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1.
The cellular prion protein (PrPC) is an N-glycosylated GPI-anchored protein usually present in lipid rafts with numerous putative functions. When it changes its conformation to a pathological isoform (then referred to as PrPSc), it is an essential part of the prion, the agent causing fatal and transmissible neurodegenerative prion diseases. There is growing evidence that toxicity and neuronal damage on the one hand and propagation/infectivity on the other hand are two distinct processes of the disease and that the GPI-anchor attachment of PrPC and PrPSc plays an important role in protein localization and in neurotoxicity. Here we review how the signal sequence of the GPI-anchor matters in PrPC localization, how an altered cellular localization of PrPC or differences in GPI-anchor composition can affect prion infection, and we discuss through which mechanisms changes on the anchorage of PrPC can modify the disease process. 相似文献
2.
Elizaveta Katorcha Natallia Makarava Regina Savtchenko Alessandra d′Azzo Ilia V. Baskakov 《PLoS pathogens》2014,10(9)
The central event underlying prion diseases involves conformational change of the cellular form of the prion protein (PrPC) into the disease-associated, transmissible form (PrPSc). PrPC 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 PrPC and the strain-specific structure of PrPSc. The current work highlights the previously unappreciated role of sialylation of PrPC glycans in prion pathogenesis, including its role in controlling prion replication rate, infectivity, cross-species barrier and PrPSc glycoform ratio. The current study demonstrates that undersialylated PrPC is selected during prion amplification in Protein Misfolding Cyclic Amplification (PMCAb) at the expense of oversialylated PrPC. As a result, PMCAb-derived PrPSc was less sialylated than brain-derived PrPSc. A decrease in PrPSc sialylation correlated with a drop in infectivity of PMCAb-derived material. Nevertheless, enzymatic de-sialylation of PrPC using sialidase was found to increase the rate of PrPSc amplification in PMCAb from 10- to 10,000-fold in a strain-dependent manner. Moreover, de-sialylation of PrPC 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 PrPC was also found to control PrPSc glycoform ratio. A decrease in PrPC sialylation levels resulted in a higher percentage of the diglycosylated glycoform in PrPSc. 2D analysis of charge distribution revealed that the sialylation status of brain-derived PrPC differed from that of spleen-derived PrPC. Knocking out lysosomal sialidase Neu1 did not change the sialylation status of brain-derived PrPC, suggesting that Neu1 is not responsible for desialylation of PrPC. The current work highlights previously unappreciated role of PrPC sialylation in prion diseases and opens multiple new research directions, including development of new therapeutic approaches. 相似文献
3.
Molecules that inhibit the formation of an abnormal isoform of prion protein (PrPSc) in prion-infected cells are candidate therapeutic agents for prion diseases. Understanding how these molecules inhibit PrPSc formation provides logical basis for proper evaluation of their therapeutic potential. In this study, we extensively analyzed the effects of the anti-PrP monoclonal antibody (mAb) 44B1, pentosan polysulfate (PPS), chlorpromazine (CPZ) and U18666A on the intracellular dynamics of a cellular isoform of prion protein (PrPC) and PrPSc in prion-infected mouse neuroblastoma cells to re-evaluate the effects of those agents. MAb 44B1 and PPS rapidly reduced PrPSc levels without altering intracellular distribution of PrPSc. PPS did not change the distribution and levels of PrPC, whereas mAb 44B1 appeared to inhibit the trafficking of cell surface PrPC to organelles in the endocytic-recycling pathway that are thought to be one of the sites for PrPSc formation. In contrast, CPZ and U18666A initiated the redistribution of PrPSc from organelles in the endocytic-recycling pathway to late endosomes/lysosomes without apparent changes in the distribution of PrPC. The inhibition of lysosomal function by monensin or bafilomycin A1 after the occurrence of PrPSc redistribution by CPZ or U18666A partly antagonized PrPSc degradation, suggesting that the transfer of PrPSc to late endosomes/lysosomes, possibly via alteration of the membrane trafficking machinery of cells, leads to PrPSc degradation. This study revealed that precise analysis of the intracellular dynamics of PrPC and PrPSc provides important information for understanding the mechanism of anti-prion agents. 相似文献
4.
The causative agent of prion diseases is the pathological isoform (PrPSc) of the host-encoded cellular prion protein (PrPC). PrPSc has an identical amino acid sequence to PrPC; thus, it has been assumed that an immune response against PrPSc could not be found in prion-affected animals. In this study, we found the anti-prion protein (PrP) antibody at the terminal stage of mouse scrapie. Several sera from mice in the terminal stage of scrapie reacted to the recombinant mouse PrP (rMPrP) molecules and brain homogenates of mouse prion diseases. These results indicate that mouse could recognize PrPC or PrPSc as antigens by the host immune system. Furthermore, immunization with rMPrP generates high titers of anti-PrP antibodies in wild-type mice. Some anti-PrP antibodies immunized with rMPrP prevent PrPSc replication in vitro. The mouse sera from terminal prion disease have several wide epitopes, although mouse sera immunized with rMPrP possess narrow epitopes. 相似文献
5.
Prion diseases are a heterogeneous class of fatal neurodegenerative disorders associated with misfolding of host cellular prion protein (PrPC) into a pathological isoform, termed PrPSc. Prion diseases affect various mammals, including humans, and effective treatments are not available. Prion diseases are distinguished from other protein misfolding disorders – such as Alzheimer’s or Parkinson’s disease – in that they are infectious. Prion diseases occur sporadically without any known exposure to infected material, and hereditary cases resulting from rare mutations in the prion protein have also been documented. The mechanistic underpinnings of prion and other neurodegenerative disorders remain poorly understood. Various proteomics techniques have been instrumental in early PrPSc detection, biomarker discovery, elucidation of PrPSc structure and mapping of biochemical pathways affected by pathogenesis. Moving forward, proteomics approaches will likely become more integrated into the clinical and research settings for the rapid diagnosis and characterization of prion pathogenesis. 相似文献
6.
Roger A. Moore Mark W. Head James W. Ironside Diane L. Ritchie Gianluigi Zanusso Young Pyo Choi Suzette A. Priola 《PLoS pathogens》2016,12(2)
Sporadic Creutzfeldt-Jakob disease (sCJD) is the most prevalent of the human prion diseases, which are fatal and transmissible neurodegenerative diseases caused by the infectious prion protein (PrPSc). The origin of sCJD is unknown, although the initiating event is thought to be the stochastic misfolding of endogenous prion protein (PrPC) into infectious PrPSc. By contrast, human growth hormone-associated cases of iatrogenic CJD (iCJD) in the United Kingdom (UK) are associated with exposure to an exogenous source of PrPSc. In both forms of CJD, heterozygosity at residue 129 for methionine (M) or valine (V) in the prion protein gene may affect disease phenotype, onset and progression. However, the relative contribution of each PrPC allotype to PrPSc in heterozygous cases of CJD is unknown. Using mass spectrometry, we determined that the relative abundance of PrPSc with M or V at residue 129 in brain specimens from MV cases of sCJD was highly variable. This result is consistent with PrPC containing an M or V at residue 129 having a similar propensity to misfold into PrPSc thus causing sCJD. By contrast, PrPSc with V at residue 129 predominated in the majority of the UK human growth hormone associated iCJD cases, consistent with exposure to infectious PrPSc containing V at residue 129. In both types of CJD, the PrPSc allotype ratio had no correlation with CJD type, age at clinical onset, or disease duration. Therefore, factors other than PrPSc allotype abundance must influence the clinical progression and phenotype of heterozygous cases of CJD. 相似文献
7.
The prion diseases occur following the conversion of the cellular prion protein (PrPC) into disease-related isoforms (PrPSc). In this study, the role of the glycosylphosphatidylinositol (GPI) anchor attached to PrPC in prion formation was examined using a cell painting technique. PrPSc formation in two prion-infected neuronal cell lines (ScGT1 and ScN2a cells) and in scrapie-infected primary cortical neurons was increased following the introduction of PrPC. In contrast, PrPC containing a GPI anchor from which the sialic acid had been removed (desialylated PrPC) was not converted to PrPSc. Furthermore, the presence of desialylated PrPC inhibited the production of PrPSc within prion-infected cortical neurons and ScGT1 and ScN2a cells. The membrane rafts surrounding desialylated PrPC contained greater amounts of sialylated gangliosides and cholesterol than membrane rafts surrounding PrPC. Desialylated PrPC was less sensitive to cholesterol depletion than PrPC and was not released from cells by treatment with glimepiride. The presence of desialylated PrPC in neurons caused the dissociation of cytoplasmic phospholipase A2 from PrP-containing membrane rafts and reduced the activation of cytoplasmic phospholipase A2. These findings show that the sialic acid moiety of the GPI attached to PrPC modifies local membrane microenvironments that are important in PrP-mediated cell signaling and PrPSc formation. These results suggest that pharmacological modification of GPI glycosylation might constitute a novel therapeutic approach to prion diseases. 相似文献
8.
Prion diseases are fatal, neurodegenerative disorders in humans and animals and are characterized by the accumulation of an abnormally folded isoform of the cellular prion protein (PrPC), denoted PrPSc, which represents the major component of infectious scrapie prions. Characterization of the mechanism of conversion of PrPC into PrPSc and identification of the intracellular site where it occurs are among the most important questions in prion biology. Despite numerous efforts, both of these questions remain unsolved. We have quantitatively analyzed the distribution of PrPC and PrPSc and measured PrPSc levels in different infected neuronal cell lines in which protein trafficking has been selectively impaired. Our data exclude roles for both early and late endosomes and identify the endosomal recycling compartment as the likely site of prion conversion. These findings represent a fundamental step towards understanding the cellular mechanism of prion conversion and will allow the development of new therapeutic approaches for prion diseases. 相似文献
9.
10.
Christopher J Silva 《朊病毒》2014,8(1):42-50
Prions are molecular pathogens, able to convert a normal cellular prion protein (PrPC) into a prion (PrPSc). The information necessary for this conversion is contained in the conformation of PrPSc. Mass spectrometry (MS) and small-molecule covalent reactions have been used to study prions. Mass spectrometry has been used to detect and quantitate prions in the attomole range (10?18 mole). MS-based analysis showed that both possess identical amino acid sequences, one disulfide bond, a GPI anchor, asparagine-linked sugar antennae, and unoxidized methionines. Mass spectrometry has been used to define elements of the secondary and tertiary structure of wild-type PrPSc and GPI-anchorless PrPSc. It has also been used to study the quaternary structure of the PrPSc multimer. Small molecule reagents react differently with the same lysine in the PrPC conformation than in the PrPSc conformation. Such differences can be detected by Western blot using mAbs with lysine-containing epitopes, such as 3F4 and 6D11. This permits the detection of PrPSc without the need for proteinase K pretreatment and can be used to distinguish among prion strains. These results illustrate how two important chemical tools, mass spectrometry and covalent modification by small molecules, are being applied to the detection and structural study of prions. Furthermore these tools are or can be applied to the study of the other protein misfolding diseases such as Alzheimer Disease, Parkinson Disease, or ALS. 相似文献
11.
Fabio Moda Chiara Vimercati Ilaria Campagnani Margherita Ruggerone Giorgio Giaccone Michela Morbin Lorena Zentilin Mauro Giacca Ileana Zucca Giuseppe Legname Fabrizio Tagliavini 《朊病毒》2012,6(4):383-390
Prion diseases are caused by a conformational modification of the cellular prion protein (PrPC) into disease-specific forms, termed PrPSc, that have the ability to interact with PrPC promoting its conversion to PrPSc. In vitro studies demonstrated that anti-PrP antibodies inhibit this process. In particular, the single chain variable fragment D18 antibody (scFvD18) showed high efficiency in curing chronically prion-infected cells. This molecule binds the PrPC region involved in the interaction with PrPSc thus halting further prion formation. These findings prompted us to test the efficiency of scFvD18 in vivo. A recombinant Adeno-Associated Viral vector serotype 9 was used to deliver scFvD18 to the brain of mice that were subsequently infected by intraperitoneal route with the mouse-adapted scrapie strain RML. We found that the treatment was safe, prolonged the incubation time of scrapie-infected animals and decreased the burden of total proteinase-resistant PrPSc in the brain, suggesting that scFvD18 interferes with prion replication in vivo. This approach is relevant for designing new therapeutic strategies for prion diseases and other disorders characterized by protein misfolding. 相似文献
12.
《朊病毒》2013,7(4):383-390
Prion diseases are caused by a conformational modification of the cellular prion protein (PrPC) into disease-specific forms, termed PrPSc, that have the ability to interact with PrPC promoting its conversion to PrPSc. In vitro studies demonstrated that anti-PrP antibodies inhibit this process. In particular, the single chain variable fragment D18 antibody (scFvD18) showed high efficiency in curing chronically prion-infected cells. This molecule binds the PrPC region involved in the interaction with PrPSc thus halting further prion formation. These findings prompted us to test the efficiency of scFvD18 in vivo. A recombinant Adeno-Associated Viral vector serotype 9 was used to deliver scFvD18 to the brain of mice that were subsequently infected by intraperitoneal route with the mouse-adapted scrapie strain RML. We found that the treatment was safe, prolonged the incubation time of scrapie-infected animals and decreased the burden of total proteinase-resistant PrPSc in the brain, suggesting that scFvD18 interferes with prion replication in vivo. This approach is relevant for designing new therapeutic strategies for prion diseases and other disorders characterized by protein misfolding. 相似文献
13.
Background
A hallmark of the prion diseases is the conversion of the host-encoded cellular prion protein (PrPC) into a disease related, alternatively folded isoform (PrPSc). The accumulation of PrPSc within the brain is associated with synapse loss and ultimately neuronal death. Novel therapeutics are desperately required to treat neurodegenerative diseases including the prion diseases.Principal Findings
Treatment with glimepiride, a sulphonylurea approved for the treatment of diabetes mellitus, induced the release of PrPC from the surface of prion-infected neuronal cells. The cell surface is a site where PrPC molecules may be converted to PrPSc and glimepiride treatment reduced PrPSc formation in three prion infected neuronal cell lines (ScN2a, SMB and ScGT1 cells). Glimepiride also protected cortical and hippocampal neurones against the toxic effects of the prion-derived peptide PrP82–146. Glimepiride treatment significantly reduce both the amount of PrP82–146 that bound to neurones and PrP82–146 induced activation of cytoplasmic phospholipase A2 (cPLA2) and the production of prostaglandin E2 that is associated with neuronal injury in prion diseases. Our results are consistent with reports that glimepiride activates an endogenous glycosylphosphatidylinositol (GPI)-phospholipase C which reduced PrPC expression at the surface of neuronal cells. The effects of glimepiride were reproduced by treatment of cells with phosphatidylinositol-phospholipase C (PI-PLC) and were reversed by co-incubation with p-chloromercuriphenylsulphonate, an inhibitor of endogenous GPI-PLC.Conclusions
Collectively, these results indicate that glimepiride may be a novel treatment to reduce PrPSc formation and neuronal damage in prion diseases. 相似文献14.
The soluble cellular prion protein (PrPC) is best known for its association with prion disease (PrD) through its conversion to a pathogenic insoluble isoform (PrPSc). However, its deleterious effects independent of PrPSc have recently been observed not only in PrD but also in Alzheimer disease (AD), two diseases which mainly affect cognition. At the same time, PrPC itself seems to have broad physiologic functions including involvement in cognitive processes. The PrPC that is believed to be soluble and monomeric has so far been the only PrP conformer observed in the uninfected brain. In 2006, we identified an insoluble PrPC conformer (termed iPrPC) in uninfected human and animal brains. Remarkably, the PrPSc-like iPrPC shares the immunoreactivity behavior and fragmentation with a newly-identified PrPSc species in a novel human PrD termed variably protease-sensitive prionopathy. Moreover, iPrPC has been observed as the major PrP species that interacts with amyloid β (Aβ) in AD. This article highlights evidence of PrP involvement in two putatively beneficial and deleterious PrP-implicated pathways in cognition and hypothesizes first, that beneficial and deleterious effects of PrPC are attributable to the chameleon-like conformation of the protein and second, that the iPrPC conformer is associated with PrD and AD.Key words: prion protein, prion disease, cognition, cognitive deficit, insoluble prion protein, Alzheimer disease, variably protease-sensitive prionopathy, dementia, memory 相似文献
15.
Magdalini Polymenidou Rita Moos Mike Scott Christina Sigurdson Yong-zhong Shi Bill Yajima Iva Hafner-Bratkovi? Roman Jerala Simone Hornemann Kurt Wuthrich Anne Bellon Martin Vey Graciela Garen Michael N. G. James Nat Kav Adriano Aguzzi 《PloS one》2008,3(12)
PrPSc, a misfolded and aggregated form of the cellular prion protein PrPC, is the only defined constituent of the transmissible agent causing prion diseases. Expression of PrPC in the host organism is necessary for prion replication and for prion neurotoxicity. Understanding prion diseases necessitates detailed structural insights into PrPC and PrPSc. Towards this goal, we have developed a comprehensive collection of monoclonal antibodies denoted POM1 to POM19 and directed against many different epitopes of mouse PrPC. Three epitopes are located within the N-terminal octarepeat region, one is situated within the central unstructured region, and four epitopes are discontinuous within the globular C-proximal domain of PrPC. Some of these antibodies recognize epitopes that are resilient to protease digestion in PrPSc. Other antibodies immunoprecipitate PrPC, but not PrPSc. A third group was found to immunoprecipitate both PrP isoforms. Some of the latter antibodies could be blocked with epitope-mimicking peptides, and incubation with an excess of these peptides allowed for immunochromatography of PrPC and PrPSc. Amino-proximal antibodies were found to react with repetitive PrPC epitopes, thereby vastly increasing their avidity. We have also created functional single-chain miniantibodies from selected POMs, which retained the binding characteristics despite their low molecular mass. The POM collection, thus, represents a unique set of reagents allowing for studies with a variety of techniques, including western blotting, ELISA, immunoprecipitation, conformation-dependent immunoassays, and plasmon surface plasmon resonance-based assays. 相似文献
16.
Sakudo A Wu G Onodera T Ikuta K 《Biochemical and biophysical research communications》2008,365(1):164-169
An abnormal isoform of prion protein (PrPSc), which is composed of the same amino acids as cellular PrP (PrPC) and has proteinase K (PK)-resistance, hypothetically converts PrPC into PrPSc. To investigate the region important for PrPSc production, we examined the levels of PrPSc in PrP gene-deficient cells (HpL3-4) expressing PrPC deleted of various regions including the octapeptide repeat region (OR) or hydrophobic region (HR). After Chandler or Obihiro prion infection, PrPSc was produced in HpL3-4 cells expressing wild-type PrPC or PrPC deleted of HR at an early stage and further reduced to below the detectable level, whereas cells expressing PrPC deleted of OR showed no PrPSc production. The results suggest that OR of PrPC is required for the early step of efficient PrPSc production. 相似文献
17.
Tsukasa Mashima Fumiko Nishikawa Yuji O. Kamatari Hiromichi Fujiwara Masayuki Saimura Takashi Nagata Tsutomu Kodaki Satoshi Nishikawa Kazuo Kuwata Masato Katahira 《Nucleic acids research》2013,41(2):1355-1362
Prion proteins (PrPs) cause prion diseases, such as bovine spongiform encephalopathy. The conversion of a normal cellular form (PrPC) of PrP into an abnormal form (PrPSc) is thought to be associated with the pathogenesis. An RNA aptamer that tightly binds to and stabilizes PrPC is expected to block this conversion and to thereby prevent prion diseases. Here, we show that an RNA aptamer comprising only 12 residues, r(GGAGGAGGAGGA) (R12), reduces the PrPSc level in mouse neuronal cells persistently infected with the transmissible spongiform encephalopathy agent. Nuclear magnetic resonance analysis revealed that R12, folded into a unique quadruplex structure, forms a dimer and that each monomer simultaneously binds to two portions of the N-terminal half of PrPC, resulting in tight binding. Electrostatic and stacking interactions contribute to the affinity of each portion. Our results demonstrate the therapeutic potential of an RNA aptamer as to prion diseases. 相似文献
18.
Yuko Ushiki‐Kaku Yoshifumi Iwamaru Kentaro Masujin Morikazu Imamura Shigeyoshi Itohara Kiyoko Ogawa‐Goto Shunji Hattori Takashi Yokoyama 《Microbiology and immunology》2013,57(11):792-796
Limited information is available about conformational differences between the abnormal isoform of prion protein (PrPSc) and cellular prion protein (PrPC) under native conditions. To clarify conformational differences between these two isoforms, PrP‐deficient mice were immunized with brain homogenates of normal and scrapie‐infected animals. All mice generated anti‐PrP antibodies. Peptide array analysis of these serum samples revealed a distinctive epitope of PrPSc consisting of QGSPGGN (PrP41–47) at the N‐terminus. This study demonstrated a conformational dissimilarity at the N‐terminus between PrPSc and PrPC, a finding that may provide novel information about conformational features of PrPSc. 相似文献
19.
Ryan Taschuk Kristen Marciniuk Pekka Määttänen Claudia Madampage Peter Hedlin Andrew Potter 《朊病毒》2014,8(1):51-59
Prions are a novel form of infectivity based on the misfolding of a self-protein (PrPC) into a pathological, infectious isomer (PrPSc). The current uncontrolled spread of chronic wasting disease in cervids, coupled with the demonstrated zoonotic nature of select livestock prion diseases, highlights the urgent need for disease management tools. While there is proof-of-principle evidence for a prion vaccine, these efforts are complicated by the challenges and risks associated with induction of immune responses to a self-protein. Our priority is to develop a PrPSc-specific prion vaccine based on epitopes that are uniquely exposed upon misfolding. These disease specific epitopes (DSEs) have the potential to enable specific targeting of the pathological species through immunotherapy. Here we review outcomes of the translation of a prion DSE into a PrPSc-specific vaccine based on the criteria of immunogenicity, safety and specificity. 相似文献
20.