Cellular prion protein offers neuroprotection in astrocytes submitted to amyloid ß oligomer toxicity.

The cellular prion protein (PrPC), in its native conformation, performs numerous cellular and cognitive functions in brain tissue. However, despite the cellular prion research in recent years, there are still questions about its participation in oxidative and neurodegenerative processes. This study aims to elucidate the involvement of PrPC in the neuroprotection cascade in the presence of oxidative stressors. For that, astrocytes from wild-type mice and knockout to PrPC were subjected to the induction of oxidative stress with hydrogen peroxide (H2O2) and with the toxic oligomer of the amyloid ß protein (AßO). We observed that the presence of PrPC showed resistance in the cell viability of astrocytes. It was also possible to monitor changes in basic levels of metals and associate them with an induced damage condition, indicating the precise role of PrPC in metal homeostasis, where the absence of PrPC leads to metallic unbalance, culminating in cellular vulnerability to oxidative stress. Increased caspase 3, p-Tau, p53, and Bcl2 may establish a relationship between a PrPC and an induced damage condition. Complementarily, it has been shown that PrPC prevents the internalization of AßO and promotes its degradation under oxidative stress induction, thus preventing protein aggregation in astrocytes. It was also observed that the presence of PrPC can be related to translocating SOD1 to cell nuclei under oxidative stress, probably controlling DNA damage. The results of this study suggest that PrPC acts against oxidative stress activating the cellular response and defense by displaying neuroprotection to neurons and ensuring the functionality of astrocytes.

The multiple functions of PrPC in physiological, cancer, and neurodegenerative contexts.

Cellular prion protein (PrPC) is a highly conserved glycoprotein, present both anchored in the cell membrane and soluble in the extracellular medium. It has a diversity of ligands and is variably expressed in numerous tissues and cell subtypes, most notably in the central nervous system (CNS). Its importance has been brought to light over the years both under physiological conditions, such as embryogenesis and immune system homeostasis, and in pathologies, such as cancer and neurodegenerative diseases. During development, PrPC plays an important role in CNS, participating in axonal growth and guidance and differentiation of glial cells, but also in other organs such as the heart, lung, and digestive system. In diseases, PrPC has been related to several types of tumors, modulating cancer stem cells, enhancing malignant properties, and inducing drug resistance. Also, in non-neoplastic diseases, such as Alzheimer's and Parkinson's diseases, PrPC seems to alter the dynamics of neurotoxic aggregate formation and, consequently, the progression of the disease. In this review, we explore in detail the multiple functions of this protein, which proved to be relevant for understanding the dynamics of organism homeostasis, as well as a promising target in the treatment of both neoplastic and degenerative diseases.

Novel quaternary structures of the human prion protein globular domain.

Prion disease is caused by the misfolding of the cellular prion protein, PrPC, into a self-templating conformer, PrPSc. Nuclear magnetic resonance (NMR) and X-ray crystallography revealed the 3D structure of the globular domain of PrPC and the possibility of its dimerization via an interchain disulfide bridge that forms due to domain swap or by non-covalent association of two monomers. On the contrary, PrPSc is composed by a complex and heterogeneous ensemble of poorly defined conformations and quaternary arrangements that are related to different patterns of neurotoxicity. Targeting PrPC with molecules that stabilize the native conformation of its globular domain emerged as a promising approach to develop anti-prion therapies. One of the advantages of this approach is employing structure-based drug discovery methods to PrPC. Thus, it is essential to expand our structural knowledge about PrPC as much as possible to aid such drug discovery efforts. In this work, we report a crystallographic structure of the globular domain of human PrPC that shows a novel dimeric form and a novel oligomeric arrangement. We use molecular dynamics simulations to explore its structural dynamics and stability and discuss potential implications of these new quaternary structures to the conversion process.

Cellular prion protein activates Caspase 3 for apoptotic defense mechanism in astrocytes.

The cellular prion protein (PrPC) is anchored in the plasma membrane of cells, and it is highly present in cells of brain tissue, exerting numerous cellular and cognitive functions. The present study proves the importance of PrPC in the cellular defense mechanism and metal homeostasis in astrocytes cells. Through experimental studies using cell lines of immortalized mice astrocytes (wild type and knockout for PrPC), we showed that PrPc is involved in the apoptosis cell death process by the activation of Caspase 3, downregulation of p53, and cell cycle maintenance. Metal homeostasis was determined by inductively coupled plasma mass spectrometry technique, indicating the crucial role of PrPC to lower intracellular calcium. The lowered calcium concentration and the Caspase 3 downregulation in the PrPC-null astrocytes resulted in a faster growth rate in cells, comparing with PrPC wild-type one. The presence of PrPC shows to be essential to cell death and healthy growth. In conclusion, our results show for the first time that astrocyte knockout cells for the cellular prion protein could modulate apoptosis-dependent cell death pathways.

A New Take on Prion Protein Dynamics in Cellular Trafficking.

The mobility of cellular prion protein (PrPC) in specific cell membrane domains and among distinct cell compartments dictates its molecular interactions and directs its cell function. PrPC works in concert with several partners to organize signaling platforms implicated in various cellular processes. The scaffold property of PrPC is able to gather a molecular repertoire to create heterogeneous membrane domains that favor endocytic events. Dynamic trafficking of PrPC through multiple pathways, in a well-orchestrated mechanism of intra and extracellular vesicular transport, defines its functional plasticity, and also assists the conversion and spreading of its infectious isoform associated with neurodegenerative diseases. In this review, we highlight how PrPC traffics across intra- and extracellular compartments and the consequences of this dynamic transport in governing cell functions and contributing to prion disease pathogenesis.

Sleep deprivation regulates availability of PrPC and Aß peptides which can impair interaction between PrPC and laminin and neuronal plasticity.

PrPC is a glycoprotein capable to interact with several molecules and mediates diverse signaling pathways. Among numerous ligands, laminin (LN) is known to promote neurite outgrowth and memory consolidation, while amyloid-beta oligomers (Aßo) trigger synaptic dysfunction. In both pathways, mGluR1 is recruited as co-receptor. The involvement of PrPC /mGluR1 in these opposite functions suggests that this complex is a key element in the regulation of synaptic activity. Considering that sleep-wake cycle is important for synaptic homeostasis, we aimed to investigate how sleep deprivation affects the expression of PrPC and its ligands, laminin, Aßo, and mGluR1, a multicomplex that can interfere with neuronal plasticity. To address this question, hippocampi of control (CT) and sleep deprived (SD) C57BL/6 mice were collected at two time points of circadian period (13 hr and 21 hr). We observed that sleep deprivation reduced PrPC and mGluR1 levels with higher effect in active state (21 hr). Sleep deprivation also caused accumulation of Aß peptides in rest period (13 hr), while laminin levels were not affected. In vitro binding assay showed that Aßo can compete with LN for PrPC binding. The influence of Aßo was also observed in neuritogenesis. LN alone promoted longer neurite outgrowth than non-treated cells in both Prnp+/+ and Prnp0/0 genotypes. Aßo alone did not show any effects, but when added together with LN, it attenuated the effects of LN only in Prnp+/+ cells. Altogether, our findings indicate that sleep deprivation regulates the availability of PrPC and Aß peptides, and based on our in vitro assays, these alterations induced by sleep deprivation can negatively affect LN-PrPC interaction, which is known to play roles in neuronal plasticity.

Effects of pH and aggregation in the human prion conversion into scrapie form: a study using molecular dynamics with excited normal modes.

There are two different prion conformations: (1) the cellular natural (PrPC) and (2) the scrapie (PrPSc), an infectious form that tends to aggregate under specific conditions. PrPC and PrPSc are widely different regarding secondary and tertiary structures. PrPSc contains more and longer ß-strands compared to PrPC. The lack of solved PrPSc structures precludes a proper understanding of the mechanisms related to the transition between cellular and scrapie forms, as well as the aggregation process. In order to investigate the conformational transition between PrPC and PrPSc, we applied MDeNM (molecular dynamics with excited normal modes), an enhanced sampling simulation technique that has been recently developed to probe large structural changes. These simulations yielded new structural rearrangements of the cellular prion that would have been difficult to obtain with standard MD simulations. We observed an increase in ß-sheet formation under low pH (≤ 4) and upon oligomerization, whose relevance was discussed on the basis of the energy landscape theory for protein folding. The characterization of intermediate structures corresponding to transition states allowed us to propose a conversion model from the cellular to the scrapie prion, which possibly ignites the fibril formation. This model can assist the design of new drugs to prevent neurological disorders related to the prion aggregation mechanism.

Loss of STI1-mediated neuronal survival and differentiation in disease-associated mutations of prion protein.

Cellular prion protein (PrPC ) is widely expressed and displays a variety of well-described functions in the central nervous system (CNS). Mutations of the PRNP gene are known to promote genetic human spongiform encephalopathies, but the components of gain- or loss-of-function mutations to PrPC remain a matter for debate. Among the proteins described to interact with PrPC is Stress-inducible protein 1 (STI1), a co-chaperonin that is secreted from astrocytes and triggers neuroprotection and neuritogenesis through its interaction with PrPC . In this work, we evaluated the impact of different PrPC pathogenic point mutations on signaling pathways induced by the STI1-PrPC interaction. We found that some of the pathogenic mutations evaluated herein induce partial or total disruption of neuritogenesis and neuroprotection mediated by mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinases 1 and 2 (ERK1/2) and protein kinase A (PKA) signaling triggered by STI1-PrPC engagement. A pathogenic mutant PrPC that lacked both neuroprotection and neuritogenesis activities fail to promote negative dominance upon wild-type PrPC . Also, a STI1-α7-nicotinic acetylcholine receptor-dependent cellular signaling was present in a PrPC mutant that maintained both neuroprotection and neuritogenesis activities similar to what has been previously observed by wild-type PrPC . These results point to a loss-of-function mechanism underlying the pathogenicity of PrPC mutations.

Loss of prion protein is associated with the development of insulin resistance and obesity.

Prion protein (PrPC) was initially described due to its involvement in transmissible spongiform encephalopathies. It was subsequently demonstrated to be a cell surface molecule involved in many physiological processes, such as vesicle trafficking. Here, we investigated the roles of PrPC in the response to insulin and obesity development. Two independent PrPC knockout (KO) and one PrPC overexpressing (TG20) mouse models were fed high-fat diets, and the development of insulin resistance and obesity was monitored. PrPC KO mice fed high-fat diets presented all of the symptoms associated with the development of insulin resistance: hyperglycemia, hyperinsulinemia, and obesity. Conversely, TG20 animals fed high-fat diets showed reduced weight and insulin resistance. Accordingly, the expression of peroxisome proliferator-activated receptor gamma (PPARγ) was reduced in PrPC KO mice and increased in TG20 animals. PrPC KO cells also presented reduced glucose uptake upon insulin stimulation, due to reduced translocation of the glucose transporter Glut4. Thus, our results suggest that PrPC reflects susceptibility to the development of insulin resistance and metabolic syndrome.

A roadmap for investigating the role of the prion protein in depression associated with neurodegenerative disease.

The physiological properties of the native, endogenous prion protein (PrP(C)) is a matter of concern, due to its pleiotropic functions and links to neurodegenerative disorders and cancer. In line with our hypothesis that the basic function of PrP(C) is to serve as a cell surface scaffold for the assembly of signaling modules, multiple interactions have been identified of PrP(C) with signaling molecules, including neurotransmitter receptors. We recently reported evidence that PrP(C) may modulate monoaminergic neurotransmission, as well as depressive-like behavior in mice. Here, we discuss how those results, together with a number of other studies, including our previous demonstration that both inflammatory and behavioral stress modulate PrP(C) content in neutrophils, suggest a distributed role of PrP(C) in clinical depression and inflammation associated with neurodegenerative diseases. An overarching understanding of the multiple interventions of PrP(C) upon physiological events may both shed light on the pathogenesis of, as well as help the identification of novel therapeutic targets for clinical depression, Prion and Alzheimer's Diseases.

Expression of Tyrosine Hydroxylase is Negatively Regulated Via Prion Protein.

Cellular prion protein (PrP(C)) is a glycoprotein of the plasma membrane that plays pleiotropic functions by interacting with multiple signaling complexes at the cell surface. Recently, a number of studies have reported the involvement of PrP(C) in dopamine metabolism and signaling, including its interactions with tyrosine hydroxylase (TH) and dopamine receptors. However, the outcomes reported by independent studies are still debatable. Therefore in this study, we investigated the effects of PrP(C) on the TH expression during the differentiation of N2a cells with dibutyryl-cAMP, a well-known cAMP analog that activates TH transcription. Upon differentiation, TH was induced with concomitant reduction of PrP(C) at protein level, but not at mRNA level. shRNA-mediated PrP(C) reduction increased the basal level of TH at both mRNA and protein levels without dibutyryl-cAMP treatment. This phenotype was reversed by re-expression of PrP(C). PrP(C) knockdown also potentiated the effect of dibutyryl-cAMP on TH expression. Our findings suggest that PrP(C) has suppressive effects on TH expression. As a consequence, altered PrP(C) functions may affect the regulation of dopamine metabolism and related neurological disorders.

Alzheimer's Aß interacts with cellular prion protein inducing neuronal membrane damage and synaptotoxicity.

A major feature of Alzheimer's disease is the accumulation of ß-amyloid (Aß) peptide in the brain. Recent studies have indicated that Aß oligomers (Aßo) can interact with the cellular prion protein (PrPc). Therefore, this interaction might be driving some of Aß toxic effects in the synaptic region. In the present study, we report that Aßo binds to PrPc in the neuronal membrane playing a role on toxic effects induced by Aß. Phospholipase C-enzymatic cleavage of PrPc from the plasma membrane attenuated the association of Aßo to the neurons. Furthermore, an anti-PrP antibody (6D11) decreased the association of Aßo to hippocampal neurons with a concomitant reduction in Aßo and PrPc co-localization. Interestingly, this antibody blocked the increase in membrane conductance and intracellular calcium induced by Aßo. Thus, the data indicate that PrPc plays a role on the membrane perforations produced by Aßo, the increase in calcium ions and the release of synaptic vesicles that subsequently leads to synaptic failure. Future studies blocking Aßo interaction with PrPc could be important for the discovery of new therapeutic strategies for Alzheimer's disease.

Interações proteína príon e seus ligantes STI1 e laminina: possíveis implicações na doença de Alzheimer / Interactions of the prion protein and its ligands STI1 and laminin: possible implications for Alzheimer’s disease

A Doença de Alzheimer (DA) é uma demência progressiva que tem como principais características a disfunção sináptica e a neurodegeneração em áreas específicas do cérebro, que levam a um quadro grave de perda de memória e outras habilidades cognitivas. Uma das principais características neuropatológicas é a deposiçãode placas amiloides extracelulares, que contêm principalmente o peptídeo beta-amiloide (Aβ), que é formado por um processamento alterado da proteína precursora amiloide (APP). Os oligômeros formados por Aβ (AβO) são considerados os elementos tóxicos mais importantes deste processo, esses se ligam às sinapses e estão estreitamente relacionados com a patogênese da DA. Recentemente, foi descrito que a proteína príon celular (PrPC) é um receptor para AβO, porém, os mecanismos envolvidos nesta interação e de que forma esta pode estar relacionada à DA ainda não foram elucidados. PrPC é uma glicoproteína ancorada à membrana plasmática que interage com diversos ligantes, como a proteína de matriz extracelular laminina e a co-chaperona STI1 (Stress Inducible Protein 1). Estas interações induzem neuroproteção, neuritogênese e modulam a formação de memória. Deste modo, torna-se interessante verificar um possível efeito neuroprotetor dos ligantes de PrPC contra a toxicidade produzida pelos AβO. Utilizando culturas neuronais, observamos que os tratamentos com AβO levam uma diminuição nos níveis da proteína sináptica sinaptofisina (Syp), devido a um aumento da degradação..

Alzheimer's disease (AD) is a progressive dementia mainly characterized by synaptic dysfunction and neurodegeneration in specific areas of the brain, leading to severe memory loss, and others cognitive inabilities. One of the main neuropathological characteristics is the formation of extracellular amyloid plaques, which mainly contain beta-amyloid peptide (Aβ). This peptide is formed by an alteration in the processing of the amyloid precursor protein (APP). The Aβ oligomers (AβO) are considered the major toxic components within this process where they bind to synapses and are closely related to the pathogenesis of the AD. Recently, it was reported that PrPC is a receptor for AβO, however, the mechanisms involved in this interaction and how this may be related to AD have not yet been elucidated. The cellular prion protein (PrPC) is a glycoprotein anchored to the plasma membrane that interacts with several ligands such as the co-chaperone STI1 (Stress inducible protein 1) and the extracellular matrix protein laminin. These interactions induce neuroprotection, neuritogenesis, and modulate memory formation. Therefore, it becomes interesting to verify a possible neuroprotective effect of PrPC ligands against the toxicity induced by AβO. We observed that the treatment of neuronal cultures with AβO lead to a decrease in the levels of synaptic protein synaptophysin (Syp) due to an increase of Syp degradation by the proteasome...

O papel da proteína prion celular na regulação da resposta à insulina / The role of cellular prion protein in the regulation of insulin signaling

A proteína Príon Celular ou PrPc, é uma molécula de superfície celular responsável por desencadear várias cascatas de transdução de sinal emediar muitos processos fisiológicos como proteção contra apoptose,indução da proliferação, adesão celular entre outros. Devido à sua grandegama de efeitos, hoje se acredita que PrPc seja um organizador de plataformas lipídicas, ou lipid rafts, na membrana celular. O receptor deinsulina é uma proteína transmembrana também presente em plataformaslipídicas e, interessantemente, a ausência de componentes destasplataformas, como caveolinas, por exemplo, leva a uma deficiência naresposta a insulina. Desta maneira, acreditávamos que a ausência de PrPcpoderia levar a uma desregulação das plataformas lipídicas e a umasinalização ineficiente do receptor de insulina. De fato, pudemos observar ainfluência de PrPc no controle da glicemia sérica, através de experimentos com camundongos que fizeram ingestão de rações com conteúdo controlado de gordura. Foi possível verificar que camundongos deficientes para PrPc não possuem controle adequado da glicemia, de peso e dos níveis de insulina. Estes resultados foram confirmados em duas cepas diferentes de camundongos deficientes para PrPc. Por outro lado, camundongos que superexpressam PrPc apresentam controle adequado de glicemia por mais tempo que camundongos do tipo-selvagem quando alimentados com dietas de alta porcentagem de gordura, sugerindo que a superexpressão de PrPc promoveria uma resistência ao diabetes tipo II, com maior sensibilidade a insulina. As análises histológicas também mostraram que os animaisdeficientes para PrPc apresentam esteatose hepática mais acentuada doque os animais do tipo selvagem, assim como hipertrofia dos adipócitos...

The cellular prion protein or PrPc is a cell surface molecule responsible fortriggering various signal transduction cascades and mediate manyphysiological processes such as protection against apoptosis, inducingproliferation, cell adhesion, among others. Due to its wide range of effects, itis believed that PrPc is a lipid raft organizer in the cell membrane. The insulinreceptor is a transmembrane protein also present in lipid rafts. Interestingly,the absence of lipid rafts’s components as caveolins, for example, leads to adeficiency in insulin response. Thus, we believed that the absence of PrPccould lead to a dysregulation of lipid rafts and inefficient insulin receptorsignaling. In fact, we observed the influence of PrPc in the control of serumglucose, through experiments with mice that ingested diets with controlled fatcontent. We found that mice deficient for PrPc do not have adequate controlof blood glucose, weight gain and insulin levels. These results wereconfirmed in two different strains of PrPc knockout mice. On the other hand,mice overexpressing PrPc exhibit adequate control of blood glucose longerthan wild-type mice when fed with high fat chow, suggesting thatoverexpression of PrPc promote resistance to Type II diabetes, with greaterinsulin sensitivity. The histological analysis also showed that PrPc deficientmice exhibit more pronounced hepatic steatosis than wild-type animals, aswell as adipocyte hypertrophy, which is characteristic of obesity. However,throughout our studies with fibroblasts derived from mice deficient for PrPc,wild-type and overexpressing PrPc, we could not prove our initial hypothesisof change in insulin receptor activity. However, we observed that micedeficient for PrPc have lesser amounts of PPAR-y, a transcription factorinvolved in adipocyte differentiation that has influence on glucose regulation

O papel da proteína prion celular na regulação da resposta à insulina / The role of cellular prion protein in the regulation of insulin signaling

A proteína Príon Celular ou PrPc, é uma molécula de superfície celular responsável por desencadear várias cascatas de transdução de sinal e mediar muitos processos fisiológicos como proteção contra apoptose, indução da proliferação, adesão celular entre outros. Devido à sua grande gama de efeitos, hoje se acredita que PrPc seja um organizador de plataformas lipídicas, ou lipid rafts, na membrana celular. O receptor de insulina é uma proteína transmembrana também presente em plataformas lipídicas e, interessantemente, a ausência de componentes destas plataformas, como caveolinas, por exemplo, leva a uma deficiência na resposta a insulina. Desta maneira, acreditávamos que a ausência de PrPc poderia levar a uma desregulação das plataformas lipídicas e a uma sinalização ineficiente do receptor de insulina. De fato, pudemos observar a influência de PrPc no controle da glicemia sérica, através de experimentos com camundongos que fizeram ingestão de rações com conteúdo controlado de gordura. Foi possível verificar que camundongos deficientes para PrPc não possuem controle adequado da glicemia, de peso e dos níveis de insulina. Estes resultados foram confirmados em duas cepas diferentes de camundongos deficientes para PrPc. Por outro lado, camundongos que superexpressam PrPc apresentam controle adequado de glicemia por mais tempo que camundongos do tipo-selvagem quando alimentados com dietas de alta porcentagem de gordura, sugerindo que a superexpressão de PrPc promoveria uma resistência ao diabetes tipo II, com maior sensibilidade a insulina. As análises histológicas também mostraram que os animais deficientes para PrPc apresentam esteatose hepática mais acentuada do que os animais do tipo selvagem, assim como hipertrofia dos adipócitos, o que é característico de obesidade. Porém ao longo dos nossos estudos com fibroblastos derivados de camundongos deficientes para PrPc, do tipo selvagem e que superexpressam PrPc, pudemos verificar...

The cellular prion protein or PrPc is a cell surface molecule responsible for triggering various signal transduction cascades and mediate many physiological processes such as protection against apoptosis, inducing proliferation, cell adhesion, among others. Due to its wide range of effects, it is believed that PrPc is a lipid raft organizer in the cell membrane. The insulin receptor is a transmembrane protein also present in lipid rafts. Interestingly, the absence of lipid rafts’s components as caveolins, for example, leads to a deficiency in insulin response. Thus, we believed that the absence of PrPc could lead to a dysregulation of lipid rafts and inefficient insulin receptor signaling. In fact, we observed the influence of PrPc in the control of serum glucose, through experiments with mice that ingested diets with controlled fat content. We found that mice deficient for PrPc do not have adequate control of blood glucose, weight gain and insulin levels. These results were confirmed in two different strains of PrPc knockout mice. On the other hand, mice overexpressing PrPc exhibit adequate control of blood glucose longer than wild-type mice when fed with high fat chow, suggesting that overexpression of PrPc promote resistance to Type II diabetes, with greater insulin sensitivity. The histological analysis also showed that PrPc deficient mice exhibit more pronounced hepatic steatosis than wild-type animals, as well as adipocyte hypertrophy, which is characteristic of obesity. However, throughout our studies with fibroblasts derived from mice deficient for PrPc, wild-type and overexpressing PrPc, we could not prove our initial hypothesis of change in insulin receptor activity. However, we observed that mice deficient for PrPc have lesser amounts of PPAR-y, a transcription factor involved in adipocyte differentiation that has influence on glucose regulation. Additionally, by flow cytometry, we found that the translocation of the glucose transporter Glut4...

The unconventional secretion of stress-inducible protein 1 by a heterogeneous population of extracellular vesicles.

The co-chaperone stress-inducible protein 1 (STI1) is released by astrocytes, and has important neurotrophic properties upon binding to prion protein (PrP(C)). However, STI1 lacks a signal peptide and pharmacological approaches pointed that it does not follow a classical secretion mechanism. Ultracentrifugation, size exclusion chromatography, electron microscopy, vesicle labeling, and particle tracking analysis were used to identify three major types of extracellular vesicles (EVs) released from astrocytes with sizes ranging from 20-50, 100-200, and 300-400 nm. These EVs carry STI1 and present many exosomal markers, even though only a subpopulation had the typical exosomal morphology. The only protein, from those evaluated here, present exclusively in vesicles that have exosomal morphology was PrP(C). STI1 partially co-localized with Rab5 and Rab7 in endosomal compartments, and a dominant-negative for vacuolar protein sorting 4A (VPS4A), required for formation of multivesicular bodies (MVBs), impaired EV and STI1 release. Flow cytometry and PK digestion demonstrated that STI1 localized to the outer leaflet of EVs, and its association with EVs greatly increased STI1 activity upon PrP(C)-dependent neuronal signaling. These results indicate that astrocytes secrete a diverse population of EVs derived from MVBs that contain STI1 and suggest that the interaction between EVs and neuronal surface components enhances STI1-PrP(C) signaling.

Laminin-γ1 chain and stress inducible protein 1 synergistically mediate PrPC-dependent axonal growth via Ca2+ mobilization in dorsal root ganglia neurons.

Prion protein (PrP(C)) is a cell surface glycoprotein that is abundantly expressed in nervous system. The elucidation of the PrP(C) interactome network and its significance on neural physiology is crucial to understanding neurodegenerative events associated with prion and Alzheimer's diseases. PrP(C) co-opts stress inducible protein 1/alpha7 nicotinic acetylcholine receptor (STI1/α7nAChR) or laminin/Type I metabotropic glutamate receptors (mGluR1/5) to modulate hippocampal neuronal survival and differentiation. However, potential cross-talk between these protein complexes and their role in peripheral neurons has never been addressed. To explore this issue, we investigated PrP(C)-mediated axonogenesis in peripheral neurons in response to STI1 and laminin-γ1 chain-derived peptide (Ln-γ1). STI1 and Ln-γ1 promoted robust axonogenesis in wild-type neurons, whereas no effect was observed in neurons from PrP(C) -null mice. PrP(C) binding to Ln-γ1 or STI1 led to an increase in intracellular Ca(2+) levels via distinct mechanisms: STI1 promoted extracellular Ca(2+) influx, and Ln-γ1 released calcium from intracellular stores. Both effects depend on phospholipase C activation, which is modulated by mGluR1/5 for Ln-γ1, but depends on, C-type transient receptor potential (TRPC) channels rather than α7nAChR for STI1. Treatment of neurons with suboptimal concentrations of both ligands led to synergistic actions on PrP(C)-mediated calcium response and axonogenesis. This effect was likely mediated by simultaneous binding of the two ligands to PrP(C). These results suggest a role for PrP(C) as an organizer of diverse multiprotein complexes, triggering specific signaling pathways and promoting axonogenesis in the peripheral nervous system.

High levels of cellular prion protein improve astrocyte development.

Prion protein (PrP(C)) has neuroprotective functions and herein we demonstrate that astrocytes from PrP(C)-over-expressing mice are more resistant to induced cell death than wild-type astrocytes. The Stress-Inducible-Protein 1 (STI1), a PrP(C) ligand, prevents cell death in both wild-type and PrP(C)-over-expressing astrocytes through the activation of protein-kinase-A. Cultured embryonic astrocytes and brain extracts from PrP(C)-over-expressing mice show higher glial fibrillary acidic protein expression and reduced vimentin and nestin levels when compared to wild-type astrocytes, suggesting faster astrocyte maturation in the former mice. Our data indicate that PrP(C) levels modulate astrocyte development, and that PrP(C)-STI1 interaction contributes to protect against astrocyte death.

Disease-associated mutations in the prion protein impair laminin-induced process outgrowth and survival.

Prions, the agents of transmissible spongiform encephalopathies, require the expression of prion protein (PrP(C)) to propagate disease. PrP(C) is converted into an abnormal insoluble form, PrP(Sc), that gains neurotoxic activity. Conversely, clinical manifestations of prion disease may occur either before or in the absence of PrP(Sc) deposits, but the loss of normal PrP(C) function contribution for the etiology of these diseases is still debatable. Prion disease-associated mutations in PrP(C) represent one of the best models to understand the impact of PrP(C) loss-of-function. PrP(C) associates with various molecules and, in particular, the interaction of PrP(C) with laminin (Ln) modulates neuronal plasticity and memory formation. To assess the functional alterations associated with PrP(C) mutations, wild-type and mutated PrP(C) proteins were expressed in a neural cell line derived from a PrP(C)-null mouse. Treatment with the laminin γ1 chain peptide (Ln γ1), which mimics the Ln binding site for PrP(C), increased intracellular calcium in cells expressing wild-type PrP(C), whereas a significantly lower response was observed in cells expressing mutated PrP(C) molecules. The Ln γ1 did not promote process outgrowth or protect against staurosporine-induced cell death in cells expressing mutated PrP(C) molecules in contrast to cells expressing wild-type PrP(C). The co-expression of wild-type PrP(C) with mutated PrP(C) molecules was able to rescue the Ln protective effects, indicating the lack of negative dominance of PrP(C) mutated molecules. These results indicate that PrP(C) mutations impair process outgrowth and survival mediated by Ln γ1 peptide in neural cells, which may contribute to the pathogenesis of genetic prion diseases.