovered in the coding region of the Caracu PRNP gene. Eight of the SNPs occurred at high frequencies in Caracu cattle (variant allele frequencies = 0.10-0.76), but were absent or only rarely observed in European and North American breeds. One of the Caracu SNPs was associated with an amino acid exchange from serine to asparagine (f = 0.17). This SNP was not detected in Holstein-Friesian, Simmental and German Gelbvieh and was only rarely detected in beef cattle (f = 0.01). We found 17 haplotypes for PRNP in the Caracu breed. PMID:16965405

Kues, W A; Ollhoff, R D; Carnwath, J W; de Souza, F P; Madeira, H M F; Niemann, H

2006-10-01

286

A novel expression system for production of soluble prion proteins in E. coli  

PubMed Central

Expression of eukaryotic proteins in Escherichia coli is challenging, especially when they contain disulfide bonds. Since the discovery of the prion protein (PrP) and its role in transmissible spongiform encephalopathies, the need to obtain large quantities of the recombinant protein for research purposes has been essential. Currently, production of recombinant PrP is achieved by refolding protocols. Here, we show that the co-expression of two different PrP with the human Quiescin Sulfhydryl OXidase (QSOX), a human chaperone with thiol/disulfide oxidase activity, in the cytoplasm of E. coli produces soluble recombinant PrP. The structural integrity of the soluble PrP has been confirmed by nuclear magnetic resonance spectroscopy, demonstrating that properly folded PrP can be easily expressed in bacteria. Furthermore, the soluble recombinant PrP produced with this method can be used for functional and structural studies. PMID:22233534

2012-01-01

287

Prolonged incubation time in sheep with prion protein containing lysine at position 171.  

PubMed

Sheep scrapie susceptibility or resistance is a function of genotype, with polymorphisms at codon 171 in the sheep prion gene playing a major role. Glutamine (Q) at codon 171 contributes to scrapie susceptibility, while arginine (R) is associated with resistance. In some breeds, lysine (K) occurs at codon 171, but its effect on scrapie resistance has not been determined. Charge and structural similarities between K and R suggest that they may contribute to prion disease susceptibility in a similar way, but studies have not been performed to confirm this. The purpose of the current study was to compare susceptibility and incubation times of AA(136)RR(154)QQ(171) (where the letter denotes the amino acid and the number the position) with AA(136)RR(154)QK(171) sheep after inoculation with scrapie. Barbado AA(136)RR(154)QQ(171) and AA(136)RR(154)QK(171) sheep were inoculated with scrapie intracerebrally to assess their susceptibility to scrapie. After inoculation, sheep were observed daily for clinical signs and were euthanized and necropsied after clinical signs were unequivocal. Tissues were collected at necropsy for immunohistochemistry and Western blot analyses. The QQ(171) sheep had clinical signs approximately 12 months after inoculation, whereas QK(171) animals had an average incubation time of 30 months to onset of clinical signs. The distribution of abnormal prion protein was similar in QQ(171) and QK(171) sheep. Results of the study indicate that sheep with a single K allele at codon 171 are susceptible to scrapie but with a prolonged incubation time. Work is currently underway to examine relative scrapie susceptibility or resistance of KK(171) sheep. PMID:22529124

Greenlee, Justin J; Zhang, Xia; Nicholson, Eric M; Kunkle, Robert A; Hamir, Amir N

2012-05-01

288

Impact of methionine oxidation as an initial event on the pathway of human prion protein conversion.  

PubMed

Prion diseases comprise a group of fatal neurodegenerative disorders characterized by the autocatalytic conversion of the cellular prion protein PrP(C) into the infectious misfolded isoform PrP(Sc). Increasing evidence supports a specific role of oxidative stress in the onset of pathogenesis. Although the associated molecular mechanisms remain to be elucidated in detail, several studies currently suggest that methionine oxidation already detected in misfolded PrP(Sc) destabilizes the native PrP fold as an early event in the conversion pathway. To obtain more insights about the specific impact of surface-exposed methionine residues on the oxidative-induced conversion of human PrP we designed, produced, and comparatively investigated two new pseudosulfoxidation mutants of human PrP 121-231 that comprises the well-folded C-terminal domain. Applying circular dichroism spectroscopy and dynamic light scattering techniques we showed that pseudosulfoxidation of all surface exposed Met residues formed a monomeric molten globule-like species with striking similarities to misfolding intermediates recently reported by other groups. However, individual pseudosulfoxidation at the polymorphic M129 site did not significantly contribute to the structural destabilization. Further metal-induced oxidation of the partly unfolded pseudosulfoxidation mutant resulted in the formation of an oligomeric state that shares a comparable size and stability with PrP oligomers detected after the application of different other triggers for structural conversion, indicating a generic misfolding pathway of PrP. The obtained results highlight the specific importance of methionine oxidation at surface exposed residues for PrP misfolding, strongly supporting the hypothesis that increased oxidative stress could be one causative event for sporadic prion diseases and other neurodegenerative disorders. PMID:24121542

Elmallah, Mohammed I Y; Borgmeyer, Uwe; Betzel, Christian; Redecke, Lars

2013-01-01

289

Prion recognition elements govern nucleation, strain specificity and species  

E-print Network

ARTICLES Prion recognition elements govern nucleation, strain specificity and species barriers Peter M. Tessier1 & Susan Lindquist2 Prions are proteins that can switch to self-perpetuating, infectious conformations. The abilities of prions to replicate, form structurally distinct strains

Lindquist, Susan

290

Sequence repeats and protein structure  

NASA Astrophysics Data System (ADS)

Repeats are frequently found in known protein sequences. The level of sequence conservation in tandem repeats correlates with their propensities to be intrinsically disordered. We employ a coarse-grained model of a protein with a two-letter amino acid alphabet, hydrophobic (H) and polar (P), to examine the sequence-structure relationship in the realm of repeated sequences. A fraction of repeated sequences comprises a distinct class of bad folders, whose folding temperatures are much lower than those of random sequences. Imperfection in sequence repetition improves the folding properties of the bad folders while deteriorating those of the good folders. Our results may explain why nature has utilized repeated sequences for their versatility and especially to design functional proteins that are intrinsically unstructured at physiological temperatures.

Hoang, Trinh X.; Trovato, Antonio; Seno, Flavio; Banavar, Jayanth R.; Maritan, Amos

2012-11-01

291

A Variational Model for Oligomer-Formation Process of GNNQQNY Peptide from Yeast Prion Protein Sup35  

E-print Network

A Variational Model for Oligomer-Formation Process of GNNQQNY Peptide from Yeast Prion Protein Sup, Tennessee; and § Zhou Pei-Yuan Center for Applied Mathematics, Tsinghua University, Beijing, China #12 that the critical nucleus size for homogeneous nucleation process is 3 peptides. Here we set the minimal step size

Zhang, Yang

292

Prion Protein is Expressed on Long-term Repopulating Hematopoietic Stem Cells and is Necessary for their Self-renewal  

E-print Network

We show that the prion protein (PrP) is expressed on the surface of bone marrow cell populations enriched in long-term repopulating hematopoietic stem cells. Affinity purification of the PrP-positive and PrP-negative ...

Lodish, Harvey F.

293

A MUTANT PRION PROTEIN SENSITIZES NEURONS TO GLUTAMATE-INDUCED EXCITOTOXICITY  

PubMed Central

Growing evidence suggests that a physiological activity of the cellular prion protein (PrPC) plays a crucial role in several neurodegenerative disorders, including prion and Alzheimer’s diseases. However, how the functional activity of PrPC is subverted to deliver neurotoxic signals remains uncertain. Transgenic mice expressing PrP with a deletion of residues 105–125 in the central region (referred to as ?CR PrP) provide important insights into this problem. Tg(?CR) mice exhibit neonatal lethality and massive degeneration of cerebellar granule neurons, a phenotype that is dose-dependently suppressed by the presence of wild-type PrP. When expressed in cultured cells, ?CR PrP induces large, ionic currents that can be detected by patch-clamping techniques. Here, we have tested the hypothesis that abnormal ion channel activity underlies the neuronal death seen in Tg(?CR) mice. We find that ?CR PrP induces abnormal ionic currents in neurons in culture and in cerebellar slices, and that this activity sensitizes the neurons to glutamate-induced, calcium-mediated death. In combination with ultrastructural and biochemical analyses, these results demonstrate a role for glutamate-induced excitotoxicity in PrP-mediated neurodegeneration. A similar mechanism may operate in other neurodegenerative disorders due to toxic, ?-rich oligomers that bind to PrPC. PMID:23392670

Biasini, Emiliano; Unterberger, Ursula; Solomon, Isaac H.; Massignan, Tania; Senatore, Assunta; Bian, Hejiao; Voigtlaender, Till; Bowman, Frederick P.; Bonetto, Valentina; Chiesa, Roberto; Luebke, Jennifer; Toselli, Paul; Harris, David A.

2013-01-01

294

Separate mechanisms act concurrently to shed and release the prion protein from the cell  

PubMed Central

The cellular prion protein (PrPC) is attached to the cell membrane via its glycosylphosphatidylinositol (GPI)-anchor and is constitutively shed into the extracellular space. Here, three different mechanisms are presented that concurrently shed PrPC from the cell. The fast ?-cleavage released a N-terminal fragment (N1) into the medium and the extreme C-terminal cleavage shed soluble full-length (FL-S) PrP and C-terminally cleaved (C1-S) fragments outside the cell. Also, a slow exosomal release of full-length (FL) and C1-fragment (C1) was demonstrated. The three separate mechanisms acting simultaneously, but with different kinetics, have to be taken into consideration when elucidating functional roles of PrPC and also when processing of PrPC is considered as a target for intervention in prion diseases. Further, in this study it was shown that metalloprotease inhibitors affected the extreme C-terminal cleavage and shedding of PrPC. The metalloprotease inhibitors did not influence the ?-cleavage or the exosomal release. Taken together, these results are important for understanding the different mechanisms acting in parallel in the shedding and cleavage of PrPC. PMID:23093798

Wik, Lotta; Klingeborn, Mikael; Willander, Hanna; Linne, Tommy

2012-01-01

295

De novo generation of prion strains  

Microsoft Academic Search

Prions are self-replicating proteins that can cause neurodegenerative disorders such as bovine spongiform encephalopathy (also known as mad cow disease). Aberrant conformations of prion proteins accumulate in the central nervous system, causing spongiform changes in the brain and eventually death. Since the inception of the prion hypothesis — which states that misfolded proteins are the infectious agents that cause these

David W. Colby

2011-01-01

296

Non-targeted Identification of Prions and Amyloid-forming Proteins from Yeast and Mammalian Cells*  

PubMed Central

The formation of amyloid aggregates is implicated both as a primary cause of cellular degeneration in multiple human diseases and as a functional mechanism for providing extraordinary strength to large protein assemblies. The recent identification and characterization of several amyloid proteins from diverse organisms argues that the amyloid phenomenon is widespread in nature. Yet identifying new amyloid-forming proteins usually requires a priori knowledge of specific candidates. Amyloid fibers can resist heat, pressure, proteolysis, and denaturation by reagents such as urea or sodium dodecyl sulfate. Here we show that these properties can be exploited to identify naturally occurring amyloid-forming proteins directly from cell lysates. This proteomic-based approach utilizes a novel purification of amyloid aggregates followed by identification by mass spectrometry without the requirement for special genetic tools. We have validated this technique by blind identification of three amyloid-based yeast prions from laboratory and wild strains and disease-related polyglutamine proteins expressed in both yeast and mammalian cells. Furthermore, we found that polyglutamine aggregates specifically recruit some stress granule components, revealing a possible mechanism of toxicity. Therefore, core amyloid-forming proteins as well as strongly associated proteins can be identified directly from cells of diverse origin. PMID:23926098

Kryndushkin, Dmitry; Pripuzova, Natalia; Burnett, Barrington G.; Shewmaker, Frank

2013-01-01

297

Radar chart deviation analysis of prion protein amino acid composition defines characteristic structural abnormalities of the N-terminal octa-peptide tandem repeat.  

PubMed

Analysis of the amino acid composition of prion protein using a newly developed program for radar-chart deviation analysis has identified an abnormality or irregularity of the N-terminal flexible domain. Aromatic amino acids Trp and His together with Gly are abnormally abounding in this N-terminal domain, in which octapeptide GQPHGGGW is connected four times in tandem. This tetrarepeat structure has been suggested to be essential for the prion protein not only to play an intrinsic functional role in the physiological condition, but also to bring on structural abnormalities in prion disease. PMID:18991771

Yokotani, Satoru; Nose, Takeru; Horiuchi, Yuji; Matsushima, Ayami; Shimohigashi, Yasuyuki

2008-01-01

298

Variant-specific prion interactions  

PubMed Central

Prions are protein conformations that “self-seed” the misfolding of their non-prion iso-forms into prion, often amyloid, conformations. The most famous prion is the mammalian PrP protein that in its prion form causes transmissible spongiform encephalopathy. Curiously there can be distinct conformational differences even between prions of the same protein propagated in the same host species. These are called prion strains or variants. For example, different PrP variants are faithfully transmitted during self-seeding and are associated with distinct disease characteristics. Variant-specific PrP prion differences include the length of the incubation period before the disease appears and the deposition of prion aggregates in distinct regions of the brain.1 Other more common neurodegenerative diseases (e.g., Alzheimer disease, Parkinson disease, type 2 diabetes and ALS) are likewise caused by the misfolding of a normal protein into a self-seeding aggregate.2-4 One of the most important unanswered questions is how the first prion-like seed arises de novo, resulting in the pathological cascade. PMID:24475372

Sharma, Jaya; Liebman, Susan W

2013-01-01

299

BSE-associated polymorphisms in the prion protein gene: an investigation.  

PubMed

The aim of this study was to determine the frequency of the 12-bp and 23-bp indel polymorphisms in the prion protein gene (PRNP) in cattle and to investigate the association between these frequencies and the occurrence of bovine spongiform encephalopathy (BSE). There was no significant difference in the 12-bp indel frequency between the BSE animals and control group. For the 23-bp indel, the BSE animals had a significantly lower + + (insins) genotype frequency and + allele frequency compared with the control animals. The - - / - - genotype frequency in the BSE animals was not significantly higher when compared with the control animals. One - allele increased the risk of BSE by a factor of 1.55 (i.e. by 55%) for the 12-bp indel and by a factor of 2.10 for the 23-bp indel. When both indels are considered, one - allele increased the risk of BSE by a factor of 1.54. PMID:24720684

Vernerova, K; Tothova, L; Mikova, A; Vodrazka, P; Simek, B; Hanusova, L; Citek, J

2014-10-01

300

Impairment of superoxide dismutase activation by N-terminally truncated prion protein (PrP) in PrP-deficient neuronal cell line  

Microsoft Academic Search

Previous studies have reported a neuroprotective role for cellular prion protein (PrPC) against apoptosis induced by serum deprivation in an immortalized prion protein gene (Prnp)-deficient neuronal cell line, but the mechanisms remain unclear. In this study, to investigate the mechanisms by which PrPC prevents apoptosis, the authors compared apoptosis of Prnp?\\/? cells with that of Prnp?\\/? cells expressing the wild-type

Akikazu Sakudo; Deug-chan Lee; Keiichi Saeki; Yuko Nakamura; Keiichi Inoue; Yoshitsugu Matsumoto; Shigeyoshi Itohara; Takashi Onodera

2003-01-01

301

Zinc Drives a Tertiary Fold in the Prion Protein with Familial Disease Mutation Sites at the Interface  

PubMed Central

The cellular prion protein PrPC consists of two domains – a flexible N-terminal domain, which participates in copper and zinc regulation, and a largely helical C-terminal domain that converts to ?-sheet in the course of prion disease. These two domains are thought to be fully independent and non-interacting. Compelling cellular and biophysical studies, however, suggest a higher order structure that is relevant to both PrPC function, as well as misfolding in disease. Here we identify a novel Zn2+ driven N-terminal – C-terminal tertiary interaction in PrPC. The C-terminal surface participating in this interaction carries the majority of the point mutations that confer familial prion disease. Investigation of mutant PrPs finds a systematic relationship between the type of mutation and the apparent strength of this newly identified domain structure. The novel structural features identified here suggest new mechanisms by which physiologic metal ions trigger PrPC trafficking and control prion disease. PMID:23290724

Spevacek, Ann R.; Evans, Eric G. B.; Miller, Jillian L.; Meyer, Heidi C.; Pelton, Jeffrey G.; Millhauser, Glenn L.

2012-01-01

302

How Does Domain Replacement Affect Fibril Formation of the Rabbit/Human Prion Proteins  

PubMed Central

Background It is known that in vivo human prion protein (PrP) have the tendency to form fibril deposits and are associated with infectious fatal prion diseases, while the rabbit PrP does not readily form fibrils and is unlikely to cause prion diseases. Although we have previously demonstrated that amyloid fibrils formed by the rabbit PrP and the human PrP have different secondary structures and macromolecular crowding has different effects on fibril formation of the rabbit/human PrPs, we do not know which domains of PrPs cause such differences. In this study, we have constructed two PrP chimeras, rabbit chimera and human chimera, and investigated how domain replacement affects fibril formation of the rabbit/human PrPs. Methodology/Principal Findings As revealed by thioflavin T binding assays and Sarkosyl-soluble SDS-PAGE, the presence of a strong crowding agent dramatically promotes fibril formation of both chimeras. As evidenced by circular dichroism, Fourier transform infrared spectroscopy, and proteinase K digestion assays, amyloid fibrils formed by human chimera have secondary structures and proteinase K-resistant features similar to those formed by the human PrP. However, amyloid fibrils formed by rabbit chimera have proteinase K-resistant features and secondary structures in crowded physiological environments different from those formed by the rabbit PrP, and secondary structures in dilute solutions similar to the rabbit PrP. The results from transmission electron microscopy show that macromolecular crowding caused human chimera but not rabbit chimera to form short fibrils and non-fibrillar particles. Conclusions/Significance We demonstrate for the first time that the domains beyond PrP-H2H3 (?-strand 1, ?-helix 1, and ?-strand 2) have a remarkable effect on fibrillization of the rabbit PrP but almost no effect on the human PrP. Our findings can help to explain why amyloid fibrils formed by the rabbit PrP and the human PrP have different secondary structures and why macromolecular crowding has different effects on fibrillization of PrPs from different species. PMID:25401497

Yan, Xu; Huang, Jun-Jie; Zhou, Zheng; Chen, Jie; Liang, Yi

2014-01-01

303

Highly infectious CJD particles lack prion protein but contain many viral-linked peptides by LC-MS/MS.  

PubMed

It is widely believed that host prion protein (PrP), without nucleic acid, converts itself into an infectious form (PrP-res) that causes transmissible encephalopathies (TSEs), such as human sporadic CJD (sCJD), endemic sheep scrapie, and epidemic BSE. There are many detailed investigations of PrP, but proteomic studies of other proteins in verified infectious TSE particles have not been pursued, even though brain homogenates without PrP retain their complete infectious titer. To define proteins that may be integral to, process, or protect an agent genome, we developed a streamlined, high-yield purification of infectious FU-CJD mouse brain particles with minimal PrP. Proteinase K (PK) abolished all residual particle PrP, but did not reduce infectivity, and viral-size particles lacking PrP were ?70S (vs. 90-120S without PK). Furthermore, over 1,500 non-PrP proteins were still present and positively identified in high titer FU-CJD particles without detectable PrP by mass spectrometry (LC-MS/MS); 114 of these peptides were linked to viral motifs in the environmental-viral database, and not evident in parallel uninfected controls. Host components were also identified in both PK and non-PK treated particles from FU-CJD mouse brain and human sCJD brain. This abundant cellular data had several surprises, including finding Huntingtin in the sCJD but not normal human brain samples. Similarly, the neural Wiskott-Aldrich sequence and multivesicular and endosome components associated with retromer APP (Alzheimer amyloid) processing were only in sCJD. These cellular findings suggest that new therapies directed at retromer-vesicular trafficking in other neurodegenerative diseases may also counteract late-onset sCJD PrP amyloid pathology. PMID:24933657

Kipkorir, Terry; Tittman, Sarah; Botsios, Sotirios; Manuelidis, Laura

2014-11-01

304

Hydrolysis of the amyloid prion protein and nonpathogenic meat and bone meal by anaerobic thermophilic prokaryotes and streptomyces subspecies.  

PubMed

Transmissible spongiform encephalopathies are caused by accumulation of highly resistant misfolded amyloid prion protein PrPres and can be initiated by penetration of such pathogen molecules from infected tissue to intact organism. Decontamination of animal meal containing amyloid prion protein is proposed thanks to the use of proteolytic enzymes secreted by thermophilic bacteria Thermoanaerobacter, Thermosipho, and Thermococcus subsp. and mesophilic soil bacteria Streptomyces subsp. Keratins alpha and beta, which resemble amyloid structures, were used as the substrates for the screening for microorganisms able to grow on keratins and producing efficient proteases specific for hydrolysis of beta-sheeted proteic structures, hence amyloids. Secretion of keratin-degrading proteases was evidenced by a zymogram method. Enzymes from thermophilic strains VC13, VC15, and S290 and Streptomyces subsp. S6 were strongly active against amyloid recombinant ovine prion protein and animal meal proteins. The studied proteases displayed broad primary specificities hydrolyzing low molecular mass peptide model substrates. Strong amyloidolytic activity of detected proteases was confirmed by experiments of hydrolysis of PrPres in SAFs produced from brain homogenates of mice infected with the 6PB1 BSE strain. The proteases from Thermoanaerobacter subsp. S290 and Streptomyces subsp. S6 are the best candidates for neutralization/elimination of amyloids in meat and bone meal and other protein-containing substances and materials. PMID:15453713

Tsiroulnikov, Kirill; Rezai, Human; Bonch-Osmolovskaya, Elisaveta; Nedkov, Peter; Gousterova, Adriana; Cueff, Valérie; Godfroy, Anne; Barbier, Georges; Métro, François; Chobert, Jean-Marc; Clayette, Pascal; Dormont, Dominique; Grosclaude, Jeanne; Haertlé, Thomas

2004-10-01

305

Molecular population genetics and evolution of a prion-like protein in Saccharomyces cerevisiae.  

PubMed Central

The prion-like behavior of Sup35p, the eRF3 homolog in the yeast Saccharomyces cerevisiae, mediates the activity of the cytoplasmic nonsense suppressor known as [PSI(+)]. Sup35p is divided into three regions of distinct function. The N-terminal and middle (M) regions are required for the induction and propagation of [PSI(+)] but are not necessary for translation termination or cell viability. The C-terminal region encompasses the termination function. The existence of the N-terminal region in SUP35 homologs of other fungi has led some to suggest that this region has an adaptive function separate from translation termination. To examine this hypothesis, we sequenced portions of SUP35 in 21 strains of S. cerevisiae, including 13 clinical isolates. We analyzed nucleotide polymorphism within this species and compared it to sequence divergence from a sister species, S. paradoxus. The N domain of Sup35p is highly conserved in amino acid sequence and is highly biased in codon usage toward preferred codons. Amino acid changes are under weak purifying selection based on a quantitative analysis of polymorphism and divergence. We also conclude that the clinical strains of S. cerevisiae are not recently derived and that outcrossing between strains in S. cerevisiae may be relatively rare in nature. PMID:11606530

Jensen, M A; True, H L; Chernoff, Y O; Lindquist, S

2001-01-01

306

Reversible Conversion of Monomeric Human Prion  

E-print Network

Reversible Conversion of Monomeric Human Prion Protein Between Native and Fibrilogenic J. P. Waltho,2 A. R. Clarke,1,4 J. Collinge1 * Prion propagation involves the conversion of cellular prion protein (PrPC ) into a disease-specific isomer, PrPSc , shifting from a predominantly -helical

Hosszu, Laszlo

307

Direct observation of multiple misfolding pathways in a single prion protein molecule  

PubMed Central

Protein misfolding is a ubiquitous phenomenon associated with a wide range of diseases. Single-molecule approaches offer a powerful tool for deciphering the mechanisms of misfolding by measuring the conformational fluctuations of a protein with high sensitivity. We applied single-molecule force spectroscopy to observe directly the misfolding of the prion protein PrP, a protein notable for having an infectious misfolded state that is able to propagate by recruiting natively folded PrP. By measuring folding trajectories of single PrP molecules held under tension in a high-resolution optical trap, we found that the native folding pathway involves only two states, without evidence for partially folded intermediates that have been proposed to mediate misfolding. Instead, frequent but fleeting transitions were observed into off-pathway intermediates. Three different misfolding pathways were detected, all starting from the unfolded state. Remarkably, the misfolding rate was even higher than the rate for native folding. A mutant PrP with higher aggregation propensity showed increased occupancy of some of the misfolded states, suggesting these states may act as intermediates during aggregation. These measurements of individual misfolding trajectories demonstrate the power of single-molecule approaches for characterizing misfolding directly by mapping out nonnative folding pathways. PMID:22421432

Yu, Hao; Liu, Xia; Neupane, Krishna; Gupta, Amar Nath; Brigley, Angela M.; Solanki, Allison; Sosova, Iveta; Woodside, Michael T.

2012-01-01

308

Melatonin-mediated ?-catenin activation protects neuron cells against prion protein-induced neurotoxicity.  

PubMed

Activation of ?-catenin in neurons regulates mitochondrial function and protects against protein misfolding disorders, including Alzheimer's disease and Huntington's disease. Melatonin, a natural secretory product of the pineal gland, exerts neuroprotective effects through the activation of ?-catenin. In this study, melatonin increased ?-catenin protein expression and activation in human neuroblastoma cell lines SH-SY5Y cells. Melatonin also inhibited PrP (106-126)-induced neurotoxicity and the inhibition attenuated by treatment of ?-catenin inhibitor ICG-001. Activation of ?-catenin blocked PrP (106-126)-mediated downregulation of anti-apoptotic protein survivin and Bcl-2. Reduction of mitochondrial membrane potential, translocation of Bax, and cytochrome c release which induced by PrP (106-126) treatment were inhibited by ?-catenin activation, which contributed to prevented PrP (106-126)-induced neuronal cell death. In conclusion, ?-catenin activation by melatonin prevented PrP (106-126)-induced neuronal cell death through regulating anti-apoptotic proteins and mitochondrial pathways. These results also suggest the therapeutic value of Wnt/?-catenin signaling in prion-related disorders as influenced by melatonin. PMID:25251028

Jeong, Jae-Kyo; Lee, Ju-Hee; Moon, Ji-Hong; Lee, You-Jin; Park, Sang-Youel

2014-11-01

309

Processing and Degradation of Exogenous Prion Protein by CD11c+ Myeloid Dendritic Cells In Vitro  

Microsoft Academic Search

The immune system plays an important role in facilitating the spread of prion infections from the periphery to the central nervous system. CD11c myeloid dendritic cells (DC) could, due to their subepithelial location and their migratory capacity, be early targets for prion infection and contribute to the spread of infection. In order to analyze mechanisms by which these cells may

Katarina M. Luhr; Robert P. A. Wallin; Hans-Gustaf Ljunggren; Peter Low; Albert Taraboulos; Krister Kristensson

2002-01-01

310

Insight into Molecular Basis of Curing of [PSI+] Prion by Overexpression of 104-kDa Heat Shock Protein (Hsp104)*  

PubMed Central

Yeast prions are a powerful model for understanding the dynamics of protein aggregation associated with a number of human neurodegenerative disorders. The AAA+ protein disaggregase Hsp104 can sever the amyloid fibrils produced by yeast prions. This action results in the propagation of “seeds” that are transmitted to daughter cells during budding. Overexpression of Hsp104 eliminates the [PSI+] prion but not other prions. Using biochemical methods we identified Hsp104 binding sites in the highly charged middle domain of Sup35, the protein determinant of [PSI+]. Deletion of a short segment of the middle domain (amino acids 129–148) diminishes Hsp104 binding and strongly affects the ability of the middle domain to stimulate the ATPase activity of Hsp104. In yeast, [PSI+] maintained by Sup35 lacking this segment, like other prions, is propagated by Hsp104 but cannot be cured by Hsp104 overexpression. These results provide new insight into the enigmatic specificity of Hsp104-mediated curing of yeast prions and sheds light on the limitations of the ability of Hsp104 to eliminate aggregates produced by other aggregation-prone proteins. PMID:22081611

Helsen, Christopher W.; Glover, John R.

2012-01-01

311

Prion Protein-Deficient Cells Show Altered Response to Oxidative Stress Due to Decreased SOD1 Activity  

Microsoft Academic Search

The cellular function of the prion protein (PrPC), a cell surface glycoprotein expressed in neurones and astrocytes, has not been elucidated. Cell culture experiments reveal that cerebellar cells lacking PrPCare more sensitive to oxidative stress and undergo cell death more readily than wild-type cells. This effect is reversible by treatment with vitamin E.In vivostudies show that the activity of Cu\\/Zn

David R. Brown; Walter J. Schulz-Schaeffer; Bernhard Schmidt; Hans A. Kretzschmar

1997-01-01

312

Different allelic effects of the codons 136 and 171 of the prion protein gene in sheep with natural scrapie  

Microsoft Academic Search

Scrapie is a transmissible degenerative disease of the central nervous system occurring naturally in sheep. It belongs to the group ofprion diseases also affecting man in which an abnormal isoform of the host-encoded prion protein (PrP) accumulating in the brain is responsible for neuronal death. Three main polymorphisms have been described in the sheep PrP gene, at positions 136, 154

C. Clouscard; P. Beaudry; J. M. Elsen; D. Milan; M. Dussaucy; C. Bounneau; F. Schelcher; J. Chatelain; J. M. Launay; J. L. Laplanche

1995-01-01

313

Introducing a Rigid Loop Structure from Deer into Mouse Prion Protein Increases Its Propensity for Misfolding In Vitro  

PubMed Central

Prion diseases are fatal neurodegenerative disorders characterized by misfolding of the cellular prion protein (PrPc) into the disease-associated isoform (PrPSc) that has increased ?-sheet content and partial resistance to proteolytic digestion. Prion diseases from different mammalian species have varying propensities for transmission upon exposure of an uninfected host to the infectious agent. Chronic Wasting Disease (CWD) is a highly transmissible prion disease that affects free ranging and farmed populations of cervids including deer, elk and moose, as well as other mammals in experimental settings. The molecular mechanisms allowing CWD to maintain comparatively high transmission rates have not been determined. Previous work has identified a unique structural feature in cervid PrP, a rigid loop between ?-sheet 2 and ?-helix 2 on the surface of the protein. This study was designed to test the hypothesis that the rigid loop has a direct influence on the misfolding process. The rigid loop was introduced into murine PrP as the result of two amino acid substitutions: S170N and N174T. Wild-type and rigid loop murine PrP were expressed in E. coli and purified. Misfolding propensity was compared for the two proteins using biochemical techniques and cell free misfolding and conversion systems. Murine PrP with a rigid loop misfolded in cell free systems with greater propensity than wild type murine PrP. In a lipid-based conversion assay, rigid loop PrP converted to a PK resistant, aggregated isoform at lower concentrations than wild-type PrP. Using both proteins as substrates in real time quaking-induced conversion, rigid loop PrP adopted a misfolded isoform more readily than wild type PrP. Taken together, these findings may help explain the high transmission rates observed for CWD within cervids. PMID:23825561

Kyle, Leah M.; John, Theodore R.; Schatzl, Hermann M.; Lewis, Randolph V.

2013-01-01

314

The 129 codon polymorphism of the Prion Protein gene influences earlier cognitive performance in Down syndrome subjects  

Microsoft Academic Search

.   Recently, a frequent prion protein gene (PRNP) polymorphism consisting of a methionine (M) for valine (V) substitution at\\u000a codon 129 has been associated with cognitive impairment in elderly individuals. Down syndrome (DS) is associated with mental\\u000a retardation and development of Alzheimer-like brain abnormalities. In the present study, we investigated the role of the PRNP\\u000a polymorphism in 122 relatively young

Roberto Del Bo; Giacomo Pietro Comi; Roberto Giorda; Marco Crimi; Federica Locatelli; Filippo Martinelli-Boneschi; Uberto Pozzoli; Enrico Castelli; Nereo Bresolin; Guglielmo Scarlato

2003-01-01

315

Strain specific resistance to murine scrapie associated with a naturally occurring human prion protein polymorphism at residue 171.  

PubMed

Transmissible spongiform encephalopathies (TSE) or prion diseases are neurodegenerative disorders associated with conversion of normal host prion protein (PrP) to a misfolded, protease-resistant form (PrPres). Genetic variations of prion protein in humans and animals can alter susceptibility to both familial and infectious prion diseases. The N171S PrP polymorphism is found mainly in humans of African descent, but its low incidence has precluded study of its possible influence on prion disease. Similar to previous experiments of others, for laboratory studies we created a transgenic model expressing the mouse PrP homolog, PrP-170S, of human PrP-171S. Since PrP polymorphisms can vary in their effects on different TSE diseases, we tested these mice with four different strains of mouse-adapted scrapie. Whereas 22L and ME7 scrapie strains induced typical clinical disease, neuropathology and accumulation of PrPres in all transgenic mice at 99-128 average days post-inoculation, strains RML and 79A produced clinical disease and PrPres formation in only a small subset of mice at very late times. When mice expressing both PrP-170S and PrP-170N were inoculated with RML scrapie, dominant-negative inhibition of disease did not occur, possibly because interaction of strain RML with PrP-170S was minimal. Surprisingly, in vitro PrP conversion using protein misfolding cyclic amplification (PMCA), did not reproduce the in vivo findings, suggesting that the resistance noted in live mice might be due to factors or conditions not present in vitro. These findings suggest that in vivo conversion of PrP-170S by RML and 79A scrapie strains was slow and inefficient. PrP-170S mice may be an example of the conformational selection model where the structure of some prion strains does not favor interactions with PrP molecules expressing certain polymorphisms. PMID:21980292

Striebel, James F; Race, Brent; Meade-White, Kimberly D; LaCasse, Rachel; Chesebro, Bruce

2011-09-01

316

Strain Specific Resistance to Murine Scrapie Associated with a Naturally Occurring Human Prion Protein Polymorphism at Residue 171  

PubMed Central

Transmissible spongiform encephalopathies (TSE) or prion diseases are neurodegenerative disorders associated with conversion of normal host prion protein (PrP) to a misfolded, protease-resistant form (PrPres). Genetic variations of prion protein in humans and animals can alter susceptibility to both familial and infectious prion diseases. The N171S PrP polymorphism is found mainly in humans of African descent, but its low incidence has precluded study of its possible influence on prion disease. Similar to previous experiments of others, for laboratory studies we created a transgenic model expressing the mouse PrP homolog, PrP-170S, of human PrP-171S. Since PrP polymorphisms can vary in their effects on different TSE diseases, we tested these mice with four different strains of mouse-adapted scrapie. Whereas 22L and ME7 scrapie strains induced typical clinical disease, neuropathology and accumulation of PrPres in all transgenic mice at 99-128 average days post-inoculation, strains RML and 79A produced clinical disease and PrPres formation in only a small subset of mice at very late times. When mice expressing both PrP-170S and PrP-170N were inoculated with RML scrapie, dominant-negative inhibition of disease did not occur, possibly because interaction of strain RML with PrP-170S was minimal. Surprisingly, in vitro PrP conversion using protein misfolding cyclic amplification (PMCA), did not reproduce the in vivo findings, suggesting that the resistance noted in live mice might be due to factors or conditions not present in vitro. These findings suggest that in vivo conversion of PrP-170S by RML and 79A scrapie strains was slow and inefficient. PrP-170S mice may be an example of the conformational selection model where the structure of some prion strains does not favor interactions with PrP molecules expressing certain polymorphisms. PMID:21980292

Striebel, James F.; Race, Brent; Meade-White, Kimberly D.; LaCasse, Rachel; Chesebro, Bruce

2011-01-01

317

A Novel, Drug-based, Cellular Assay for the Activity of Neurotoxic Mutants of the Prion Protein*  

PubMed Central

In prion diseases, the infectious isoform of the prion protein (PrPSc) may subvert a normal, physiological activity of the cellular isoform (PrPC). A deletion mutant of the prion protein (?105–125) that produces a neonatal lethal phenotype when expressed in transgenic mice provides a window into the normal function of PrPC and how it can be corrupted to produce neurotoxic effects. We report here the surprising and unexpected observation that cells expressing ?105–125 PrP and related mutants are hypersensitive to the toxic effects of two classes of antibiotics (aminoglycosides and bleomycin analogues) that are commonly used for selection of stably transfected cell lines. This unusual phenomenon mimics several essential features of ?105–125 PrP toxicity seen in transgenic mice, including rescue by co-expression of wild type PrP. Cells expressing ?105–125 PrP are susceptible to drug toxicity within minutes, suggesting that the mutant protein enhances cellular accumulation of these cationic compounds. Our results establish a screenable cellular phenotype for the activity of neurotoxic forms of PrP, and they suggest possible mechanisms by which these molecules could produce their pathological effects in vivo. PMID:19940127

Massignan, Tania; Stewart, Richard S.; Biasini, Emiliano; Solomon, Isaac H.; Bonetto, Valentina; Chiesa, Roberto; Harris, David A.

2010-01-01

318

Phenotypic characterization of cells participating in transport of prion protein aggregates across the intestinal mucosa of sheep  

PubMed Central

The oral route is considered to be the main entry site of several transmissible spongiform encephalopathies or prion diseases of animals and man. Following natural and experimental oral exposure to scrapie, sheep first accumulate disease associated prion protein (PrPd) in Peyer’s patch (PP) lymphoid follicles. In this study, recombinant ovine prion protein (rPrP) was inoculated into gut loops of young lambs and the transportation across the intestinal wall studied. In particular, the immunohistochemical phenotypes of cells bearing the inoculated prion protein were investigated. The rPrP was shown to be transported across the villi of the gut, into the lacteals and submucosal lymphatics, mimicking the transport route of PrPd from scrapie brain inoculum observed in a previous intestinal loop experiment. The cells bearing the inoculated rPrP were mainly mononuclear cells, and multicolor immunofluorescence procedures were used to show that the rPrP bearing cells were professional antigen presenting cells expressing Major histocompatibility complex II (MHCII). In addition, the rPrP bearing cells labeled with CD205, CD11b and the macrophage marker CD68, and not with the dendritic cell markers CD11c and CD209. Others have reported that cells expressing CD205 and CD11b in the absence of CD11c have been shown to induce T cell tolerance or regulatory T cells. Based on this association, it was speculated that the rPrP and by extension PrPd and scrapie infective material may exploit the physiological process of macromolecular uptake across the gut, and that this route of entry may have implications for immune surveillance. PMID:22437736

Piercey Akesson, Caroline; Press, Charles McL.; Tranulis, Michael A.; Jeffrey, Martin; Aleksandersen, Mona; Landsverk, Thor; Espenes, Arild

2012-01-01

319

Prions and Prion-Like Pathogens in Neurodegenerative Disorders  

PubMed Central

Prions are unique elements in biology, being able to transmit biological information from one organism to another in the absence of nucleic acids. They have been identified as self-replicating proteinaceous agents responsible for the onset of rare and fatal neurodegenerative disorders—known as transmissible spongiform encephalopathies, or prion diseases—which affect humans and other animal species. More recently, it has been proposed that other proteins associated with common neurodegenerative disorders, such as Alzheimer’s and Parkinson’s disease, can self-replicate like prions, thus sustaining the spread of neurotoxic entities throughout the nervous system. Here, we review findings that have contributed to expand the prion concept, and discuss if the involved toxic species can be considered bona fide prions, including the capacity to infect other organisms, or whether these pathogenic aggregates share with prions only the capability to self-replicate.

Peggion, Caterina; Sorgato, Maria Catia; Bertoli, Alessandro

2014-01-01

320

Circumventing Tolerance to the Prion Protein (PrP): Vaccination with PrP-Displaying Retrovirus Particles Induces Humoral Immune Responses against the Native Form of Cellular PrP  

Microsoft Academic Search

Passive immunization with antibodies directed against the cellular form of the prion protein (PrPC) can protect against prion disease. However, active immunization with recombinant prion protein has so far failed to induce antibodies directed against native PrPC expressed on the cell surface. To develop an antiprion vac- cine, a retroviral display system presenting either the full-length mouse PrP (PrP209) or

Daphne Nikles; Patricia Bach; Klaus Boller; Christoph A. Merten; Fabio Montrasio; Frank L. Heppner; Adriano Aguzzi; Klaus Cichutek; Ulrich Kalinke; Christian J. Buchholz

2005-01-01

321

Metabotropic glutamate receptor 5 is a coreceptor for Alzheimer a? oligomer bound to cellular prion protein.  

PubMed

Soluble amyloid-? oligomers (A?o) trigger Alzheimer's disease (AD) pathophysiology and bind with high affinity to cellular prion protein (PrP(C)). At the postsynaptic density (PSD), extracellular A?o bound to lipid-anchored PrP(C) activates intracellular Fyn kinase to disrupt synapses. Here, we screened transmembrane PSD proteins heterologously for the ability to couple A?o-PrP(C) with Fyn. Only coexpression of the metabotropic glutamate receptor, mGluR5, allowed PrP(C)-bound A?o to activate Fyn. PrP(C) and mGluR5 interact physically, and cytoplasmic Fyn forms a complex with mGluR5. A?o-PrP(C) generates mGluR5-mediated increases of intracellular calcium in Xenopus oocytes and in neurons, and the latter is also driven by human AD brain extracts. In addition, signaling by A?o-PrP(C)-mGluR5 complexes mediates eEF2 phosphorylation and dendritic spine loss. For mice expressing familial AD transgenes, mGluR5 antagonism reverses deficits in learning, memory, and synapse density. Thus, A?o-PrP(C) complexes at the neuronal surface activate mGluR5 to disrupt neuronal function. PMID:24012003

Um, Ji Won; Kaufman, Adam C; Kostylev, Mikhail; Heiss, Jacqueline K; Stagi, Massimiliano; Takahashi, Hideyuki; Kerrisk, Meghan E; Vortmeyer, Alexander; Wisniewski, Thomas; Koleske, Anthony J; Gunther, Erik C; Nygaard, Haakon B; Strittmatter, Stephen M

2013-09-01

322

MRS in Early and Presymptomatic Carriers of a Novel Octapeptide Repeat Insertion in the Prion Protein Gene  

PubMed Central

To evaluate the proton MR spectroscopy (1H MRS) changes in carriers of a novel octapeptide repeat insertion in the Prion Protein Gene (PRNP) and family history of frontotemporal dementia with ataxia. Four at-risk mutation carriers and 13 controls were compared using single voxel, short TE, 1H MRS from the posterior cingulate gyrus. The mutation carriers had an increased choline/creatine, p=0.003 and increased myoinositol/creatine ratio, p=0.003. 1H MRS identified differences in markers of glial activity and choline metabolism in pre- and early symptomatic carriers of a novel PRNP gene octapeptide insertion. These findings expand the possible diagnostic utility of 1H MRS in familial prion disorders. PMID:22612156

McDade, Eric M; Boeve, Bradley F.; Fields, Julie A; Kumar, Neeraj; Rademakers, Rosa; Baker, Matt C.; Knopman, David S.; Petersen, Ronald C.; Jack, Clifford R.; Kantarci, Kejal

2012-01-01

323

MRS in early and presymptomatic carriers of a novel octapeptide repeat insertion in the prion protein gene.  

PubMed

To evaluate the proton magnetic resonance (MR) spectroscopy ((1) H MRS) changes in carriers of a novel octapeptide repeat insertion in the prion protein gene (PRNP) and family history of frontotemporal dementia with ataxia. Four at-risk mutation carriers and 13 controls were compared using single voxel, short TE, (1) H MRS from the posterior cingulate gyrus. The mutation carriers had an increased choline/creatine, P = .003 and increased myoinositol/creatine ratio, P = .003. (1) H MRS identified differences in markers of glial activity and choline metabolism in pre- and early-symptomatic carriers of a novel PRNP gene octapeptide insertion. These findings expand the possible diagnostic utility of (1) H MRS in familial prion disorders. PMID:22612156

McDade, Eric M; Boeve, Bradley F; Fields, Julie A; Kumar, Neeraj; Rademakers, Rosa; Baker, Matt C; Knopman, Bsc David S; Petersen, Ronald C; Jack, Clifford R; Kantarci, Kejal

2013-07-01

324

Redox behaviors of the neurotoxic portion in human prion protein, HuPrP(106-126)  

NASA Astrophysics Data System (ADS)

A peptide fragment of human prion protein, HuPrP(106-126), has been reported to mimic the pathological features underlying prion diseases. Although the actual neurotoxic mechanism of HuPrP(106-126) has not been elucidated, several hypotheses has been proposed based on the role for copper. In this study, to understand the toxic function of HuPrP(106-126) from a viewpoint of electrochemical competence, we investigated redox properties of copper ion complexes with four different binding motifs of a model of HuPrP(106-126) based on density functional theory calculations. We found that the HuPrP(106-126)-derived models exhibited diverse redox activities that depended on copper-binding conformations.

Yamamoto, Norifumi; Kuwata, Kazuo

2010-09-01

325

Double replacement gene targeting for the production of a series of mouse strains with different prion protein gene alterations  

SciTech Connect

We have developed a double replacement gene targeting strategy which enables the production of a series of mouse strains bearing different subtle alterations to endogenous genes. This is a two-step process in which a region of the gene of interest is first replaced with a selectable marker to produce an inactivated allele, which is then re-targeted with a second vector to reconstruct the inactivated allele, concomitantly introducing an engineered mutation. Five independent embryonic stem cell lines have been produced bearing different targeted alterations to the prion protein gene, including one which raises the level of expression. We have constructed mice bearing the codon 101 proline to leucine substitution linked to the human familial prion disease, Gerstmann-Straussler-Scheinker syndrome. We anticipate that this procedure will have applications to the study of human inherited diseases and the development of therapies. 43 refs., 6 figs., 1 tab.

Moore, R.C.; Redhead, N.J.; Selfridge, J. [Univ. of Edinburgh (United Kingdom)] [and others] [Univ. of Edinburgh (United Kingdom); and others

1995-09-01

326

Original article Prion gene (PRNP) haplotype variation  

E-print Network

Original article Prion gene (PRNP) haplotype variation in United States goat breeds (Open Access not examined before at PRNP. scrapie / goat / polymorphism / resistance / prion 1. INTRODUCTION Scrapie that encodes prion protein have been associated with differential resistance and susceptibility to scrapie

Boyer, Edmond

327

Prion strain discrimination using luminescent conjugated polymers  

E-print Network

Prion strain discrimination using luminescent conjugated polymers Christina J Sigurdson1,6, K Peter of prions may reflect conformational variability of PrPSc, a disease-associated, aggregated variant of the cellular prion protein, PrPC. Here we used luminescent conjugated polymers (LCPs), which emit conformation

Cai, Long

328

Sequence Requirements of Spider Silk Elastic Proteins.  

National Technical Information Service (NTIS)

Sequence for flagelliform silk proteins from Nephila, Argiope, and Araneus species have been obtained. These sequences show significant sequence differences although the changes are not predicted to affect the secondary structure of the protein and theref...

R. V. Lewis

2001-01-01

329

Coexpression of wild-type and mutant prion proteins alters their cellular localization and partitioning into detergent-resistant membranes.  

PubMed

Transmissible spongiform encephalopathies (TSEs) are a group of diseases of infectious, sporadic and genetic origin, found in higher organisms and caused by the pathological form of the prion protein. The inheritable subgroup of TSEs is linked to insertional or point mutations in the prion gene prnp, which favour its misfolding and are passed on to offspring in an autosomal-dominant fashion. The large majority of patients with these diseases are heterozygous for the prnp gene, leading to the coexpression of the wild-type (wt) (PrP(C)) and the mutant forms (PrPmut) in the carriers of these mutations. To mimic this situation in vitro, we produced Fischer rat thyroid cells coexpressing PrPwt alongside mutant versions of mouse PrP including A117V, E200K and T182A relevant to the human TSE diseases Gestmann-Sträussler-Scheinker (GSS) disease and familial Creutzfeldt-Jakob disease (fCJD). We found that coexpression of mutant PrP with wt proteins does not affect the glycosylation pattern or the biochemical characteristics of either protein. However, FRET and co-immunoprecipitation experiments suggest an interaction occurring between the wt and mutant proteins. Furthermore, by comparing the intracellular localization and detergent-resistant membrane (DRM) association in single- and double-expressing clones, we found changes in the intracellular/surface ratio and an increased sequestration of both proteins in DRMs, a site believed to be involved in the pathological conversion (or protection thereof) of the prion protein. We, therefore, propose that the mutant forms alter the subcellular localization and the membrane environment of the wt protein in co-transfected cells. These effects may play a role in the development of these diseases. PMID:18410485

Schiff, Edwin; Campana, Vincenza; Tivodar, Simona; Lebreton, Stéphanie; Gousset, Karine; Zurzolo, Chiara

2008-07-01

330

Dynamically-expressed prion-like proteins form a cuticle in the pharynx of Caenorhabditis elegans.  

PubMed

In molting animals, a cuticular extracellular matrix forms the first barrier to infection and other environmental insults. In the nematode Caenorhabditis elegans there are two types of cuticle: a well-studied collagenous cuticle lines the body, and a poorly-understood chitinous cuticle lines the pharynx. In the posterior end of the pharynx is the grinder, a tooth-like cuticular specialization that crushes food prior to transport to the intestine for digestion. We here show that the grinder increases in size only during the molt. To gain molecular insight into the structure of the grinder and pharyngeal cuticle, we performed a microarray analysis to identify mRNAs increased during the molt. We found strong transcriptional induction during the molt of 12 of 15 previously identified abu genes encoding Prion-like (P) glutamine (Q) and asparagine (N) rich PQN proteins, as well as 15 additional genes encoding closely related PQN proteins. abu/pqn genes, which we name the abu/pqn paralog group (APPG) genes, were expressed in pharyngeal cells and the proteins encoded by two APPG genes we tested localized to the pharyngeal cuticle. Deleting the APPG gene abu-14 caused abnormal pharyngeal cuticular structures and knocking down other APPG genes resulted in abnormal cuticular function. We propose that APPG proteins promote the assembly and function of a unique cuticular structure. The strong developmental regulation of the APPG genes raises the possibility that such genes would be identified in transcriptional profiling experiments in which the animals' developmental stage is not precisely staged. PMID:25361578

George-Raizen, Julia B; Shockley, Keith R; Trojanowski, Nicholas F; Lamb, Annesia L; Raizen, David M

2014-01-01

331

Dynamically-expressed prion-like proteins form a cuticle in the pharynx of Caenorhabditis elegans  

PubMed Central

ABSTRACT In molting animals, a cuticular extracellular matrix forms the first barrier to infection and other environmental insults. In the nematode Caenorhabditis elegans there are two types of cuticle: a well-studied collagenous cuticle lines the body, and a poorly-understood chitinous cuticle lines the pharynx. In the posterior end of the pharynx is the grinder, a tooth-like cuticular specialization that crushes food prior to transport to the intestine for digestion. We here show that the grinder increases in size only during the molt. To gain molecular insight into the structure of the grinder and pharyngeal cuticle, we performed a microarray analysis to identify mRNAs increased during the molt. We found strong transcriptional induction during the molt of 12 of 15 previously identified abu genes encoding Prion-like (P) glutamine (Q) and asparagine (N) rich PQN proteins, as well as 15 additional genes encoding closely related PQN proteins. abu/pqn genes, which we name the abu/pqn paralog group (APPG) genes, were expressed in pharyngeal cells and the proteins encoded by two APPG genes we tested localized to the pharyngeal cuticle. Deleting the APPG gene abu-14 caused abnormal pharyngeal cuticular structures and knocking down other APPG genes resulted in abnormal cuticular function. We propose that APPG proteins promote the assembly and function of a unique cuticular structure. The strong developmental regulation of the APPG genes raises the possibility that such genes would be identified in transcriptional profiling experiments in which the animals' developmental stage is not precisely staged. PMID:25361578

George-Raizen, Julia B.; Shockley, Keith R.; Trojanowski, Nicholas F.; Lamb, Annesia L.; Raizen, David M.

2014-01-01

332

A Prion Protein Fragment Primes Type 1 Astrocytes to Proliferation Signals from Microglia  

Microsoft Academic Search

Gliosis is a hallmark of prion disease. A neurotoxic prion peptide (PrP106-126) induces astrocyte proliferation in the presence of microglia. This peptide also directly enhances microglial proliferation in culture. We have investigated this further to understand the method by which factors released by microglia and PrP106-126 work together to enhance astrocyte proliferation. PrP106-126 in the presence of microglia specifically enhanced

David R. Brown; Bernhard Schmidt; Hans A. Kretzschmar

1998-01-01

333

Prion Problem Space  

NSDL National Science Digital Library

This problem space introduces basic skills in protein structure exploration, utilizing prions -- relatively small proteins that display dramatically alternate conformations for similar primary structures. We will learn to search databases for protein structures, explore the Cn3D software, and propose questions that may be answered with these tools.

Stephen Everse (University of Vermont College of Medicine;Biochemistry)

2005-12-16

334

Btn3 is a negative regulator of Btn2-mediated endosomal protein trafficking and prion curing in yeast  

PubMed Central

Yeast Btn2 facilitates the retrieval of specific proteins from late endosomes (LEs) to the Golgi, a process that may be adversely affected in Batten disease patients. We isolated the putative yeast orthologue of a human complex I deficiency gene, designated here as BTN3, as encoding a Btn2-interacting protein and negative regulator. First, yeast overexpressing BTN3 phenocopy the deletion of BTN2 and mislocalize certain trans-Golgi proteins, like Kex2 and Yif1, to the LE and vacuole, respectively. In contrast, the deletion of BTN3 results in a tighter pattern of protein localization to the Golgi. Second, BTN3 overexpression alters Btn2 localization from the IPOD compartment, which correlates with a sharp reduction in Btn2-mediated [URE3] prion curing. Third, Btn3 and the Snc1 v-SNARE compete for the same binding domain on Btn2, and this competition controls Btn2 localization and function. The inhibitory effects upon protein retrieval and prion curing suggest that Btn3 sequesters Btn2 away from its substrates, thus down-regulating protein trafficking and aggregation. Therefore Btn3 is a novel negative regulator of intracellular protein sorting, which may be of importance in the onset of complex I deficiency and Batten disease in humans. PMID:21441304

Kanneganti, Vydehi; Kama, Rachel; Gerst, Jeffrey E.

2011-01-01

335

Evidence for Retrogene Origins of the Prion Gene Family  

PubMed Central

The evolutionary origin of prion genes, only known to exist in the vertebrate lineage, had remained elusive until recently. Following a lead from interactome investigations of the murine prion protein, our previous bioinformatic analyses revealed the evolutionary descent of prion genes from an ancestral ZIP metal ion transporter. However, the molecular mechanism of evolution remained unexplored. Here we present a computational investigation of this question based on sequence, intron-exon, synteny and pseudogene analyses. Our data suggest that during the emergence of metazoa, a cysteine-flanked core domain was modularly inserted, or arose de novo, in a preexisting ZIP ancestor gene to generate a prion-like ectodomain in a subbranch of ZIP genes. Approximately a half-billion years later, a genomic insertion of a spliced transcript coding for such a prion-like ZIP ectodomain may have created the prion founder gene. We document that similar genomic insertions involving ZIP transcripts, and probably relying on retropositional elements, have indeed occurred more than once throughout evolution. PMID:22046361

Ehsani, Sepehr; Tao, Renzhu; Pocanschi, Cosmin L.; Ren, Hezhen; Harrison, Paul M.; Schmitt-Ulms, Gerold

2011-01-01

336

Loss of prion protein leads to age-dependent behavioral abnormalities and changes in cytoskeletal protein expression.  

PubMed

The cellular prion protein (PrPC) is a highly conserved protein whose exact physiological role remains elusive. In the present study, we investigated age-dependent behavioral abnormalities in PrPC-knockout (Prnp0/0) mice and wild-type (WT) controls. Prnp0/0 mice showed age-dependent behavioral deficits in memory performance, associative learning, basal anxiety, and nest building behavior. Using a hypothesis-free quantitative proteomic investigation, we found that loss of PrPC affected the levels of neurofilament proteins in an age-dependent manner. In order to understand the biochemical basis of these observations, we analyzed the phosphorylation status of neurofilament heavy chain (NF-H). We found a reduction in NF-H phosphorylation in both Prnp0/0 mice and in PrPC-deficient cells. The expression of Fyn and phospho-Fyn, a potential regulator for NF phosphorylation, was associated with PrPC ablation. The number of ?-tubulin III-positive neurons in the hippocampus was diminished in Prnp0/0 mice relative to WT mice. These data indicate that PrPC plays an important role in cytoskeletal organization, brain function, and age-related neuroprotection. Our work represents the first direct biochemical link between these proteins and the observed behavioral phenotypes. PMID:24604355

Schmitz, Matthias; Greis, Catharina; Ottis, Philipp; Silva, Christopher J; Schulz-Schaeffer, Walter J; Wrede, Arne; Koppe, Katharina; Onisko, Bruce; Requena, Jesús R; Govindarajan, Nambirajan; Korth, Carsten; Fischer, Andre; Zerr, Inga

2014-12-01

337

Recruitment of cellular prion protein to mitochondrial raft-like microdomains contributes to apoptosis execution  

PubMed Central

We examined the possibility that cellular prion protein (PrPC) plays a role in the receptor-mediated apoptotic pathway. We first found that CD95/Fas triggering induced a redistribution of PrPC to the mitochondria of T lymphoblastoid CEM cells via a mechanism that brings into play microtubular network integrity and function. In particular, we demonstrated that PrPC was redistributed to raft-like microdomains at the mitochondrial membrane, as well as at endoplasmic reticulum-mitochondria–associated membranes. Our in vitro experiments also demonstrated that, although PrPC had such an effect on mitochondria, it induced the loss of mitochondrial membrane potential and cytochrome c release only after a contained rise of calcium concentration. Finally, the involvement of PrPC in apoptosis execution was also analyzed in PrPC-small interfering RNA–transfected cells, which were found to be significantly less susceptible to CD95/Fas–induced apoptosis. Taken together, these results suggest that PrPC might play a role in the complex multimolecular signaling associated with CD95/Fas receptor–mediated apoptosis. PMID:22031292

Mattei, Vincenzo; Matarrese, Paola; Garofalo, Tina; Tinari, Antonella; Gambardella, Lucrezia; Ciarlo, Laura; Manganelli, Valeria; Tasciotti, Vincenzo; Misasi, Roberta; Malorni, Walter; Sorice, Maurizio

2011-01-01

338

Solution Structure and Dynamics of the I214V Mutant of the Rabbit Prion Protein  

PubMed Central

Background The conformational conversion of the host-derived cellular prion protein (PrPC) into the disease-associated scrapie isoform (PrPSc) is responsible for the pathogenesis of transmissible spongiform encephalopathies (TSEs). Various single-point mutations in PrPCs could cause structural changes and thereby distinctly influence the conformational conversion. Elucidation of the differences between the wild-type rabbit PrPC (RaPrPC) and various mutants would be of great help to understand the ability of RaPrPC to be resistant to TSE agents. Methodology/Principal Findings We determined the solution structure of the I214V mutant of RaPrPC(91–228) and detected the backbone dynamics of its structured C-terminal domain (121–228). The I214V mutant displays a visible shift of surface charge distribution that may have a potential effect on the binding specificity and affinity with other chaperones. The number of hydrogen bonds declines dramatically. Urea-induced transition experiments reveal an obvious decrease in the conformational stability. Furthermore, the NMR dynamics analysis discloses a significant increase in the backbone flexibility on the pico- to nanosecond time scale, indicative of lower energy barrier for structural rearrangement. Conclusions/Significance Our results suggest that both the surface charge distribution and the intrinsic backbone flexibility greatly contribute to species barriers for the transmission of TSEs, and thereby provide valuable hints for understanding the inability of the conformational conversion for RaPrPC. PMID:20949107

Xiong, Minqian; Peng, Yu; Yao, Wenming; Hong, Jing; Lin, Donghai

2010-01-01

339

Copper attachment to prion protein at a non-octarepeat site  

NASA Astrophysics Data System (ADS)

Prion protein (PrP) plays a causative role in a group of neurodegenerative diseases, which include "mad cow disease" or its human form variant Creutzfeld-Jacob disease. Normal function of PrP remains unknown, but it is now well established that PrP can efficiently bind copper ions and this ability has been linked to its function. The primary binding sites are located in the so-called octarepeat region located between residues 60-91. While these are by now well characterized, the sites located outside these region remain mostly undetermined. In this work, we investigate the properties of Cu binding site located at His 111 using recently developed hybrid Kohn-Sham/orbital-free density functional simulations. Experimental data indicate that copper is coordinated by either four nitrogens or three nitrogens and one oxygen. We investigate both possibilities, comparing their energetics and attachment geometries. Similarities and differences with other binding sites and implications for PrP function will also be discussed.

Hodak, Miroslav; Bernholc, Jerry

2011-03-01

340

Conformational diversity in prion protein variants influences intermolecular [beta]-sheet formation  

SciTech Connect

A conformational transition of normal cellular prion protein (PrP{sup C}) to its pathogenic form (PrP{sup Sc}) is believed to be a central event in the transmission of the devastating neurological diseases known as spongiform encephalopathies. The common methionine/valine polymorphism at residue 129 in the PrP influences disease susceptibility and phenotype. We report here seven crystal structures of human PrP variants: three of wild-type (WT) PrP containing V129, and four of the familial variants D178N and F198S, containing either M129 or V129. Comparison of these structures with each other and with previously published WT PrP structures containing M129 revealed that only WT PrPs were found to crystallize as domain-swapped dimers or closed monomers; the four mutant PrPs crystallized as non-swapped dimers. Three of the four mutant PrPs aligned to form intermolecular {beta}-sheets. Several regions of structural variability were identified, and analysis of their conformations provides an explanation for the structural features, which can influence the formation and conformation of intermolecular {beta}-sheets involving the M/V129 polymorphic residue.

Lee, Seungjoo; Antony, Lizamma; Hartmann, Rune; Knaus, Karen J.; Surewicz, Krystyna; Surewicz, Witold K.; Yee, Vivien C. (Case Western); (Cleveland Clinic)

2010-04-19

341

Phthalocyanine tetrasulfonates bind to multiple sites on natively-folded prion protein.  

PubMed

The phthalocyanine tetrasulfonates (PcTS), a class of cyclic tetrapyrroles, bind to the mammalian prion protein, PrP. Remarkably, they can act as anti-scrapie agents to prevent the formation and spread of infectious, misfolded PrP. While the effects of phthalocyanines on the diseased state have been investigated, the interaction between PcTS and PrP has not yet been extensively characterized. Here we use multiple, complementary assays (surface plasmon resonance, isothermal titration calorimetry, fluorescence correlation spectroscopy, and tryptophan fluorescence quenching) to characterize the binding of PcTS to natively-folded hamster PrP(90-232), in order to determine binding constants, ligand stoichiometry, influence of buffer ionic strength, and the effects of chelated metal ions. We found that binding strength depends strongly on chelated metal ions, with Al(3+)-PcTS binding the weakest and free-base PcTS the strongest of the three types tested (Al(3+), Zn(2+), and free-base). Buffer ionic strength also affected the binding, with K(d) increasing along with salt concentration. The binding isotherms indicated the presence of at least two different binding sites with micromolar affinities and a total stoichiometry of ~4-5 PcTS molecules per PrP molecule. PMID:22480824

Dee, Derek R; Gupta, Amar Nath; Anikovskiy, Max; Sosova, Iveta; Grandi, Elena; Rivera, Laura; Vincent, Abhilash; Brigley, Angela M; Petersen, Nils O; Woodside, Michael T

2012-06-01

342

Lipid rafts: linking prion protein to zinc transport and amyloid-? toxicity in Alzheimer's disease.  

PubMed

Dysregulation of neuronal zinc homeostasis plays a major role in many processes related to brain aging and neurodegenerative diseases, including Alzheimer's disease (AD). Yet, despite the critical role of zinc in neuronal function, the cellular mechanisms underpinning its homeostatic control are far from clear. We reported that the cellular prion protein (PrP(C)) is involved in the uptake of zinc into neurons. This PrP(C)-mediated zinc influx required the metal-binding octapeptide repeats in PrP(C) and the presence of the zinc permeable AMPA channel with which PrP(C) directly interacted. Together with the observation that PrP(C) is evolutionarily related to the ZIP family of zinc transporters, these studies indicate that PrP(C) plays a key role in neuronal zinc homeostasis. Therefore, PrP(C) could contribute to cognitive health and protect against age-related zinc dyshomeostasis but PrP(C) has also been identified as a receptor for amyloid-? oligomers which accumulate in the brains of those with AD. We propose that the different roles that PrP(C) has are due to its interaction with different ligands and/or co-receptors in lipid raft-based signaling/transport complexes. PMID:25364748

Watt, Nicole T; Griffiths, Heledd H; Hooper, Nigel M

2014-01-01

343

Lipid rafts: linking prion protein to zinc transport and amyloid-? toxicity in Alzheimer's disease  

PubMed Central

Dysregulation of neuronal zinc homeostasis plays a major role in many processes related to brain aging and neurodegenerative diseases, including Alzheimer's disease (AD). Yet, despite the critical role of zinc in neuronal function, the cellular mechanisms underpinning its homeostatic control are far from clear. We reported that the cellular prion protein (PrPC) is involved in the uptake of zinc into neurons. This PrPC-mediated zinc influx required the metal-binding octapeptide repeats in PrPC and the presence of the zinc permeable AMPA channel with which PrPC directly interacted. Together with the observation that PrPC is evolutionarily related to the ZIP family of zinc transporters, these studies indicate that PrPC plays a key role in neuronal zinc homeostasis. Therefore, PrPC could contribute to cognitive health and protect against age-related zinc dyshomeostasis but PrPC has also been identified as a receptor for amyloid-? oligomers which accumulate in the brains of those with AD. We propose that the different roles that PrPC has are due to its interaction with different ligands and/or co-receptors in lipid raft-based signaling/transport complexes. PMID:25364748

Watt, Nicole T.; Griffiths, Heledd H.; Hooper, Nigel M.

2014-01-01

344

Influence of specific HSP70 domains on fibril formation of the yeast prion protein Ure2  

PubMed Central

Ure2p is the protein determinant of the Saccharomyces cerevisiae prion state [URE3]. Constitutive overexpression of the HSP70 family member SSA1 cures cells of [URE3]. Here, we show that Ssa1p increases the lag time of Ure2p fibril formation in vitro in the presence or absence of nucleotide. The presence of the HSP40 co-chaperone Ydj1p has an additive effect on the inhibition of Ure2p fibril formation, whereas the Ydj1p H34Q mutant shows reduced inhibition alone and in combination with Ssa1p. In order to investigate the structural basis of these effects, we constructed and tested an Ssa1p mutant lacking the ATPase domain, as well as a series of C-terminal truncation mutants. The results indicate that Ssa1p can bind to Ure2p and delay fibril formation even in the absence of the ATPase domain, but interaction of Ure2p with the substrate-binding domain is strongly influenced by the C-terminal lid region. Dynamic light scattering, quartz crystal microbalance assays, pull-down assays and kinetic analysis indicate that Ssa1p interacts with both native Ure2p and fibril seeds, and reduces the rate of Ure2p fibril elongation in a concentration-dependent manner. These results provide new insights into the structural and mechanistic basis for inhibition of Ure2p fibril formation by Ssa1p and Ydj1p. PMID:23530260

Xu, Li-Qiong; Wu, Si; Buell, Alexander K.; Cohen, Samuel I. A.; Chen, Li-Jun; Hu, Wan-Hui; Cusack, Sarah A.; Itzhaki, Laura S.; Zhang, Hong; Knowles, Tuomas P. J.; Dobson, Christopher M.; Welland, Mark E.; Jones, Gary W.; Perrett, Sarah

2013-01-01

345

Synthetic amyloid-? oligomers impair long-term memory independently of cellular prion protein  

PubMed Central

Inability to form new memories is an early clinical sign of Alzheimer’s disease (AD). There is ample evidence that the amyloid-? (A?) peptide plays a key role in the pathogenesis of this disorder. Soluble, bio-derived oligomers of A? are proposed as the key mediators of synaptic and cognitive dysfunction, but more tractable models of A??mediated cognitive impairment are needed. Here we report that, in mice, acute intracerebroventricular injections of synthetic A?1–42 oligomers impaired consolidation of the long-term recognition memory, whereas mature A?1–42 fibrils and freshly dissolved peptide did not. The deficit induced by oligomers was reversible and was prevented by an anti-A? antibody. It has been suggested that the cellular prion protein (PrPC) mediates the impairment of synaptic plasticity induced by A?. We confirmed that A?1–42 oligomers interact with PrPC, with nanomolar affinity. However, PrP-expressing and PrP knock-out mice were equally susceptible to this impairment. These data suggest that A?1–42 oligomers are responsible for cognitive impairment in AD and that PrPC is not required. PMID:20133875

Balducci, Claudia; Beeg, Marten; Stravalaci, Matteo; Bastone, Antonio; Sclip, Alessandra; Biasini, Emiliano; Tapella, Laura; Colombo, Laura; Manzoni, Claudia; Borsello, Tiziana; Chiesa, Roberto; Gobbi, Marco; Salmona, Mario; Forloni, Gianluigi

2010-01-01

346

Alzheimer Amyloid-? Oligomer Bound to Post-Synaptic Prion Protein Activates Fyn to Impair Neurons  

PubMed Central

SUMMARY Amyloid-beta (A?) oligomers are thought to trigger Alzheimer’s disease (AD) pathophysiology. Cellular Prion Protein (PrPC) selectively binds oligomeric A? and can mediate AD-related phenotypes. Here, we examined the specificity, distribution and signaling from A?/PrP complexes, seeking to explain how they might alter the function of NMDA receptors in neurons. PrPC is enriched in post-synaptic densities, and A?/PrPC interaction leads to Fyn kinase activation. Soluble A? assemblies derived from human AD brain interact with PrPC to activate Fyn. A? engagement of PrPC/Fyn signaling yields phosphorylation of the NR2B subunit of NMDA-receptors, which is coupled to an initial increase and then loss of surface NMDA-receptors. A?-induced LDH release and dendritic spine loss require both PrPC and Fyn, and human familial AD transgene-induced convulsive seizures do not occur in mice lacking PrPC. These results delineate an A? oligomer signal transduction pathway requiring PrPC and Fyn to alter synaptic function with relevance to AD. PMID:22820466

Um, Ji Won; Nygaard, Haakon B.; Heiss, Jacqueline K.; Kostylev, Mikhail A.; Stagi, Massimiliano; Vortmeyer, Alexander; Wisniewski, Thomas; Gunther, Erik C.; Strittmatter, Stephen M.

2012-01-01

347

Overcoming barriers and thresholds - signaling of oligomeric A? through the prion protein to Fyn  

PubMed Central

Evidence has been mounting for an involvement of the prion protein (PrP) in a molecular pathway assumed to play a critical role in the etiology of Alzheimer disease. A currently popular model sees oligomeric amyloid ? (oA?) peptides bind directly to PrP to emanate a signal that causes activation of the cytoplasmic tyrosine kinase Fyn, an essential player in a cascade of events that ultimately leads to NMDA receptor-mediated excitotoxicity and hyper-phosphorylation of tau. The model does not reveal, however, how extracellular binding of oA? to PrP is communicated across the plasma membrane barrier to affect activation of Fyn. A scenario whereby PrP may adapt a transmembrane topology to affect Fyn activation in the absence of additional partners is currently not supported by evidence. A survey of known candidate PrP interactors leads to a small number of molecules that are known to acquire a transmembrane topology and understood to contribute to Fyn activation. Because multiple signaling pathways converge onto Fyn, a realistic model needs to take into account a reality of Fyn acting as a hub that integrates signals from multiple inhibitory and activating effectors. To clarify the role of PrP in oA?-dependent excitotoxicity, future studies may need to incorporate experimental designs that can probe the contributions of Fyn modulator pathways and rely on analogous readouts, rather than threshold effects, known to underlie excitotoxic signaling. PMID:23856335

2013-01-01

348

Cellular prion protein is essential for oligomeric amyloid-?-induced neuronal cell death  

PubMed Central

In Alzheimer disease (AD), amyloid-? (A?) oligomer is suggested to play a critical role in imitating neurodegeneration, although its pathogenic mechanism remains to be determined. Recently, the cellular prion protein (PrPC) has been reported to be an essential co-factor in mediating the neurotoxic effect of A? oligomer. However, these previous studies focused on the synaptic plasticity in either the presence or the absence of PrPC and no study to date has reported whether PrPC is required for the neuronal cell death, the most critical element of neurodegeneration in AD. Here, we show that Prnp?/? mice are resistant to the neurotoxic effect of A? oligomer in vivo and in vitro. Furthermore, application of an anti-PrPC antibody or PrPC peptide prevents A? oligomer-induced neurotoxicity. These findings are the first to demonstrate that PrPC is required for A? oligomer-induced neuronal cell death, the pathology essential to cognitive loss. PMID:22100763

Kudo, Wataru; Lee, Hyun-Pil; Zou, Wen-Quan; Wang, Xinglong; Perry, George; Zhu, Xiongwei; Smith, Mark A.; Petersen, Robert B.; Lee, Hyoung-gon

2012-01-01

349

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

PubMed Central

Transmissible spongiform encephalopathies (TSEs) or prion diseases are associated with accumulations of disease specific PrP (PrPd) 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 PrPd in the adrenal medulla of scrapie affected sheep using light and electron microscopy. Using immunogold electron microscopy, non-fibrillar forms of PrPd were shown to accumulate mainly in association with chromaffin cells, occasional nerve endings and macrophages. PrPd accumulation was associated with distinctive membrane changes of chromaffin cells including increased electron density, abnormal linearity and invaginations. Internalisation of PrPd from the chromaffin cell plasma membrane occurred in association with granule recycling following hormone exocytosis. PrPd 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 PrPd is at the level of plasma membranes. However, the precise nature of PrPd-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 PrPd with exocytic granules of the hormone trafficking system may provide an additional source of infectivity in blood. PMID:23469286

McGovern, Gillian; Jeffrey, Martin

2013-01-01

350

Early Embryonic Gene Expression Profiling of Zebrafish Prion Protein (Prp2) Morphants  

PubMed Central

Background The Prion protein (PRNP/Prp) plays a crucial role in transmissible spongiform encephalopathies (TSEs) like Creutzfeldt-Jakob disease (CJD), scrapie and mad cow disease. Notwithstanding the importance in human and animal disease, fundamental aspects of PRNP/Prp function and transmission remains unaccounted for. Methodology/Principal Findings The zebrafish (Danio rerio) genome contains three Prp encoding genes assigned prp1, prp2 and prp3. Currently, the second paralogue is believed to be the most similar to the mammalian PRNP gene in structure and function. Functional studies of the PRNP gene ortholog was addressed by prp2 morpholino (MO) knockdown experiments. Investigation of Prp2 depleted embryos revealed high mortality and apoptosis at 24 hours post fertilization (hpf) as well as impaired brain and neuronal development. In order to elucidate the underlying mechanisms, a genome-wide transcriptome analysis was carried out in viable 24 hpf morphants. The resulting changes in gene expression profiles revealed 249 differently expressed genes linked to biological processes like cell death, neurogenesis and embryonic development. Conclusions/Significance The current study contributes to the understanding of basic Prp functions and demonstrates that the zebrafish is an excellent model to address the role of Prp in vertebrates. The gene knockdown of prp2 indicates an essential biological function for the zebrafish ortholog with a morphant phenotype that suggests a neurodegenerative action and gene expression effects which are apoptosis related and effects gene networks controlling neurogenesis and embryo development. PMID:21042590

Nourizadeh-Lillabadi, Rasoul; Seilø Torgersen, Jacob; Vestrheim, Olav; König, Melanie; Aleström, Peter; Syed, Mohasina

2010-01-01

351

Functional Depletion of Mahogunin by Cytosolically Exposed Prion Protein Contributes to Neurodegeneration  

PubMed Central

The pathways leading from aberrant Prion protein (PrP) metabolism to neurodegeneration are poorly understood. Some familial PrP mutants generate increased CtmPrP, a transmembrane isoform associated with disease. In other disease situations, a potentially toxic cytosolic form (termed cyPrP) might be produced. However, the mechanisms by which CtmPrP or cyPrP cause selective neuronal dysfunction are unknown. Here, we show that both CtmPrP and cyPrP can interact with and disrupt the function of Mahogunin (Mgrn), a cytosolic ubiquitin ligase whose loss causes spongiform neurodegeneration. Cultured cells and transgenic mice expressing either CtmPrP-producing mutants or cyPrP partially phenocopy Mgrn depletion, displaying aberrant lysosomal morphology and loss of Mgrn in selected brain regions. These effects were rescued by either Mgrn overexpression, competition for PrP binding sites, or preventing cytosolic exposure of PrP. Thus, transient or partial exposure of PrP to the cytosol leads to inappropriate Mgrn sequestration that contributes to neuronal dysfunction and disease. PMID:19524515

Chakrabarti, Oishee; Hegde, Ramanujan S.

2009-01-01

352

Evidence for degradation of abnormal prion protein in tissues from sheep with scrapie during composting  

PubMed Central

This study investigated whether the abnormal prion protein (PrPSc) in tissues from sheep with scrapie would be destroyed by composting. Tissues from sheep naturally infected with scrapie were placed within fiberglass mesh bags and buried in compost piles for 108 d in experiment 1 or 148 d in experiment 2. The temperature in the compost piles rose quickly; it was above 60°C for about 2 wk and then slowly declined to the ambient temperature. Before composting, PrPSc was detected in all the tissues by Western blotting. In experiment 1, PrPSc was not detected after composting in the tissue remnants or the surrounding sawdust. In experiment 2, 1 of 5 specimens tested negative after composting, whereas PrPSc was detected in the other 4 bags, though in reduced amounts compared with those before composting. Tissue weights were reduced during composting. Analysis of the tissue remnants for microbial 16S ribosomal DNA demonstrated that there were more diverse microbes involved in experiment 1 than in experiment 2 and that the guanine and cytosine content of the microbial 16S DNA was higher in the specimens of experiment 1 than in those of experiment 2, which suggests greater dominance of thermophilic microbes in experiment 1. These results indicate that composting may have value as a means for degrading PrPSc in carcasses and other wastes. PMID:17193880

Huang, Hongsheng; Spencer, J. Lloyd; Soutyrine, Andrei; Guan, Jeiwen; Rendulich, Jasmine; Balachandran, Aru

2007-01-01

353

Dynamic prions revealed by magic.  

PubMed

Prion proteins can be propagated as amyloid fibrils with several different conformational variants. By providing structural information at atomic level for two such variants of a yeast prion, Frederick and colleagues, in this issue of Chemistry & Biology, reveal how conformational flexibility can generate phenotypic diversity. PMID:24560165

Tuite, Mick F; Howard, Mark J; Xue, Wei-Feng

2014-02-20

354

Prion Protein-mediated Toxicity of Amyloid-? Oligomers Requires Lipid Rafts and the Transmembrane LRP1*  

PubMed Central

Soluble oligomers of the amyloid-? (A?) peptide cause neurotoxicity, synaptic dysfunction, and memory impairments that underlie Alzheimer disease (AD). The cellular prion protein (PrPC) was recently identified as a high affinity neuronal receptor for A? oligomers. We report that fibrillar A? oligomers recognized by the OC antibody, which have been shown to correlate with the onset and severity of AD, bind preferentially to cells and neurons expressing PrPC. The binding of A? oligomers to cell surface PrPC, as well as their downstream activation of Fyn kinase, was dependent on the integrity of cholesterol-rich lipid rafts. In SH-SY5Y cells, fluorescence microscopy and co-localization with subcellular markers revealed that the A? oligomers co-internalized with PrPC, accumulated in endosomes, and subsequently trafficked to lysosomes. The cell surface binding, internalization, and downstream toxicity of A? oligomers was dependent on the transmembrane low density lipoprotein receptor-related protein-1 (LRP1). The binding of A? oligomers to cell surface PrPC impaired its ability to inhibit the activity of the ?-secretase BACE1, which cleaves the amyloid precursor protein to produce A?. The green tea polyphenol (?)-epigallocatechin gallate and the red wine extract resveratrol both remodeled the fibrillar conformation of A? oligomers. The resulting nonfibrillar oligomers displayed significantly reduced binding to PrPC-expressing cells and were no longer cytotoxic. These data indicate that soluble, fibrillar A? oligomers bind to PrPC in a conformation-dependent manner and require the integrity of lipid rafts and the transmembrane LRP1 for their cytotoxicity, thus revealing potential targets to alleviate the neurotoxic properties of A? oligomers in AD. PMID:23386614

Rushworth, Jo V.; Griffiths, Heledd H.; Watt, Nicole T.; Hooper, Nigel M.

2013-01-01

355

Salivary prions in sheep and deer  

PubMed Central

Scrapie of sheep and chronic wasting disease (CWD) of cervids are transmissible prion diseases. Milk and placenta have been identified as sources of scrapie prions but do not explain horizontal transmission. In contrast, CWD prions have been reported in saliva, urine and feces, which are thought to be responsible for horizontal transmission. While the titers of CWD prions have been measured in feces, levels in saliva or urine are unknown. Because sheep produce ?17 L/day of saliva and scrapie prions are present in tongue and salivary glands of infected sheep, we asked if scrapie prions are shed in saliva. We inoculated transgenic (Tg) mice expressing ovine prion protein, Tg(OvPrP) mice, with saliva from seven Cheviot sheep with scrapie. Six of seven samples transmitted prions to Tg(OvPrP) mice with titers of ?0.5 to 1.7 log ID50 U/ml. Similarly, inoculation of saliva samples from two mule deer with CWD transmitted prions to Tg(ElkPrP) mice with titers of ?1.1 to ?0.4 log ID50 U/ml. Assuming similar shedding kinetics for salivary prions as those for fecal prions of deer, we estimated the secreted salivary prion dose over a 10-mo period to be as high as 8.4 log ID50 units for sheep and 7.0 log ID50 units for deer. These estimates are similar to 7.9 log ID50 units of fecal CWD prions for deer. Because saliva is mostly swallowed, salivary prions may reinfect tissues of the gastrointestinal tract and contribute to fecal prion shedding. Salivary prions shed into the environment provide an additional mechanism for horizontal prion transmission. PMID:22453179

Tamguney, Gultekin; Richt, Jurgen A; Hamir, Amir N; Greenlee, Justin J; Miller, Michael W; Wolfe, Lisa L; Sirochman, Tracey M; Young, Alan J; Glidden, David V; Johnson, Natrina L; Giles, Kurt; DeArmond, Stephen J

2012-01-01

356

Prion transmission  

PubMed Central

Prion diseases range from being highly infectious, for example scrapie and CWD, which show facile transmission between susceptible individuals, to showing negligible horizontal transmission, such as BSE and CJD, which are spread via food or iatrogenically, respectively. Scrapie and CWD display considerable in vivo dissemination, with PrPSc and infectivity being found in a range of peripheral tissues. This in vivo dissemination appears to facilitate the recently reported excretion of prion through multiple routes such as from skin, feces, urine, milk, nasal secretions, saliva and placenta. Furthermore, excreted scrapie and CWD agent is detected within environmental samples such as water and on the surfaces of inanimate objects. The cycle of “uptake of prion from the environment—widespread in vivo prion dissemination—prion excretion—prion persistence in the environment” is likely to explain the facile transmission and maintenance of these diseases within wild and farmed populations over many years. PMID:20948292

Maddison, Ben C

2010-01-01

357

Twilight zone of protein sequence alignments  

Microsoft Academic Search

Abstract: l findings are applicable to automaticdatabase searches.Keywords: alignment quality analysis\\/evolutionary conservation\\/genome analysis\\/protein sequence alignment\\/sequence spacehoppingIntroductionProtein sequence alignments in twilight zoneProtein sequences fold into unique three-dimensional (3D)structures. However, proteins with similar sequences adoptsimilar structures (Zuckerkandl and Pauling, 1965; Doolittle,1981; Doolittle, 1986; Chothia and Lesk, 1986). Indeed, mostprotein...

Burkhard Rost

1999-01-01

358

Prions: generation and spread versus neurotoxicity.  

PubMed

Neurodegenerative diseases are characterized by the aggregation of misfolded proteins in the brain. Among these disorders are the prion diseases, which are transmissible, and in which the misfolded proteins ("prions") are also the infectious agent. Increasingly, it appears that misfolded proteins in Alzheimer and Parkinson diseases and the tauopathies also propagate in a "prion-like" manner. However, the association between prion formation, spread, and neurotoxicity is not clear. Recently, we showed that in prion disease, protein misfolding leads to neurodegeneration through dysregulation of generic proteostatic mechanisms, specifically, the unfolded protein response. Genetic and pharmacological manipulation of the unfolded protein response was neuroprotective despite continuing prion replication, hence dissociating this from neurotoxicity. The data have clear implications for treatment across the spectrum of these disorders, targeting pathogenic processes downstream of protein misfolding. PMID:24860100

Halliday, Mark; Radford, Helois; Mallucci, Giovanna R

2014-07-18

359

Modeling Amyloid-Beta as Homogeneous Dodecamers and in Complex with Cellular Prion Protein  

PubMed Central

Soluble amyloid beta (A?) peptide has been linked to the pathology of Alzheimer’s disease. A variety of soluble oligomers have been observed to be toxic, ranging from dimers to protofibrils. No tertiary structure has been identified as a single biologically relevant form, though many models are comprised of highly ordered ?-sheets. Evidence exists for much less ordered toxic oligomers. The mechanism of toxicity remains highly debated and probably involves multiple pathways. Interaction of A? oligomers with the N-terminus of the cellular form of the prion protein (PrPc) has recently been proposed. The intrinsically disordered nature of this protein and the highly polymorphic nature of A? oligomers make structural resolution of the complex exceptionally challenging. In this study, molecular dynamics simulations are performed for dodecameric assemblies of A? comprised of monomers having a single, short antiparallel ?-hairpin at the C-terminus. The resulting models, devoid of any intermolecular hydrogen bonds, are shown to correlate well with experimental data and are found to be quite stable within the hydrophobic core, whereas the ?-helical N-termini transform to a random coil state. This indicates that highly ordered assemblies are not required for stability and less ordered oligomers are a viable component in the population of soluble oligomers. In addition, a tentative model is proposed for the association of A? dimers with a double deletion mutant of the intrinsically disordered N-terminus of PrPc. This may be useful as a conceptual working model for the binding of higher order oligomers and in the design of further experiments. PMID:23145167

Gallion, Steven L.

2012-01-01

360

Prion proteins with pathogenic and protective mutations show similar structure and dynamics  

PubMed Central

Conformational change in the prion protein (PrP) is thought to be responsible for a group of rare but fatal neurodegenerative diseases of humans and other animals, including Creutzfeldt-Jakob disease and bovine spongiform encephalopathy. However, little is known about the mechanism by which normal cellular PrPs initiate and propagate the conformational change. Here, we studied backbone dynamics of the inherited pathogenic mutants (P101L and H186R), protective mutants (Q167R and Q218K), and wild type mouse PrP(89?230) at pH 5.5 and 3.5. Mutations result in minor chemical shift changes around the mutation sites except that H186R induces large chemical shift changes at distal regions. At lowered pH values, the C-terminal half of the second helix is significantly disordered for the wild type and all mutant proteins, while other parts of the protein are essentially unaffected. This destabilization is accompanied by protonation of the partially exposed histidine H186 in the second helix of the wild type protein. This region in the mutant protein H186R is disordered even at pH 5.5. The wild type and mutant proteins have similar ?s conformational exchange near the two ?-strands and have similar ns internal motions in several regions including the C-terminal half of the second helix, but only wild type and P101L have extensive ns internal motions throughout the helices. These motions mostly disappear at lower pH. Our findings raise the possibility that the pathogenic or dominant negative mutations exert their effects on some non-native intermediate form such as PrP* after conversion of cellular PrP (PrPC) into the pathogenic isoform PrPSc has been initiated; additionally, formation of PrPSc might begin within the C-terminal folded region rather than in the disordered N-terminal region. PMID:19618915

Bae, Sung-Hun; Legname, Giuseppe; Serban, Ana; Prusiner, Stanley B.; Wright, Peter E.; Dyson, H. Jane

2009-01-01

361

Defining the Conformational Features of Anchorless, Poorly Neuroinvasive Prions  

PubMed Central

Infectious prions cause diverse clinical signs and form an extraordinary range of structures, from amorphous aggregates to fibrils. How the conformation of a prion dictates the disease phenotype remains unclear. Mice expressing GPI-anchorless or GPI-anchored prion protein exposed to the same infectious prion develop fibrillar or nonfibrillar aggregates, respectively, and show a striking divergence in the disease pathogenesis. To better understand how a prion's physical properties govern the pathogenesis, infectious anchorless prions were passaged in mice expressing anchorless prion protein and the resulting prions were biochemically characterized. Serial passage of anchorless prions led to a significant decrease in the incubation period to terminal disease and altered the biochemical properties, consistent with a transmission barrier effect. After an intraperitoneal exposure, anchorless prions were only weakly neuroinvasive, as prion plaques rarely occurred in the brain yet were abundant in extracerebral sites such as heart and adipose tissue. Anchorless prions consistently showed very high stability in chaotropes or when heated in SDS, and were highly resistant to enzyme digestion. Consistent with the results in mice, anchorless prions from a human patient were also highly stable in chaotropes. These findings reveal that anchorless prions consist of fibrillar and highly stable conformers. The additional finding from our group and others that both anchorless and anchored prion fibrils are poorly neuroinvasive strengthens the hypothesis that a fibrillar prion structure impedes efficient CNS invasion. PMID:23637596

Bett, Cyrus; Kurt, Tim D.; Lucero, Melanie; Trejo, Margarita; Rozemuller, Annemieke J.; Kong, Qingzhong; Nilsson, K. Peter R.; Masliah, Eliezer; Oldstone, Michael B.; Sigurdson, Christina J.

2013-01-01

362

Spontaneous generation of prion infectivity in fatal familial insomnia knock-in mice  

E-print Network

A crucial tenet of the prion hypothesis is that misfolding of the prion protein (PrP) induced by mutations associated with familial prion disease is, in an otherwise normal mammalian brain, sufficient to generate the ...

Faas, Henryk

363

Endoplasmic reticulum stress induces PRNP prion protein gene expression in breast cancer  

PubMed Central

Introduction High prion protein (PrP) levels are associated with breast, colon and gastric cancer resistance to treatment and with a poor prognosis for the patients. However, little is known about the underlying molecular mechanism(s) regulating human PrP gene (PRNP) expression in cancers. Because endoplasmic reticulum (ER) stress is associated with solid tumors, we investigated a possible regulation of PRNP gene expression by ER stress. Methods Published microarray databases of breast cancer tissues and breast carcinoma cell lines were analyzed for PrP mRNA and ER stress marker immunoglobulin heavy chain binding protein (BiP) levels. Breast cancer tissue microarrays (TMA) were immunostained for BiP and PrP. Breast carcinoma MCF-7, MDA-MB-231, HS578T and HCC1500 cells were treated with three different ER stressors - Brefeldin A, Tunicamycin, Thapsigargin - and levels of PrP mRNA or protein assessed by RT-PCR and Western blot analyses. A human PRNP promoter-luciferase reporter was used to assess transcriptional activation by ER stressors. Site-directed mutagenesis identified the ER stress response elements (ERSE). Chromatin immunoprecipitation (ChIP) analyses were done to identify the ER stress-mediated transcriptional regulators. The role of cleaved activating transcription factor 6? (?ATF6?) and spliced X-box protein-1 (sXBP1) in PRNP gene expression was assessed with over-expression or silencing techniques. The role of PrP protection against ER stress was assessed with PrP siRNA and by using Prnp null cell lines. Results We find that mRNA levels of BiP correlated with PrP transcript levels in breast cancer tissues and breast carcinoma cell lines. PrP mRNA levels were enriched in the basal subtype and were associated with poor prognosis in breast cancer patients. Higher PrP and BiP levels correlated with increasing tumor grade in TMA. ER stress was a positive regulator of PRNP gene transcription in MCF-7 cells and luciferase reporter assays identified one ER stress response element (ERSE) conserved among primates and rodents and three primate-specific ERSEs that regulated PRNP gene expression. Among the various transactivators of the ER stress-regulated unfolded protein response (UPR), ATF6? and XBP1 transactivated PRNP gene expression, but the ability of these varied in different cell types. Functionally, PrP delayed ER stress-induced cell death. Conclusions These results establish PRNP as a novel ER stress-regulated gene that could increase survival in breast cancers. PMID:23497519

2013-01-01

364

Genetic predictions of prion disease susceptibility in carnivore species based on variability of the prion gene coding region.  

PubMed

Mammalian species vary widely in their apparent susceptibility to prion diseases. For example, several felid species developed prion disease (feline spongiform encephalopathy or FSE) during the bovine spongiform encephalopathy (BSE) epidemic in the United Kingdom, whereas no canine BSE cases were detected. Whether either of these or other groups of carnivore species can contract other prion diseases (e.g. chronic wasting disease or CWD) remains an open question. Variation in the host-encoded prion protein (PrP(C)) largely explains observed disease susceptibility patterns within ruminant species, and may explain interspecies differences in susceptibility as well. We sequenced and compared the open reading frame of the PRNP gene encoding PrP(C) protein from 609 animal samples comprising 29 species from 22 genera of the Order Carnivora; amongst these samples were 15 FSE cases. Our analysis revealed that FSE cases did not encode an identifiable disease-associated PrP polymorphism. However, all canid PrPs contained aspartic acid or glutamic acid at codon 163 which we propose provides a genetic basis for observed susceptibility differences between canids and felids. Among other carnivores studied, wolverine (Gulo gulo) and pine marten (Martes martes) were the only non-canid species to also express PrP-Asp163, which may impact on their prion diseases susceptibility. Populations of black bear (Ursus americanus) and mountain lion (Puma concolor) from Colorado showed little genetic variation in the PrP protein and no variants likely to be highly resistant to prions in general, suggesting that strain differences between BSE and CWD prions also may contribute to the limited apparent host range of the latter. PMID:23236380

Stewart, Paula; Campbell, Lauren; Skogtvedt, Susan; Griffin, Karen A; Arnemo, Jon M; Tryland, Morten; Girling, Simon; Miller, Michael W; Tranulis, Michael A; Goldmann, Wilfred

2012-01-01

365

The tyrosine kinase inhibitor imatinib mesylate delays prion neuroinvasion by inhibiting prion propagation in the periphery  

Microsoft Academic Search

Prion diseases are fatal neurodegenerative disorders with no effective therapy. A hallmark of prion disease is the conversion\\u000a of the normal cellular form of prion protein PrPC into a disease-associated isoform PrPSc. The authors recently have shown that a tyrosine kinase inhibitor, imatinib mesylate, induces clearance of PrPSc via specific inhibition of c-Abl in prion-infected cell culture models. In this

Seong-Wook Yun; Alexa Ertmer; Eckhard Flechsig; Sabine Gilch; Peter Riederer; Manfred Gerlach; Hermann M. Schätzl; Michael A. Klein

2007-01-01

366

A case of scrapie in a sheep carrying the lysine-171 allele of the prion protein gene.  

PubMed

Susceptibility to scrapie in sheep depends on the host PrP genotype. No data about the linkage of the rare ARK allele to differential scrapie susceptibility are currently available. Several tissues isolated from sheep from an Italian scrapie outbreak and carrying the ARK allele were examined for the presence of the pathological prion protein. A weak positivity was detected only by Western blot in the brainstem of one ARK/ARH sheep. This result shows that the ARK allele does not confer full resistance against scrapie and that the allele needs to be studied further before it can be considered for breeding purposes. PMID:16575479

Acutis, P L; Martucci, F; Mazza, M; Peletto, S; Iulini, B; Corona, C; Bozzetta, E; Casalone, C; Caramelli, M

2006-09-01

367

Detection of prion protein in the cerebrospinal fluid of elk (Cervus canadensis nelsoni) with chronic wasting disease using protein misfolding cyclic amplification.  

PubMed

Cerebrospinal fluid (CSF) has been examined as a possible source for preclinical diagnosis of prion diseases in hamsters and sheep. The present report describes the detection of chronic wasting disease (CWD) in the CSF of elk and evaluates its usefulness as an antemortem test for CWD. The CSF from 6 captive and 31 free-ranging adult elk was collected at necropsy and evaluated for the presence of the abnormal isoform of the prion protein that has been associated with CWD (PrP(CWD)) via protein misfolding cyclic amplification. Additionally, the obex from each animal was examined by immunohistochemistry (IHC). Four out of 6 captive animals were CWD-positive and euthanized due to signs of terminal CWD. The remaining 2 were CWD negative. None of the 31 free-range animals showed overt signs of CWD, but 12 out of 31 tested positive for CWD by IHC. Protein misfolding cyclic amplification detected PrP(CWD) from 3 of the 4 captive animals showing clinical signs of CWD and none of the nonclinical animals that were CWD positive by IHC. The data suggests that CWD prions can be detected in the CSF of elk, but only relatively late in the course of the disease. PMID:22621952

Nichols, Tracy A; Spraker, Terry R; Gidlewski, Tom; Powers, Jenny G; Telling, Glenn C; VerCauteren, Kurt C; Zabel, Mark D

2012-07-01

368

Effect of electrostatics on aggregation of prion protein Sup35 peptide  

NASA Astrophysics Data System (ADS)

Self-assembly of misfolded proteins into ordered fibrillar structures is a fundamental property of a wide range of proteins and peptides. This property is also linked with the development of various neurodegenerative diseases such as Alzheimer’s and Parkinson’s. Environmental conditions modulate the misfolding and aggregation processes. We used a peptide, CGNNQQNY, from yeast prion protein Sup35, as a model system to address effects of environmental conditions on aggregate formation. The GNNQQNY peptide self-assembles in fibrils with structural features that are similar to amyloidogenic proteins. Atomic force microscopy (AFM) and thioflavin T (ThT) fluorescence assay were employed to follow the aggregation process at various pHs and ionic strengths. We also used single molecule AFM force spectroscopy to probe interactions between the peptides under various conditions. The ThT fluorescence data showed that the peptide aggregates fast at pH values approaching the peptide isoelectric point (pI = 5.3) and the kinetics is 10 times slower at acidic pH (pH 2.0), suggesting that electrostatic interactions contribute to the peptide self-assembly into aggregates. This hypothesis was tested by experiments performed at low (11 mM) and high (150 mM) ionic strengths. Indeed, the aggregation lag time measured at pH 2 at low ionic strength (11 mM) is 195 h, whereas the lag time decreases ˜5 times when the ionic strength is increased to 150 mM. At conditions close to the pI value, pH 5.6, the aggregation lag time is 12 ± 6 h under low ionic strength, and there is minimal change to the lag time at 150 mM NaCl. The ionic strength also influences the morphology of aggregates visualized with AFM. In pH 2.0 and at high ionic strength, the aggregates are twofold taller than those formed at low ionic strength. In parallel, AFM force spectroscopy studies revealed minimal contribution of electrostatics to dissociation of transient peptide dimers.

Portillo, Alexander M.; Krasnoslobodtsev, Alexey V.; Lyubchenko, Yuri L.

2012-04-01

369

Assessing the Susceptibility of Transgenic Mice Overexpressing Deer Prion Protein to Bovine Spongiform Encephalopathy  

PubMed Central

Several transgenic mouse models have been developed which facilitate the transmission of chronic wasting disease (CWD) of cervids and allow prion strain discrimination. The present study was designed to assess the susceptibility of the prototypic mouse line, Tg(CerPrP)1536+/?, to bovine spongiform encephalopathy (BSE) prions, which have the ability to overcome species barriers. Tg(CerPrP)1536+/? mice challenged with red deer-adapted BSE resulted in 90% to 100% attack rates, and BSE from cattle failed to transmit, indicating agent adaptation in the deer. PMID:24257620

Vickery, Christopher M.; Lockey, Richard; Holder, Thomas M.; Thorne, Leigh; Beck, Katy E.; Wilson, Christina; Denyer, Margaret; Sheehan, John; Marsh, Sarah; Webb, Paul R.; Dexter, Ian; Norman, Angela; Popescu, Emma; Schneider, Amanda; Holden, Paul; Griffiths, Peter C.; Plater, Jane M.; Dagleish, Mark P.; Martin, Stuart; Telling, Glenn C.; Simmons, Marion M.

2014-01-01

370

Dynamic changes and surveillance function of prion protein expression in gastric cancer drug resistance  

PubMed Central

AIM: To explore the dynamic changes of prion protein (PrPc) in the process of gastric cancer drug resistance and the role of PrPc expression in the prognosis of gastric cancer patients receiving chemotherapy. METHODS: A series of gastric cancer cell lines resistant to different concentrations of adriamycin was established, and the expression of PrPc, Bcl-2 and Bax was detected in these cells. Apoptosis was determined using Annexin V staining. Western blotting and immunohistochemistry were performed to detect the expression of PrPc in patients receiving chemotherapy and to explore the role of PrPc expression in predicting the chemosensitivity and the outcome of gastric cancer patients receiving chemotherapy. Follow-up was performed for 2 years. RESULTS: PrPc expression was increased with the increase in drug resistance. Bcl-2, together with PrPc, increased the level of anti-apoptosis of cancer cells. Increased PrPc expression predicted the enhanced level of anti-apoptosis and resistance to anticancer drugs. PrPc expression could be used as a marker for predicting the efficacy of chemotherapy and the prognosis of gastric cancer. Increased PrPc expression predicted both poor chemosensitivity and a low 2-year survival rate. Contrarily, low PrPc expression predicted favorable chemosensitivity and a relatively high 2-year survival rate. CONCLUSION: PrPc expression is associated with histological types and differentiation of gastric cancer cells; The PrPc expression level might be a valuable marker in predicting the efficacy of chemotherapy and the prognosis of gastric cancer patients receiving chemotherapy. PMID:22046086

Wang, Ji-Heng; Du, Jing-Ping; Zhang, Ying-Hai; Zhao, Xiao-Jun; Fan, Ru-Ying; Wang, Zhi-Hong; Wu, Zi-Tao; Han, Ying

2011-01-01

371

The prion protein ligand, stress-inducible phosphoprotein 1, regulates amyloid-? oligomer toxicity.  

PubMed

In Alzheimer's disease (AD), soluble amyloid-? oligomers (A?Os) trigger neurotoxic signaling, at least partially, via the cellular prion protein (PrP(C)). However, it is unknown whether other ligands of PrP(C) can regulate this potentially toxic interaction. Stress-inducible phosphoprotein 1 (STI1), an Hsp90 cochaperone secreted by astrocytes, binds to PrP(C) in the vicinity of the A?O binding site to protect neurons against toxic stimuli. Here, we investigated a potential role of STI1 in A?O toxicity. We confirmed the specific binding of A?Os and STI1 to the PrP and showed that STI1 efficiently inhibited A?O binding to PrP in vitro (IC50 of ?70 nm) and also decreased A?O binding to cultured mouse primary hippocampal neurons. Treatment with STI1 prevented A?O-induced synaptic loss and neuronal death in mouse cultured neurons and long-term potentiation inhibition in mouse hippocampal slices. Interestingly, STI1-haploinsufficient neurons were more sensitive to A?O-induced cell death and could be rescued by treatment with recombinant STI1. Noteworthy, both A?O binding to PrP(C) and PrP(C)-dependent A?O toxicity were inhibited by TPR2A, the PrP(C)-interacting domain of STI1. Additionally, PrP(C)-STI1 engagement activated ?7 nicotinic acetylcholine receptors, which participated in neuroprotection against A?O-induced toxicity. We found an age-dependent upregulation of cortical STI1 in the APPswe/PS1dE9 mouse model of AD and in the brains of AD-affected individuals, suggesting a compensatory response. Our findings reveal a previously unrecognized role of the PrP(C) ligand STI1 in protecting neurons in AD and suggest a novel pathway that may help to offset A?O-induced toxicity. PMID:24133259

Ostapchenko, Valeriy G; Beraldo, Flavio H; Mohammad, Amro H; Xie, Yu-Feng; Hirata, Pedro H F; Magalhaes, Ana C; Lamour, Guillaume; Li, Hongbin; Maciejewski, Andrzej; Belrose, Jillian C; Teixeira, Bianca L; Fahnestock, Margaret; Ferreira, Sergio T; Cashman, Neil R; Hajj, Glaucia N M; Jackson, Michael F; Choy, Wing-Yiu; MacDonald, John F; Martins, Vilma R; Prado, Vania F; Prado, Marco A M

2013-10-16

372

Prion protein genotypes of Italian sheep breeds with lysine-171 and phenylalanine-141 detection.  

PubMed

Amino acid polymorphisms of the prion protein gene influence sheep susceptibility to classical and atypical scrapie. Substitutions at codons 136, 154 and 171 play an important role in classical scrapie. Codon 141 leucine to phenylalanine mutation (AFRQ) has been recognized as an increased risk factor for atypical scrapie. In addition a rare allele with lysine at codon 171 (ARK) has been detected in Mediterranean sheep breeds. The presence of ARK poses two problems: the determination of its frequency and its possible interference with genotyping output of routine methods lacking specific detection capacity for ARK. The aim of our work was the development of a routine genotyping method with the capacity to identify ARK and AFRQ in addition to the normally detected alleles and to determine the frequencies of all these alleles in 5 main Italian breeds: Sarda (n=2494), Bergamasca (n=2686), Appenninica (n=297), Comisana (n=361) and Massese (n=402). A multiplex primer extension assay targeting the six single nucleotide polymorphisms of interest was developed. Allele frequencies revealed a very low level of ARR in Bergamasca (6.91%) as opposed to the other breeds, very diverse levels of AFRQ ranging from absence in Comisana to 10.70% in Massese and a restricted presence of ARK. This allele has only been detected in Bargamasca with a significant 3.67% and marginally in Appenninica (0.34%). These results underline the need for adequate routine methods for genotyping of breeds with alleles that can interfere with typing of important codons such as the case of ARK for codon 171. PMID:19157728

Pongolini, Stefano; Bergamini, Federica; Iori, Alessandra; Migliore, Sergio; Corradi, Attilio; Bassi, Stefano

2009-05-28

373

A Nine Amino Acid Domain is Essential for Mutant Prion Protein Toxicity  

PubMed Central

Transgenic mice expressing PrP molecules with several different internal deletions display spontaneous neurodegenerative phenotypes that can be dose-dependently suppressed by co-expression of wild-type PrP. Each of these deletions, including the largest one (?32–134), retains nine amino acids immediately following the signal peptide cleavage site (residues 23–31; KKRPKPGGW). These residues have been implicated in several biological functions of PrP, including endocytic trafficking and binding of glycosaminoglycans. We report here on our experiments to test the role of this domain in the toxicity of deleted forms of PrP. We find that transgenic mice expressing ?23–134 PrP display no clinical symptoms or neuropathology, in contrast to mice expressing ?32–134 PrP, suggesting that residues 23–31 are essential for the toxic phenotype. Using a newly developed cell culture assay, we narrow the essential region to amino acids 23–26, and we show that mutant PrP toxicity is not related to the role of the N-terminal residues in endocytosis or binding to endogenous glycosaminoglycans. However, we find that mutant PrP toxicity is potently inhibited by application of exogenous glycosaminoglycans, suggesting that the latter molecules block an essential interaction between the N-terminus of PrP and a membrane-associated target site. Our results demonstrate that a short segment containing positively charged amino acids at the N-terminus of PrP plays an essential role in mediating PrP-related neurotoxicity. This finding identifies a protein domain that may serve as a drug target for amelioration of prion neurotoxicity. PMID:21957261

Westergard, Laura; Turnbaugh, Jessie A.; Harris, David A.

2011-01-01

374

Molecular advances in understanding inherited prion diseases  

Microsoft Academic Search

The prion diseases are neurodegenerative disorders that have attracted great interest because of the possible link between\\u000a bovine spongiform encephalopathy (BSE) and variant Creutzfeldt-Jakob disease (CTD) in humans. Possible transmission of these\\u000a diseases has been linked to a single protein termed the prion protein. This protein is an abnormal isoform of a normal synaptic\\u000a glycoprotein. The majority of prion diseases

David R. Brown

2002-01-01

375

Chronic Lymphocytic Inflammation Specifies the Organ Tropism of Prions  

NASA Astrophysics Data System (ADS)

Prions typically accumulate in nervous and lymphoid tissues. Because proinflammatory cytokines and immune cells are required for lymphoid prion replication, we tested whether inflammatory conditions affect prion pathogenesis. We administered prions to mice with five inflammatory diseases of the kidney, pancreas, or liver. In all cases, chronic lymphocytic inflammation enabled prion accumulation in otherwise prion-free organs. Inflammatory foci consistently correlated with lymphotoxin up-regulation and ectopic induction of FDC-M1+ cells expressing the normal cellular prion protein PrPC. By contrast, inflamed organs of mice lacking lymphotoxin-? or its receptor did not accumulate the abnormal isoform PrPSc, nor did they display infectivity upon prion inoculation. By expanding the tissue distribution of prions, chronic inflammatory conditions may act as modifiers of natural and iatrogenic prion transmission.

Heikenwalder, Mathias; Zeller, Nicolas; Seeger, Harald; Prinz, Marco; Klöhn, Peter-Christian; Schwarz, Petra; Ruddle, Nancy H.; Weissmann, Charles; Aguzzi, Adriano

2005-02-01

376

Mammalian Prion protein expression in yeast; a model for transmembrane insertion.  

PubMed

The prion protein (PrP), a GPI-anchored glycoprotein, is inefficiently secreted by mammalian microsomes, 50% being found as transmembrane (TM) proteins with the central TM1 segment spanning the membrane. TM1 hydrophobicity is marginal for lateral membrane insertion, which is primarily driven by hydrophobic interaction between the ER translocon and substrates in transit. Most inserted TM1 has its N-terminus in the ER lumen (Ntm orientation), as expected for arrest of normal secretion. However, 20% is found in inverted Ctm orientation. These are minor species in vivo, presumably a consequence of efficient quality control. PrP mutations that increase TM1 hydrophobicity result in increased Ctm insertion, both in vitro and in mouse brain, and a strong correlation is found between CtmPrP insertion and neuropathology in transgenic mice; a copper-dependent pathogenicity mechanism is suggested. PrP fusions with a C-terminal epitope tag, when expressed in yeast cells at moderate levels, appear to interact efficiently with the translocon, providing a useful model for testing the effects of PrP mutations on TM insertion and orientation. However, secretion of PrP by the mammalian translocon requires the TRAP complex, absent in yeast, where essentially all PrP ends up as TM species, 85-90% Ntm and 10-15% Ctm. Although yeast is, therefore, an incomplete mimic of mammalian PrP trafficking, effects on Ctm insertion of mutations increasing TM1 hydrophobicity closely reflect those seen in vitro. Electrostatic substrate-translocon interactions are a major determinant of TM protein insertion orientation and the yeast model was used to investigate the role of the large negative charge difference across TM1, a likely cause of translocation delay that would favor TM insertion and Ctm orientation. An increase in ?Ch from -5 to -7 caused a marked increase in Ctm insertion, while a decrease to -3 or -1 allowed 35 and about 65% secretion, respectively. Utility of the yeast model and the role of this charge difference in driving PrP membrane insertion are confirmed. PMID:24141197

Tipper, Donald; Martinez-Vilchez, Immaculada; Markgren, Lucas; Kagalwala, Din Z

2013-01-01

377

Antagonistic interactions between yeast [PSI(+)] and [URE3] prions and curing of [URE3] by Hsp70 protein chaperone Ssa1p but not by Ssa2p.  

PubMed

The yeast [PSI(+)], [URE3], and [PIN(+)] genetic elements are prion forms of Sup35p, Ure2p, and Rnq1p, respectively. Overexpression of Sup35p, Ure2p, or Rnq1p leads to increased de novo appearance of [PSI(+)], [URE3], and [PIN(+)], respectively. This inducible appearance of [PSI(+)] was shown to be dependent on the presence of [PIN(+)] or [URE3] or overexpression of other yeast proteins that have stretches of polar residues similar to the prion-determining domains of the known prion proteins. In a similar manner, [PSI(+)] and [URE3] facilitate the appearance of [PIN(+)]. In contrast to these positive interactions, here we find that in the presence of [PIN(+)], [PSI(+)] and [URE3] repressed each other's propagation and de novo appearance. Elevated expression of Hsp104 and Hsp70 (Ssa2p) had little effect on these interactions, ruling out competition between the two prions for limiting amounts of these protein chaperones. In contrast, we find that constitutive overexpression of SSA1 but not SSA2 cured cells of [URE3], uncovering a specific interaction between Ssa1p and [URE3] and a functional distinction between these nearly identical Hsp70 isoforms. We also find that Hsp104 abundance, which critically affects [PSI(+)] propagation, is elevated when [URE3] is present. Our results are consistent with the notion that proteins that have a propensity to form prions may interact with heterologous prions but, as we now show, in a negative manner. Our data also suggest that differences in how [PSI(+)] and [URE3] interact with Hsp104 and Hsp70 may contribute to their antagonistic interactions. PMID:11997496

Schwimmer, Christine; Masison, Daniel C

2002-06-01

378

Scrapie replication in lymphoid tissues depends on prion protein-expressing follicular dendritic cells  

Microsoft Academic Search

The immune system is central in the pathogenesis of scrapie and other transmissible spongiform encephalopathies (TSEs) or 'prion' diseases. After infecting by peripheral (intraperitoneal or oral) routes, most TSE agents replicate in spleen and lymph nodes before neuroinvasion. Characterization of the cells supporting replication in these tissues is essential to understanding early pathogenesis and may indicate potential targets for therapy,

K. L. Brown; K. Stewart; D. L. Ritchie; N. A. Mabbott; A. Williams; H. Fraser; W. I. Morrison; M. E. Bruce

1999-01-01

379

Statistical Mechanics of Prion Diseases  

SciTech Connect

We present a two-dimensional, lattice based, protein-level statistical mechanical model for prion diseases (e.g., mad cow disease) with concomitant prion protein misfolding and aggregation. Our studies lead us to the hypothesis that the observed broad incubation time distribution in epidemiological data reflect fluctuation dominated growth seeded by a few nanometer scale aggregates, while much narrower incubation time distributions for innoculated lab animals arise from statistical self-averaging. We model ''species barriers'' to prion infection and assess a related treatment protocol.

Slepoy, A.; Singh, R. R. P.; Pazmandi, F.; Kulkarni, R. V.; Cox, D. L.

2001-07-30

380

Evidence for a Protein Mutator in Yeast: Role of the Hsp70Related Chaperone Ssb in Formation, Stability, and Toxicity of the (PSI) Prion  

Microsoft Academic Search

Propagation of the yeast protein-based non-Mendelian element (PSI), a prion-like form of the release factor Sup35, was shown to be regulated by the interplay between chaperone proteins Hsp104 and Hsp70. While overproduction of Hsp104 protein cures cells of (PSI), overproduction of the Ssa1 protein of the Hsp70 family protects (PSI) from the curing effect of Hsp104. Here we demonstrate that

YURY O. CHERNOFF; GARY P. NEWNAM; JAIJIT KUMAR; KIM ALLEN; AMY D. ZINK

1999-01-01

381

NMR structure of the bank vole prion protein at 20 degrees C contains a structured loop of residues 165-171.  

PubMed

The recent introduction of bank vole (Clethrionomys glareolus) as an additional laboratory animal for research on prion diseases revealed an important difference when compared to the mouse and the Syrian hamster, since bank voles show a high susceptibility to infection by brain homogenates from a wide range of diseased species such as sheep, goats, and humans. In this context, we determined the NMR structure of the C-terminal globular domain of the recombinant bank vole prion protein (bvPrP) [bvPrP(121-231)] at 20 degrees C. bvPrP(121-231) has the same overall architecture as other mammalian PrPs, with three alpha-helices and an antiparallel beta-sheet, but it differs from PrP of the mouse and most other mammalian species in that the loop connecting the second beta-strand and helix alpha2 is precisely defined at 20 degrees C. This is similar to the previously described structures of elk PrP and the designed mouse PrP (mPrP) variant mPrP[S170N,N174T](121-231), whereas Syrian hamster PrP displays a structure that is in-between these limiting cases. Studies with the newly designed variant mPrP[S170N](121-231), which contains the same loop sequence as bvPrP, now also showed that the single-amino-acid substitution S170N in mPrP is sufficient for obtaining a well-defined loop, thus providing the rationale for this local structural feature in bvPrP. PMID:18773909

Christen, Barbara; Pérez, Daniel R; Hornemann, Simone; Wüthrich, Kurt

2008-11-01

382

Acid-induced Molten Globule State of a Prion Protein: CRUCIAL ROLE OF STRAND 1-HELIX 1-STRAND 2 SEGMENT.  

PubMed

The conversion of a cellular prion protein (PrP(C)) to its pathogenic isoform (PrP(Sc)) is a critical event in the pathogenesis of prion diseases. Pathogenic conversion is usually associated with the oligomerization process; therefore, the conformational characteristics of the pre-oligomer state may provide insights into the conversion process. Previous studies indicate that PrP(C) is prone to oligomer formation at low pH, but the conformation of the pre-oligomer state remains unknown. In this study, we systematically analyzed the acid-induced conformational changes of PrP(C) and discovered a unique acid-induced molten globule state at pH 2.0 termed the "A-state." We characterized the structure of the A-state using far/near-UV CD, 1-anilino-8-naphthalene sulfonate fluorescence, size exclusion chromatography, and NMR. Deuterium exchange experiments with NMR detection revealed its first unique structure ever reported thus far; i.e. the Strand 1-Helix 1-Strand 2 segment at the N terminus was preferentially unfolded, whereas the Helix 2-Helix 3 segment at the C terminus remained marginally stable. This conformational change could be triggered by the protonation of Asp(144), Asp(147), and Glu(196), followed by disruption of key salt bridges in PrP(C). Moreover, the initial population of the A-state at low pH (pH 2.0-5.0) was well correlated with the rate of the ?-rich oligomer formation, suggesting that the A-state is the pre-oligomer state. Thus, the specific conformation of the A-state would provide crucial insights into the mechanisms of oligomerization and further pathogenic conversion as well as facilitating the design of novel medical chaperones for treating prion diseases. PMID:25217639

Honda, Ryo P; Yamaguchi, Kei-Ichi; Kuwata, Kazuo

2014-10-31

383

Mouse models for studying the formation and propagation of prions.  

PubMed

Prions are self-propagating protein conformers that cause a variety of neurodegenerative disorders in humans and animals. Mouse models have played key roles in deciphering the biology of prions and in assessing candidate therapeutics. The development of transgenic mice that form prions spontaneously in the brain has advanced our understanding of sporadic and genetic prion diseases. Furthermore, the realization that many proteins can become prions has necessitated the development of mouse models for assessing the potential transmissibility of common neurodegenerative diseases. As the universe of prion diseases continues to expand, mouse models will remain crucial for interrogating these devastating illnesses. PMID:24860095

Watts, Joel C; Prusiner, Stanley B

2014-07-18

384

Animal prion diseases.  

PubMed

Prion diseases occur in many animal species, most notably in ruminants. While scrapie in sheep has been recognised for three centuries and goat scrapie has been recognised for decades, BSE in cattle is a relatively novel disease which was first diagnosed in the UK in the mid 1980s. BSE was most likely caused through dietary exposure to animal feed contaminated with prions and disease was subsequently transmitted to people. The BSE epidemic is almost at an end, but the recent identification of so called atypical forms of BSE and scrapie pose many questions about the possible spectrum of prion diseases in animals and their transmissibility to other species, including humans.The pathogenesis of animal prion diseases has been studied both in natural infections and in experimental animal models. Detection of infectivity is greatly helped by suitable rodent models, in particular transgenic mice. Clinically infected animals show characteristic neuropathology in the brain and spinal cord which is accompanied by the accumulation of a conformationally altered, protease-resistant host protein. The post-mortem diagnosis is based on the detection of this protein, PrP(Sc), but despite recent impressive developments a routine ante-mortem diagnostic test has proved elusive.There is no treatment for prion diseases in animals, but disease outbreaks are controlled through a mixture of movement restrictions on holdings, culling of affected animals and herds and, for classical scrapie in sheep, selective breeding for genetic resistance. Prions are very stable and can remain in the environment for prolonged periods. This poses serious practical questions with regard to the decontamination of infected premises. The control of BSE specifically through restrictions in animal feeding practises has been successful, but the changing spectrum of these diseases plus the economic pressures to relax feed bans and reduce levels of surveillance will require constant vigilance to safeguard animal and public health. PMID:23225014

Windl, Otto; Dawson, Mike

2012-01-01

385

Detection of bovine spongiform encephalopathy, ovine scrapie prion-related protein (PrPSc) and normal PrPc by monoclonal antibodies raised to copper-refolded prion protein.  

PubMed Central

Prion-related protein (PrP) is a glycosylphosphatidylinositol-linked cell-surface protein expressed by a wide variety of cells, including those of the nervous system and the immune system. Several functions of normal cellular PrP (PrPc) have been proposed that may be associated with the capacity of this protein to bind copper. In the present study, we describe the generation of a panel of monoclonal antibodies raised to copper-refolded PrP, which may be used to analyse the normal and disease-associated forms of this protein. The anti-PrP monoclonal antibodies were reactive by Western blot and ELISA with recombinant murine PrPc refolded in the presence or absence of either copper or manganese, and with the disease-susceptible allelic form V136R154Q171 ('VRQ'; where single-letter amino-acid notation has been used) and disease-resistant allelic form A136R154R171 ('ARR') of recombinant ovine PrPc. FACS analysis of lymphoid cells using these monoclonal antibodies showed that wild-type non-activated mouse lymphocytes expressed little, if any, PrPc. These monoclonal antibodies were shown to react with the unglycosylated and monoglycosylated forms of PrPSc (abnormal disease-specific conformation of PrP) in prion-infected tissue samples from all of the different species tested by Western blot. In addition, this analysis allowed one to make a distinction between bovine spongiform encephalopathy ('BSE') and scrapie PrPSc) isolates from experimentally infected sheep on the basis of their different electrophoretic mobilities. PMID:12429022

Thackray, Alana M; Madec, Jean-Yves; Wong, Edmond; Morgan-Warren, Robert; Brown, David R; Baron, Thierry; Bujdoso, Raymond

2003-01-01

386

A novel copper(II) coordination at His186 in full-length murine prion protein  

SciTech Connect

To explore Cu(II) ion coordination by His{sup 186} in the C-terminal domain of full-length prion protein (moPrP), we utilized the magnetic dipolar interaction between a paramagnetic metal, Cu(II) ion, and a spin probe introduced in the neighborhood of the postulated binding site by the spin labeling technique (SDSL technique). Six moPrP mutants, moPrP(D143C), moPrP(Y148C), moPrP(E151C), moPrP(Y156C), moPrP(T189C), and moPrP(Y156C,H186A), were reacted with a methane thiosulfonate spin probe and a nitroxide residue (R1) was created in the binding site of each one. Line broadening of the ESR spectra was induced in the presence of Cu(II) ions in moPrP(Y148R1), moPrP(Y151R1), moPrP(Y156R1), and moPrP(T189R1) but not moPrP(D143R1). This line broadening indicated the presence of electron-electron dipolar interaction between Cu(II) and the nitroxide spin probe, suggesting that each interspin distance was within 20 A. The interspin distance ranges between Cu(II) and the spin probes of moPrP(Y148R1), moPrP(Y151R1), moPrP(Y156R1), and moPrP(T189R1) were estimated to be 12.1 A, 18.1 A, 10.7 A, and 8.4 A, respectively. In moPrP(Y156R1,H186A), line broadening between Cu(II) and the spin probe was not observed. These results suggest that a novel Cu(II) binding site is involved in His186 in the Helix2 region of the C-terminal domain of moPrP{sup C}.

Watanabe, Yasuko [Laboratory of Radiation Biology, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan)] [Laboratory of Radiation Biology, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan); Hiraoka, Wakako [Laboratory of Biophysics, School of Science and Technology, Meiji University, Kawasaki 214-8571 (Japan)] [Laboratory of Biophysics, School of Science and Technology, Meiji University, Kawasaki 214-8571 (Japan); Igarashi, Manabu; Ito, Kimihito [Department of Global Epidemiology, Hokkaido University Research Center for Zoonosis Control, Sapporo 001-0020 (Japan)] [Department of Global Epidemiology, Hokkaido University Research Center for Zoonosis Control, Sapporo 001-0020 (Japan); Shimoyama, Yuhei [Soft-Matter Physics Laboratory, Graduate School of Emergent Science, Muroran Institute of Technology, Muroran 050-8585 (Japan)] [Soft-Matter Physics Laboratory, Graduate School of Emergent Science, Muroran Institute of Technology, Muroran 050-8585 (Japan); Horiuchi, Motohiro [Laboratory of Prion Diseases, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan)] [Laboratory of Prion Diseases, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan); Yamamori, Tohru; Yasui, Hironobu; Kuwabara, Mikinori [Laboratory of Radiation Biology, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan)] [Laboratory of Radiation Biology, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan); Inagaki, Fuyuhiko [Laboratory of Structural Biology, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812 (Japan)] [Laboratory of Structural Biology, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812 (Japan); Inanami, Osamu, E-mail: inanami@vetmed.hokudai.ac.jp [Laboratory of Radiation Biology, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan)] [Laboratory of Radiation Biology, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan)

2010-04-09

387

The kinetics of proteinase K digestion of linear prion polymers  

E-print Network

The kinetics of proteinase K digestion of linear prion polymers Joanna Masel* and Vincent A. A are caused by a protein-only infectious agent, known as a prion. It is not clear how a protein can be capable of replicating itself, and the mechanism remains controversial. One in£uential model hypothesizes that prions

388

A virtual lab for exploring the yeast prion  

E-print Network

A virtual lab for exploring the ¢¡¤£¦¥¨§© yeast prion Jacqueline L. Whalley , Mick F. Tuite within the cell of a prion protein in yeast. The biological background to the project is outlined for this transformation process. Abnormal forms of proteins which have this infectious property and known as prion

Kent, University of

389

Different allelic effects of the codons 136 and 171 of the prion protein gene in sheep with natural scrapie.  

PubMed

Scrapie is a transmissible degenerative disease of the central nervous system occurring naturally in sheep. It belongs to the group of prion diseases also affecting man in which an abnormal isoform of the host-encoded prion protein (PrP) accumulating in the brain is responsible for neuronal death. Three main polymorphisms have been described in the sheep PrP gene, at positions 136, 154 and 171. A strong association between susceptibility/resistance to natural scrapie and a dimorphism at codon 136 of the ovine PrP gene has been reported in several breeds, including Romanov. This dimorphism, however, is not found in all scrapie-affected breeds. We have compared the PrP genotypes of Lacaune sheep obtained from enzootically affected flocks with those of apparently healthy sheep. A third variant at codon 171 was also evidenced. The results were compared with those obtained in a single experimental Romanov flock orally challenged with nematode parasites in which scrapie suddenly appeared and killed 80% of the sheep. We present evidence that, even in different epizootological circumstances, the major genetic factor controlling the susceptibility/resistance to natural scrapie in sheep, is represented by codon 171 genotype of the PrP gene. We also suggest that a modification of the allelic effects of codon 136 can occur in heavily infected animals. PMID:7636494

Clouscard, C; Beaudry, P; Elsen, J M; Milan, D; Dussaucy, M; Bounneau, C; Schelcher, F; Chatelain, J; Launay, J M; Laplanche, J L

1995-08-01

390

Targeted mutation of the gene encoding prion protein in zebrafish reveals a conserved role in neuron excitability.  

PubMed

The function of the cellular prion protein (PrP(C)) in healthy brains remains poorly understood, in part because Prnp knockout mice are viable. On the other hand, transient knockdown of Prnp homologs in zebrafish (including two paralogs, prp1 and prp2) has suggested that PrP(C) is required for CNS development, cell adhesion, and neuroprotection. It has been argued that zebrafish Prp2 is most similar to mammalian PrP(C), yet it has remained intransigent to the most thorough confirmations of reagent specificity during knockdown. Thus we investigated the role of prp2 using targeted gene disruption via zinc finger nucleases. Prp2(-/-) zebrafish were viable and did not display overt developmental phenotypes. Back-crossing female prp2(-/-) fish ruled out a role for maternal mRNA contributions. Prp2(-/-) larvae were found to have increased seizure-like behavior following exposure to the convulsant pentylenetetrazol (PTZ), as compared to wild type fish. In situ recordings from intact hindbrains demonstrated that prp2 regulates closing of N-Methyl-d-aspartate (NMDA) receptors, concomitant with neuroprotection during glutamate excitotoxicity. Overall, the knockout of Prp2 function in zebrafish independently confirmed hypothesized roles for PrP, identifying deeply conserved functions in post-developmental regulation of neuron excitability that are consequential to the etiology of prion and Alzheimer diseases. PMID:23523635

Fleisch, Valerie C; Leighton, Patricia L A; Wang, Hao; Pillay, Laura M; Ritzel, R Gary; Bhinder, Ganive; Roy, Birbickram; Tierney, Keith B; Ali, Declan W; Waskiewicz, Andrew J; Allison, W Ted

2013-07-01

391

Coupled action of ?-glutamyl transpeptidase-glutathione and keratinase effectively degrades feather keratin and surrogate prion protein, Sup 35NM.  

PubMed

Recombinant Escherichia coli HB101 harboring keratinase rKP2 from Pseudomonas aeruginosa KS-1 degraded 2% chicken feather in LB-Amp medium in 24h. SEM analysis and detailed studies revealed that bacterial colonization of feather was a pre-requisite for degradation of feather by keratinase. The mechanism of sulfitolysis revealed involvement of free cystinyl group as a source of redox during colonization as DTNB inhibited feather degradation by rKP2. Involvement of GGT-GSH system in contribution of free cystinyl group for redox was established by using GGT knockout recombinant E. coli strain that failed to degrade feather inspite of successful colonization and keratinase production. Short term experiments further confirmed enhanced protein release from feather keratin in presence of GGT-GSH redox. In the presence of similar redox, rKP2 also degraded surrogate prion protein, Sup 35NM in 15 min at 37°C, pH 7.0. PMID:22776236

Sharma, Richa; Gupta, Rani

2012-09-01

392

Fungal prion HET-s as a model for structural complexity and self-propagation in prions  

PubMed Central

The highly ordered and reproducible structure of the fungal prion HET-s makes it an excelle