scholarly journals The Cellular Prion Protein Increases the Uptake and Toxicity of TDP-43 Fibrils

Viruses ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1625
Author(s):  
Carlo Scialò ◽  
Luigi Celauro ◽  
Marco Zattoni ◽  
Thanh Hoa Tran ◽  
Edoardo Bistaffa ◽  
...  
Keyword(s):  
Prion Protein ◽  
Neuroblastoma Cells ◽  
Cellular Prion Protein ◽  
Cellular Systems ◽  
Human Form ◽  
Downstream Effector ◽  
Common Receptor ◽  
Overexpressing Cell ◽  
Almost All ◽  
In Cells

Cytoplasmic aggregation of the primarily nuclear TAR DNA-binding protein 43 (TDP-43) affects neurons in most amyotrophic lateral sclerosis (ALS) and approximately half of frontotemporal lobar degeneration (FTLD) cases. The cellular prion protein, PrPC, has been recognized as a common receptor and downstream effector of circulating neurotoxic species of several proteins involved in neurodegeneration. Here, capitalizing on our recently adapted TDP-43 real time quaking induced reaction, we set reproducible protocols to obtain standardized preparations of recombinant TDP-43 fibrils. We then exploited two different cellular systems (human SH-SY5Y and mouse N2a neuroblastoma cells) engineered to express low or high PrPC levels to investigate the link between PrPC expression on the cell surface and the internalization of TDP-43 fibrils. Fibril uptake was increased in cells overexpressing either human or mouse prion protein. Increased internalization was associated with detrimental consequences in all PrP-overexpressing cell lines but was milder in cells expressing the human form of the prion protein. As described for other amyloids, treatment with TDP-43 fibrils induced a reduction in the accumulation of the misfolded form of PrPC, PrPSc, in cells chronically infected with prions. Our results expand the list of misfolded proteins whose uptake and detrimental effects are mediated by PrPC, which encompass almost all pathological amyloids involved in neurodegeneration.

scholarly journals Dimerization of the cellular prion protein inhibits propagation of scrapie prions

2018 ◽  
Vol 293 (21) ◽  
pp. 8020-8031 ◽  
Author(s):  
Anna D. Engelke ◽  
Anika Gonsberg ◽  
Simrika Thapa ◽  
Sebastian Jung ◽  
Sarah Ulbrich ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Prion Protein ◽  
Conformational Transition ◽  
Prion Diseases ◽  
Neuroblastoma Cells ◽  
Direct Interaction ◽  
Cellular Prion Protein ◽  
Dominant Negative ◽  
Dimerization Domain ◽  
Hydrophobic Domain

A central step in the pathogenesis of prion diseases is the conformational transition of the cellular prion protein (PrPC) into the scrapie isoform, denoted PrPSc. Studies in transgenic mice have indicated that this conversion requires a direct interaction between PrPC and PrPSc; however, insights into the underlying mechanisms are still missing. Interestingly, only a subfraction of PrPC is converted in scrapie-infected cells, suggesting that not all PrPC species are suitable substrates for the conversion. On the basis of the observation that PrPC can form homodimers under physiological conditions with the internal hydrophobic domain (HD) serving as a putative dimerization domain, we wondered whether PrP dimerization is involved in the formation of neurotoxic and/or infectious PrP conformers. Here, we analyzed the possible impact on dimerization of pathogenic mutations in the HD that induce a spontaneous neurodegenerative disease in transgenic mice. Similarly to wildtype (WT) PrPC, the neurotoxic variant PrP(AV3) formed homodimers as well as heterodimers with WTPrPC. Notably, forced PrP dimerization via an intermolecular disulfide bond did not interfere with its maturation and intracellular trafficking. Covalently linked PrP dimers were complex glycosylated, GPI-anchored, and sorted to the outer leaflet of the plasma membrane. However, forced PrPC dimerization completely blocked its conversion into PrPSc in chronically scrapie-infected mouse neuroblastoma cells. Moreover, PrPC dimers had a dominant-negative inhibition effect on the conversion of monomeric PrPC. Our findings suggest that PrPC monomers are the major substrates for PrPSc propagation and that it may be possible to halt prion formation by stabilizing PrPC dimers.

Domain-Specific Activation of Death-Associated Intracellular Signalling Cascades by the Cellular Prion Protein in Neuroblastoma Cells

2015 ◽  
Vol 53 (7) ◽  
pp. 4438-4448 ◽  
Author(s):  
Silvia Vilches ◽  
Cristina Vergara ◽  
Oriol Nicolás ◽  
Ágata Mata ◽  
José A. del Río ◽  
...  
Keyword(s):  
Prion Protein ◽  
Neuroblastoma Cells ◽  
Cellular Prion Protein ◽  
Intracellular Signalling ◽  
Domain Specific ◽  
Signalling Cascades

scholarly journals The uptake of tau amyloid fibrils is facilitated by the cellular prion protein and hampers prion propagation in cultured cells

2020 ◽  
Author(s):  
Elena De Cecco ◽  
Luigi Celauro ◽  
Silvia Vanni ◽  
Micaela Grandolfo ◽  
Adriano Aguzzi ◽  
...  
Keyword(s):  
Prion Protein ◽  
Amyloid Fibrils ◽  
Cultured Cells ◽  
Neuroblastoma Cells ◽  
Cellular Prion Protein ◽  
Brain Diseases ◽  
Gene Encoding ◽  
Prion Propagation ◽  
Prion Conversion ◽  
Tau Fibrils

AbstractTauopathies are prevalent, invariably fatal brain diseases for which no cure is available. Tauopathies progressively affect the brain through cell-to-cell transfer of tau protein amyloids, yet the spreading mechanisms are unknown. Here we show that the cellular prion protein (PrPC) facilitates the uptake of tau aggregates by cultured cells, possibly by acting as an endocytic receptor. In mouse neuroblastoma cells, we found that tau amyloids bind to PrPC; internalization of tau fibrils was reduced in isogenic cells devoid of the gene encoding PrPC. Antibodies against N-proximal epitopes of PrPC impaired the binding of tau amyloids and decreased their uptake. Surprisingly, exposure of chronically prion-infected cells to tau amyloids reduced the accumulation of aggregated prion protein; this effect lasted for more than 72 hours after amyloid removal. These results point to bidirectional interactions between the two proteins: whilst PrPC mediates the entrance of tau fibrils in cells, PrPSc buildup is greatly reduced in their presence, possibly because of an impairment in the prion conversion process.

scholarly journals The cellular prion protein interacts with and promotes the activity of Na,K-ATPases

PLoS ONE ◽  
2021 ◽  
Vol 16 (11) ◽  
pp. e0258682
Author(s):  
Declan Williams ◽  
Mohadeseh Mehrabian ◽  
Hamza Arshad ◽  
Shehab Eid ◽  
Christopher Sackmann ◽  
...  
Keyword(s):  
Prion Protein ◽  
Prion Diseases ◽  
Protein A ◽  
Neuroblastoma Cells ◽  
Cellular Prion Protein ◽  
Affinity Capture ◽  
Endoproteolytic Cleavage ◽  
Relative Quantitation ◽  
Steady State Levels ◽  
Uptake Activity

The prion protein (PrP) is best known for its ability to cause fatal neurodegenerative diseases in humans and animals. Here, we revisited its molecular environment in the brain using a well-developed affinity-capture mass spectrometry workflow that offers robust relative quantitation. The analysis confirmed many previously reported interactions. It also pointed toward a profound enrichment of Na,K-ATPases (NKAs) in proximity to cellular PrP (PrPC). Follow-on work validated the interaction, demonstrated partial co-localization of the ATP1A1 and PrPC, and revealed that cells exposed to cardiac glycoside (CG) inhibitors of NKAs exhibit correlated changes to the steady-state levels of both proteins. Moreover, the presence of PrPC was observed to promote the ion uptake activity of NKAs in a human co-culture paradigm of differentiated neurons and glia cells, and in mouse neuroblastoma cells. Consistent with this finding, changes in the expression of 5’-nucleotidase that manifest in wild-type cells in response to CG exposure can also be observed in untreated PrPC-deficient cells. Finally, the endoproteolytic cleavage of the glial fibrillary acidic protein, a hallmark of late-stage prion disease, can also be induced by CGs, raising the prospect that a loss of NKA activity may contribute to the pathobiology of prion diseases.

scholarly journals Toxicological Evaluation of Anti-Scrapie Trimethoxychalcones and Oxadiazoles

2015 ◽  
Vol 87 (2 suppl) ◽  
pp. 1421-1434 ◽  
Author(s):  
CLAUDIA P. FIGUEIREDO ◽  
NATALIA C. FERREIRA ◽  
GISELLE F. PASSOS ◽  
ROBSON DA COSTA ◽  
FERNANDA S. NEVES ◽  
...  
Keyword(s):  
Prion Protein ◽  
Aromatic Compounds ◽  
Prion Diseases ◽  
Intraperitoneal Administration ◽  
Neuroblastoma Cells ◽  
Cellular Prion Protein ◽  
Transmissible Spongiform Encephalopathies ◽  
Toxicological Evaluation ◽  
Spongiform Encephalopathies

An altered form of the cellular prion protein, the PrPScor PrPRes, is implicated in the occurrence of the still untreatable transmissible spongiform encephalopathies. We have previously synthesized and characterized aromatic compounds that inhibit protease-resistant prion protein (PrPRes) accumulation in scrapie-infected cells. These compounds belong to different chemical classes, including acylhydrazones, chalcones and oxadiazoles. Some of the active compounds were non-toxic to neuroblastoma cells in culture and seem to possess drugable properties, since they are in agreement with the Lipinski´s rule of 5 and present desirable pharmacokinetic profiles as predicted in silico. Before the evaluation of the in vivo efficacy of the aromatic compounds in scrapie-infected mice, safety assessment in healthy mice is needed. Here we used Swiss mice to evaluate the acute toxicity profile of the six most promising anti-prionic compounds, the 2,4,5-trimethoxychalcones (J1, J8, J20 and J35) and the 1,3,4-oxadiazoles (Y13 and Y17). One single oral administration (300 mg/kg) of J1, J8, J20, J35, Y13 and Y17 or repeated intraperitoneal administration (10 mg/kg, 3 times a week, for 4 weeks) of J1, J8 and J35, did not elicit toxicity in mice. We strongly believe that the investigated trimethoxychalcones and oxadiazoles are interesting compounds to be further analyzed in vivo against prion diseases.

scholarly journals Neurotropic influenza A virus infection causes prion protein misfolding into infectious prions in neuroblastoma cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Hideyuki Hara ◽  
Junji Chida ◽  
Keiji Uchiyama ◽  
Agriani Dini Pasiana ◽  
Etsuhisa Takahashi ◽  
...  
Keyword(s):  
Prion Protein ◽  
Influenza A Virus ◽  
Protein Misfolding ◽  
Influenza A ◽  
Prion Diseases ◽  
Neuroblastoma Cells ◽  
Cellular Prion Protein ◽  
Infected Cells ◽  
Hit And Run

AbstractMisfolding of the cellular prion protein, PrPC, into the amyloidogenic isoform, PrPSc, which forms infectious protein aggregates, the so-called prions, is a key pathogenic event in prion diseases. No pathogens other than prions have been identified to induce misfolding of PrPC into PrPSc and propagate infectious prions in infected cells. Here, we found that infection with a neurotropic influenza A virus strain (IAV/WSN) caused misfolding of PrPC into PrPSc and generated infectious prions in mouse neuroblastoma cells through a hit-and-run mechanism. The structural and biochemical characteristics of IAV/WSN-induced PrPSc were different from those of RML and 22L laboratory prions-evoked PrPSc, and the pathogenicity of IAV/WSN-induced prions were also different from that of RML and 22L prions, suggesting IAV/WSN-specific formation of PrPSc and infectious prions. Our current results may open a new avenue for the role of viral infection in misfolding of PrPC into PrPSc and formation of infectious prions.

scholarly journals Cell-surface retention of PrPC by anti-PrP antibody prevents protease-resistant PrP formation

2004 ◽  
Vol 85 (11) ◽  
pp. 3473-3482 ◽  
Author(s):  
Chan-Lan Kim ◽  
Ayako Karino ◽  
Naotaka Ishiguro ◽  
Morikazu Shinagawa ◽  
Motoyoshi Sato ◽  
...  
Keyword(s):  
Cell Surface ◽  
Prion Protein ◽  
Flow Cytometric Analysis ◽  
Degradation Pathway ◽  
Surface Flow ◽  
Cellular Prion Protein ◽  
Terminal Portion ◽  
Persistently Infected ◽  
Prion Propagation ◽  
In Cells

The C-terminal portion of the prion protein (PrP), corresponding to a protease-resistant core fragment of the abnormal isoform of the prion protein (PrPSc), is essential for prion propagation. Antibodies to the C-terminal portion of PrP are known to inhibit PrPSc accumulation in cells persistently infected with prions. Here it was shown that, in addition to monoclonal antibodies (mAbs) to the C-terminal portion of PrP, a mAb recognizing the octapeptide repeat region in the N-terminal part of PrP that is dispensable for PrPSc formation reduced PrPSc accumulation in cells persistently infected with prions. The 50 % effective dose was as low as ∼1 nM, and, regardless of their epitope specificity, the inhibitory mAbs shared the ability to bind cellular prion protein (PrPC) expressed on the cell surface. Flow cytometric analysis revealed that mAbs that bound to the cell surface during cell culture were not internalized even after their withdrawal from the growth medium. Retention of the mAb–PrPC complex on the cell surface was also confirmed by the fact that internalization was enhanced by treatment of cells with dextran sulfate. These results suggested that anti-PrP mAb antagonizes PrPSc formation by interfering with the regular PrPC degradation pathway.

scholarly journals Cellular prion protein mediates early apoptotic proteome alternation and phospho-modification in human neuroblastoma cells

2017 ◽  
Vol 8 (1) ◽  
pp. e2557-e2557 ◽  
Author(s):  
Saima Zafar ◽  
Christina Behrens ◽  
Hassan Dihazi ◽  
Matthias Schmitz ◽  
Inga Zerr ◽  
...  
Keyword(s):  
Prion Protein ◽  
Neuroblastoma Cells ◽  
Cellular Prion Protein ◽  
Human Neuroblastoma Cells ◽  
Human Neuroblastoma

scholarly journals Global analysis of protein degradation in prion infected cells

2019 ◽  
Author(s):  
Charles Hutti ◽  
Kevin A. Welle ◽  
Jennifer R. Hryhorenko ◽  
Sina Ghaemmaghami
Keyword(s):  
Protein Degradation ◽  
Prion Protein ◽  
Degradation Kinetics ◽  
Global Analysis ◽  
Neuroblastoma Cells ◽  
Cellular Prion Protein ◽  
Lysosomal Hydrolases ◽  
Prion Infection ◽  
Degradation Rates ◽  
Infected Cells

ABSTRACTPrion diseases are rare neurological disorders caused by the misfolding of the cellular prion protein (PrPC) into cytotoxic fibrils (PrPSc). Intracellular PrPSc aggregates primarily accumulate within late endosomes and lysosomes, organelles that participate in the degradation and turnover of a large subset of the proteome. Thus, intracellular accumulation of PrPSc aggregates have the potential to globally influence protein degradation kinetics within an infected cell. We analyzed the proteome-wide effect of prion infection on protein degradation rates in N2a neuroblastoma cells by dynamic stable isotopic labeling with amino acids in cell culture (dSILAC) and bottom-up proteomics. The analysis quantified the degradation rates of more than 4,700 proteins in prion-infected and uninfected cells. As expected, the degradation rate of the prion protein is significantly decreased upon aggregation in infected cells. In contrast, the degradation kinetics of the remainder of the N2a proteome generally increases upon prion infection. This effect occurs concurrently with increases in the cellular activities of autophagy and lysosomal hydrolases. The resulting enhancement in proteome flux may play a role in the survival of N2a cells upon prion infection.

scholarly journals A glycolipid-anchored prion protein is endocytosed via clathrin-coated pits.

1994 ◽  
Vol 125 (6) ◽  
pp. 1239-1250 ◽  
Author(s):  
S L Shyng ◽  
J E Heuser ◽  
D A Harris
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Prion Protein ◽  
Transmembrane Protein ◽  
Primary Cultures ◽  
Neuroblastoma Cells ◽  
Cellular Prion Protein ◽  
Coated Vesicles ◽  
Coated Pits ◽  
Hypertonic Medium

The cellular prion protein (PrPc) is a glycolipid-anchored, cell surface protein of unknown function, a posttranslationally modified isoform of which PrPSc is involved in the pathogenesis of Creutzfeldt-Jakob disease, scrapie, and other spongiform encephalopathies. We have shown previously that chPrP, a chicken homologue of mammalian PrPC, constitutively cycles between the cell surface and an endocytic compartment, with a transit time of approximately 60 min in cultured neuroblastoma cells. We now report that endocytosis of chPrP is mediated by clathrin-coated pits. Immunogold labeling of neuroblastoma cells demonstrates that the concentration of chPrP within 0.05 microns of coated pits is 3-5 times higher than over other areas of the plasma membrane. Moreover, gold particles can be seen within coated vesicles and deeply invaginated coated pits that are in the process of pinching off from the plasma membrane. ChPrP is also localized to coated pits in primary cultures of neurons and glia, and is found in coated vesicles purified from chicken brain. Finally, internalization of chPrP is reduced by 70% after neuroblastoma cells are incubated in hypertonic medium, a treatment that inhibits endocytosis by disrupting clathrin lattices. Caveolae, plasmalemmal invaginations in which several other glycolipid-anchored proteins are concentrated, are not seen in neuroblastoma cells analyzed by thin-section or deep-etch electron microscopy. Moreover, these cells do not express detectable levels of caveolin, a caveolar coat protein. Since chPrP lacks a cytoplasmic domain that could interact directly with the intracellular components of clathrin-coated pits, we propose that the polypeptide chain of chPrP associates with the extracellular domain of a transmembrane protein that contains a coated pit internalization signal.

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