RESUMEN
The α-synuclein protein can adopt several different conformations that cause neurodegeneration. Different α-synuclein conformers cause at least three distinct α-synucleinopathies: multiple system atrophy (MSA), dementia with Lewy bodies (DLB), and Parkinson's disease (PD). In earlier studies, we transmitted MSA to transgenic (Tg) mice and cultured HEK cells both expressing mutant α-synuclein (A53T) but not to cells expressing α-synuclein (E46K). Now, we report that DLB is caused by a strain of α-synuclein prions that is distinct from MSA. Using cultured HEK cells expressing mutant α-synuclein (E46K), we found that DLB prions could be transmitted to these HEK cells. Our results argue that a third strain of α-synuclein prions likely causes PD, but further studies are needed to identify cells and/or Tg mice that express a mutant α-synuclein protein that is permissive for PD prion replication. Our findings suggest that other α-synuclein mutants should give further insights into α-synuclein prion replication, strain formation, and disease pathogenesis, all of which are likely required to discover effective drugs for the treatment of PD as well as the other α-synucleinopathies.
Asunto(s)
Demencia/metabolismo , Enfermedad por Cuerpos de Lewy/metabolismo , Atrofia de Múltiples Sistemas/metabolismo , Priones/metabolismo , alfa-Sinucleína/metabolismo , Anciano , Línea Celular , Femenino , Humanos , Masculino , Persona de Mediana Edad , Enfermedad de Parkinson/metabolismo , Sinucleinopatías/metabolismoRESUMEN
In multiple system atrophy (MSA), the α-synuclein protein misfolds into a self-templating prion conformation that spreads throughout the brain, leading to progressive neurodegeneration. While the E46K mutation in α-synuclein causes familial Parkinson's disease (PD), we previously discovered that this mutation blocks in vitro propagation of MSA prions. Recent studies by others indicate that α-synuclein adopts a misfolded conformation in MSA in which a Greek key motif is stabilized by an intramolecular salt bridge between residues E46 and K80. Hypothesizing that the E46K mutation impedes salt bridge formation and, therefore, exerts a selective pressure that can modulate α-synuclein strain propagation, we asked whether three distinct α-synuclein prion strains could propagate in TgM47+/- mice, which express human α-synuclein with the E46K mutation. Following intracranial injection of these strains, TgM47+/- mice were resistant to MSA prion transmission, whereas recombinant E46K preformed fibrils (PFFs) transmitted neurological disease to mice and induced the formation of phosphorylated α-synuclein neuropathology. In contrast, heterotypic seeding following wild-type (WT) PFF-inoculation resulted in preclinical α-synuclein prion propagation. Moreover, when we inoculated TgM20+/- mice, which express WT human α-synuclein, with E46K PFFs, we observed delayed transmission kinetics with an incomplete attack rate. These findings suggest that the E46K mutation constrains the number of α-synuclein prion conformations that can propagate in TgM47+/- mice, expanding our understanding of the selective pressures that impact α-synuclein prion replication.
Asunto(s)
Atrofia de Múltiples Sistemas , Priones , Humanos , Ratones , Animales , alfa-Sinucleína/genética , Priones/genética , Ratones Transgénicos , MutaciónRESUMEN
Various forms of Parkinson's disease, including its common sporadic form, are characterized by prominent α-synuclein (αSyn) aggregation in affected brain regions. However, the role of αSyn in the pathogenesis and evolution of the disease remains unclear, despite vast research efforts of more than a quarter century. A better understanding of the role of αSyn, either primary or secondary, is critical for developing disease-modifying therapies. Previous attempts to hone this research have been challenged by experimental limitations, but recent technological advances may facilitate progress. The Scientific Issues Committee of the International Parkinson and Movement Disorder Society (MDS) charged a panel of experts in the field to discuss current scientific priorities and identify research strategies with potential for a breakthrough. © 2024 The Author(s). Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.
RESUMEN
In several neurodegenerative disorders, proteins that typically exhibit an α-helical structure misfold into an amyloid conformation rich in ß-sheet content. Through a self-templating mechanism, these amyloids are able to induce additional protein misfolding, facilitating their propagation throughout the central nervous system. This disease mechanism was originally identified for the prion protein (PrP), which misfolds into PrPSc in a number of disorders, including variant Creutzfeldt-Jakob disease (vCJD) and bovine spongiform encephalopathy (BSE). More recently, the prion mechanism of disease was expanded to include other proteins that rely on this self-templating mechanism to cause progressive degeneration, including α-synuclein misfolding in Parkinson's disease (PD). Several studies now suggest that PD patients can be subcategorized based on where in the body misfolded α-synuclein originates, either the brain or the gut, similar to patients developing sporadic CJD or vCJD. In this review, we discuss the human and animal model data indicating that α-synuclein and PrPSc misfolding originates in the gut in body-first PD and vCJD, and summarize the data identifying the role of the autonomic nervous system in the gut-brain axis of both diseases.
Asunto(s)
Encéfalo/metabolismo , Síndrome de Creutzfeldt-Jakob/metabolismo , Enfermedad de Parkinson/metabolismo , Animales , Encéfalo/patología , Síndrome de Creutzfeldt-Jakob/patología , Modelos Animales de Enfermedad , Humanos , Neuronas/metabolismo , Neuronas/patología , Enfermedad de Parkinson/patología , Proteínas Priónicas/metabolismoRESUMEN
Multiple system atrophy (MSA), a progressive neurodegenerative disease characterized by autonomic dysfunction and motor impairment, is caused by the self-templated misfolding of the protein α-synuclein. With no treatment currently available, we sought to characterize the spread of α-synuclein in a transgenic mouse model of MSA prion propagation to support drug discovery programs for synucleinopathies. Brain homogenates from MSA patient samples or mouse-passaged MSA were inoculated either by standard freehand injection or stereotactically into TgM83+/- mice, which express human α-synuclein with the A53T mutation. Following disease onset, brains from the mice were tested for biologically active α-synuclein prions using a cell-based assay and examined for α-synuclein neuropathology. Inoculation studies using homogenates prepared from brain regions lacking detectable α-synuclein neuropathology transmitted neurological disease to mice. Terminal animals contained similar concentrations of α-synuclein prions; however, a time-course study where mice were terminated every five days through disease progression revealed that the kinetics of α-synuclein prion replication in the mice were variable. Stereotactic inoculation into the thalamus reduced variability in disease onset in the mice, although incubation times were consistent with standard inoculations. Using human samples with and without neuropathological lesions, we observed that α-synuclein prion formation precedes neuropathology in the brain, suggesting that disease in patients is not limited to brain regions containing neuropathological lesions.
Asunto(s)
Encéfalo/metabolismo , Atrofia de Múltiples Sistemas/metabolismo , Mutación Puntual , alfa-Sinucleína/metabolismo , Animales , Encéfalo/patología , Femenino , Humanos , Cinética , Masculino , Ratones , Ratones Transgénicos , Atrofia de Múltiples Sistemas/genética , Atrofia de Múltiples Sistemas/patología , Priones/genética , Priones/metabolismo , alfa-Sinucleína/genéticaRESUMEN
Synucleinopathies are a group of clinically and neuropathologically distinct protein misfolding diseases caused by unique α-synuclein conformations, or strains. While multiple atomic resolution cryo-electron microscopy structures of α-synuclein fibrils are now deposited in Protein Data Bank, significant gaps in the biological consequences arising from each conformation have yet to be unraveled. Mutations in the α-synuclein gene (SNCA), cofactors, and the solvation environment contribute to the formation and maintenance of each disease-causing strain. This review highlights the impact of each of these factors on α-synuclein misfolding and discusses the implications of the resulting structural variability on therapeutic development.
Asunto(s)
Deficiencias en la Proteostasis , Sinucleinopatías , Biología , Microscopía por Crioelectrón , Humanos , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismoRESUMEN
In multiple system atrophy (MSA), the protein α-synuclein misfolds into a prion conformation that self-templates and causes progressive neurodegeneration. While many point mutations in the α-synuclein gene, SNCA, have been identified as the cause of heritable Parkinson's disease (PD), none have been identified as causing MSA. To examine whether MSA prions can transmit disease to mice expressing wild-type (WT) human α-synuclein, we inoculated transgenic (Tg) mice denoted TgM20+/- with brain homogenates prepared from six different deceased MSA patients. All six samples transmitted CNS disease to the mice, with an average incubation period of ~ 280 days. Interestingly, TgM20+/- female mice developed disease > 60 days earlier than their male counterparts. Brains from terminal mice contained phosphorylated α-synuclein throughout the hindbrain, consistent with the distribution of α-synuclein inclusions in MSA patients. In addition, using our α-syn-YFP cell lines, we detected α-synuclein prions in brain homogenates prepared from terminal mice that retained MSA strain properties. To our knowledge, the studies described here are the first to show that MSA prions transmit neurological disease to mice expressing WT SNCA and that the rate of transmission is sex dependent. By comparison, TgM20+/- mice inoculated with WT preformed fibrils (PFFs) developed severe neurological disease in ~ 210 days and exhibited robust α-synuclein neuropathology in both limbic regions and the hindbrain. Brain homogenates from these animals exhibited biological activities that are distinct from those found in MSA-inoculated mice when tested in the α-syn-YFP cell lines. Differences between brains from MSA-inoculated and WT PFF-inoculated mice potentially argue that α-synuclein prions from MSA patients are distinct from the PFF inocula and that PFFs do not replicate MSA strain biology.
Asunto(s)
Atrofia de Múltiples Sistemas , Priones , Animales , Femenino , Humanos , Cuerpos de Inclusión/patología , Masculino , Ratones , Ratones Transgénicos , Atrofia de Múltiples Sistemas/patología , Priones/genética , Priones/metabolismo , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismoAsunto(s)
Atrofia de Múltiples Sistemas , Enfermedad de Parkinson , Priones , Humanos , alfa-SinucleínaRESUMEN
Synucleinopathies are a group of neurodegenerative disorders caused by the misfolding and self-templating of the protein α-synuclein, or the formation of α-synuclein prions. Each disorder differs by age of onset, presenting clinical symptoms, α-synuclein inclusion morphology, and neuropathological distribution. Explaining this disease-specific variability, the strain hypothesis postulates that each prion disease is encoded by a distinct conformation of the misfolded protein, and therefore, each synucleinopathy is caused by a unique α-synuclein structure. This review discusses the current data supporting the role of α-synuclein strains in disease heterogeneity. Several in vitro and in vivo models exist for evaluating strain behavior, however, as the focus of this article is to compare strains across synucleinopathy patients, our discussion predominantly focuses on the two models most commonly used for this purpose: the α-syn140*A53T-YFP cell line and the TgM83+/- mouse model. Here we define each strain based on biochemical stability, ability to propagate in α-syn140-YFP cell lines, and incubation period, inclusion morphology and distribution, and neurological signs in TgM83+/- mice.
Asunto(s)
Enfermedades Neurodegenerativas/genética , Sinucleinopatías/genética , alfa-Sinucleína/genética , Animales , Línea Celular , Humanos , Ratones , Ratones Noqueados , Ratones TransgénicosRESUMEN
In the neurodegenerative disease multiple system atrophy (MSA), α-synuclein misfolds into a self-templating conformation to become a prion. To compare the biological activity of α-synuclein prions in MSA and Parkinson's disease (PD), we developed nine α-synuclein-YFP cell lines expressing point mutations responsible for inherited PD. MSA prions robustly infected wild-type, A30P, and A53T α-synuclein-YFP cells, but they were unable to replicate in cells expressing the E46K mutation. Coexpression of the A53T and E46K mutations was unable to rescue MSA prion infection in vitro, establishing that MSA α-synuclein prions are conformationally distinct from the misfolded α-synuclein in PD patients. This observation may have profound implications for developing treatments for neurodegenerative diseases.
Asunto(s)
Atrofia de Múltiples Sistemas/genética , Enfermedad de Parkinson/genética , Mutación Puntual , Priones/genética , Animales , Línea Celular , Células HEK293 , Humanos , Ratones Transgénicos , Priones/metabolismo , Priones/patogenicidad , Pliegue de Proteína , alfa-Sinucleína/química , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismoRESUMEN
Previously, we reported that intracranial inoculation of brain homogenate from multiple system atrophy (MSA) patient samples produces neurological disease in the transgenic (Tg) mouse model TgM83+/-, which uses the prion protein promoter to express human α-synuclein harboring the A53T mutation found in familial Parkinson's disease (PD). In our studies, we inoculated MSA and control patient samples into Tg mice constructed using a P1 artificial chromosome to express wild-type (WT), A30P, and A53T human α-synuclein on a mouse α-synuclein knockout background [Tg(SNCA+/+)Nbm, Tg(SNCA*A30P+/+)Nbm, and Tg(SNCA*A53T+/+)Nbm]. In contrast to studies using TgM83+/- mice, motor deficits were not observed by 330-400 days in any of the Tg(SNCA)Nbm mice after inoculation with MSA brain homogenates. However, using a cell-based bioassay to measure α-synuclein prions, we found brain homogenates from Tg(SNCA*A53T+/+)Nbm mice inoculated with MSA patient samples contained α-synuclein prions, whereas control mice did not. Moreover, these α-synuclein aggregates retained the biological and biochemical characteristics of the α-synuclein prions in MSA patient samples. Intriguingly, Tg(SNCA*A53T+/+)Nbm mice developed α-synuclein pathology in neurons and astrocytes throughout the limbic system. This finding is in contrast to MSA-inoculated TgM83+/- mice, which develop exclusively neuronal α-synuclein aggregates in the hindbrain that cause motor deficits with advanced disease. In a crossover experiment, we inoculated TgM83+/- mice with brain homogenate from two MSA patient samples or one control sample first inoculated, or passaged, in Tg(SNCA*A53T+/+)Nbm animals. Additionally, we performed the reverse experiment by inoculating Tg(SNCA*A53T+/+)Nbm mice with brain homogenate from the same two MSA samples and one control sample first passaged in TgM83+/- animals. The TgM83+/- mice inoculated with mouse-passaged MSA developed motor dysfunction and α-synuclein prions, whereas the mouse-passaged control sample had no effect. Similarly, the mouse-passaged MSA samples induced α-synuclein prion formation in Tg(SNCA*A53T+/+)Nbm mice, but the mouse-passaged control sample did not. The confirmed transmission of α-synuclein prions to a second synucleinopathy model and the ability to propagate prions between two distinct mouse lines while retaining strain-specific properties provides compelling evidence that MSA is a prion disease.
Asunto(s)
Atrofia de Múltiples Sistemas/patología , Enfermedades por Prión/patología , Enfermedades por Prión/transmisión , Priones/metabolismo , alfa-Sinucleína/metabolismo , Animales , Humanos , Ratones , Ratones TransgénicosRESUMEN
Tau prions are thought to aggregate in the central nervous system, resulting in neurodegeneration. Among the tauopathies, Alzheimer's disease (AD) is the most common, whereas argyrophilic grain disease (AGD), corticobasal degeneration (CBD), chronic traumatic encephalopathy (CTE), Pick's disease (PiD), and progressive supranuclear palsy (PSP) are less prevalent. Brain extracts from deceased individuals with PiD, a neurodegenerative disorder characterized by three-repeat (3R) tau prions, were used to infect HEK293T cells expressing 3R tau fused to yellow fluorescent protein (YFP). Extracts from AGD, CBD, and PSP patient samples, which contain four-repeat (4R) tau prions, were transmitted to HEK293 cells expressing 4R tau fused to YFP. These studies demonstrated that prion propagation in HEK cells requires isoform pairing between the infecting prion and the recipient substrate. Interestingly, tau aggregates in AD and CTE, containing both 3R and 4R isoforms, were unable to robustly infect either 3R- or 4R-expressing cells. However, AD and CTE prions were able to replicate in HEK293T cells expressing both 3R and 4R tau. Unexpectedly, increasing the level of 4R isoform expression alone supported the propagation of both AD and CTE prions. These results allowed us to determine the levels of tau prions in AD and CTE brain extracts.
Asunto(s)
Enfermedad de Alzheimer/metabolismo , Encefalopatía Traumática Crónica/metabolismo , Proteínas tau/metabolismo , Enfermedad de Alzheimer/genética , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Línea Celular , Encefalopatía Traumática Crónica/genética , Células HEK293 , Humanos , Proteínas Luminiscentes/química , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Mutación , Enfermedad de Pick/genética , Enfermedad de Pick/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Parálisis Supranuclear Progresiva/genética , Parálisis Supranuclear Progresiva/metabolismo , Regulación hacia Arriba , Proteínas tau/química , Proteínas tau/genéticaRESUMEN
In multiple system atrophy (MSA), progressive neurodegeneration results from the protein α-synuclein misfolding into a self-templating prion conformation that spreads throughout the brain. MSA prions are transmissible to transgenic (Tg) mice expressing mutated human α-synuclein (TgM83+/-), inducing neurological disease following intracranial inoculation with brain homogenate from deceased patient samples. Noting the similarities between α-synuclein prions and PrP scrapie (PrPSc) prions responsible for Creutzfeldt-Jakob disease (CJD), we investigated MSA transmission under conditions known to result in PrPSc transmission. When peripherally exposed to MSA via the peritoneal cavity, hind leg muscle, and tongue, TgM83+/- mice developed neurological signs accompanied by α-synuclein prions in the brain. Iatrogenic CJD, resulting from PrPSc prion adherence to surgical steel instruments, has been investigated by incubating steel sutures in contaminated brain homogenate before implantation into mouse brain. Mice studied using this model for MSA developed disease, whereas wire incubated in control homogenate had no effect on the animals. Notably, formalin fixation did not inactivate α-synuclein prions. Formalin-fixed MSA patient samples also transmitted disease to TgM83+/- mice, even after incubating in fixative for 244 months. Finally, at least 10% sarkosyl was found to be the concentration necessary to partially inactivate MSA prions. These results demonstrate the robustness of α-synuclein prions to denaturation. Moreover, they establish the parallel characteristics between PrPSc and α-synuclein prions, arguing that clinicians should exercise caution when working with materials that might contain α-synuclein prions to prevent disease.
Asunto(s)
Atrofia de Múltiples Sistemas/metabolismo , Priones/metabolismo , Animales , Transporte Biológico , Encéfalo/metabolismo , Encéfalo/patología , Detergentes/farmacología , Modelos Animales de Enfermedad , Fijadores , Formaldehído , Células HEK293 , Humanos , Ratones Transgénicos , Atrofia de Múltiples Sistemas/patología , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Mutación , Priones/administración & dosificación , Agregado de Proteínas , Estabilidad Proteica/efectos de los fármacos , Sarcosina/análogos & derivados , Sarcosina/farmacología , Acero Inoxidable , alfa-Sinucleína/administración & dosificación , alfa-Sinucleína/efectos adversos , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismoRESUMEN
Increasingly, evidence argues that many neurodegenerative diseases, including progressive supranuclear palsy (PSP), are caused by prions, which are alternatively folded proteins undergoing self-propagation. In earlier studies, PSP prions were detected by infecting human embryonic kidney (HEK) cells expressing a tau fragment [TauRD(LM)] fused to yellow fluorescent protein (YFP). Here, we report on an improved bioassay using selective precipitation of tau prions from human PSP brain homogenates before infection of the HEK cells. Tau prions were measured by counting the number of cells with TauRD(LM)-YFP aggregates using confocal fluorescence microscopy. In parallel studies, we fused α-synuclein to YFP to bioassay α-synuclein prions in the brains of patients who died of multiple system atrophy (MSA). Previously, MSA prion detection required â¼120 d for transmission into transgenic mice, whereas our cultured cell assay needed only 4 d. Variation in MSA prion levels in four different brain regions from three patients provided evidence for three different MSA prion strains. Attempts to demonstrate α-synuclein prions in brain homogenates from Parkinson's disease patients were unsuccessful, identifying an important biological difference between the two synucleinopathies. Partial purification of tau and α-synuclein prions facilitated measuring the levels of these protein pathogens in human brains. Our studies should facilitate investigations of the pathogenesis of both tau and α-synuclein prion disorders as well as help decipher the basic biology of those prions that attack the CNS.
Asunto(s)
Enfermedades Neurodegenerativas/metabolismo , Priones/metabolismo , alfa-Sinucleína/metabolismo , Animales , Células HEK293 , Humanos , Ratones , Enfermedades Neurodegenerativas/patologíaRESUMEN
Prions are proteins that adopt alternative conformations that become self-propagating; the PrP(Sc) prion causes the rare human disorder Creutzfeldt-Jakob disease (CJD). We report here that multiple system atrophy (MSA) is caused by a different human prion composed of the α-synuclein protein. MSA is a slowly evolving disorder characterized by progressive loss of autonomic nervous system function and often signs of parkinsonism; the neuropathological hallmark of MSA is glial cytoplasmic inclusions consisting of filaments of α-synuclein. To determine whether human α-synuclein forms prions, we examined 14 human brain homogenates for transmission to cultured human embryonic kidney (HEK) cells expressing full-length, mutant human α-synuclein fused to yellow fluorescent protein (α-syn140*A53T-YFP) and TgM83(+/-) mice expressing α-synuclein (A53T). The TgM83(+/-) mice that were hemizygous for the mutant transgene did not develop spontaneous illness; in contrast, the TgM83(+/+) mice that were homozygous developed neurological dysfunction. Brain extracts from 14 MSA cases all transmitted neurodegeneration to TgM83(+/-) mice after incubation periods of â¼120 d, which was accompanied by deposition of α-synuclein within neuronal cell bodies and axons. All of the MSA extracts also induced aggregation of α-syn*A53T-YFP in cultured cells, whereas none of six Parkinson's disease (PD) extracts or a control sample did so. Our findings argue that MSA is caused by a unique strain of α-synuclein prions, which is different from the putative prions causing PD and from those causing spontaneous neurodegeneration in TgM83(+/+) mice. Remarkably, α-synuclein is the first new human prion to be identified, to our knowledge, since the discovery a half century ago that CJD was transmissible.
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Atrofia de Múltiples Sistemas/metabolismo , Trastornos Parkinsonianos/metabolismo , Priones/metabolismo , alfa-Sinucleína/metabolismo , Anciano , Animales , Encéfalo/patología , Exones , Femenino , Células HEK293 , Humanos , Inmunohistoquímica , Masculino , Ratones , Ratones Transgénicos , Microscopía Fluorescente , Persona de Mediana Edad , Atrofia de Múltiples Sistemas/genética , Enfermedades Neurodegenerativas/metabolismo , Fosforilación , Polimorfismo de Nucleótido Simple , Ubiquinona/análogos & derivados , Ubiquinona/metabolismo , alfa-Sinucleína/genéticaAsunto(s)
Enfermedades Neurodegenerativas/patología , Enfermedades Neurodegenerativas/terapia , Humanos , Enfermedades Neurodegenerativas/diagnóstico , Medicina de Precisión , Deficiencias en la Proteostasis/diagnóstico , Deficiencias en la Proteostasis/patología , Deficiencias en la Proteostasis/terapiaRESUMEN
Prion diseases are a group of neurodegenerative disorders caused by misfolding of proteins into pathogenic conformations that self-template to spread disease. Although this mechanism is largely associated with the prion protein (PrP) in classical prion diseases, a growing literature indicates that other proteins, including α-synuclein, rely on a similar disease mechanism. Notably, α-synuclein misfolds into distinct conformations, or strains, that cause discrete clinical disorders including multiple system atrophy (MSA) and Parkinson's disease (PD). Because the recognized similarities between PrP and α-synuclein are increasing, this review article draws from research on PrP to identify the host and strain factors that impact disease pathogenesis, predominantly in rodent models, and focuses on key considerations for future research on α-synuclein prions.
Asunto(s)
Enfermedades por Prión , alfa-Sinucleína , alfa-Sinucleína/metabolismo , Animales , Humanos , Enfermedades por Prión/metabolismo , Priones/metabolismoRESUMEN
Pre-formed fibrils (PFFs) made from recombinant α-synuclein are broadly used throughout the field in cellular and animal models of Parkinson's disease. However, their ability to successfully recapitulate disease biology is a controversial topic. In this article, two researchers debate this issue with Amanda Woerman taking the view that PFFs are a model of synucleinopathy but not Parkinson's disease, while Kelvin Luk defends their use as an important tool in the field.
Parkinson's disease patients develop accumulations of misfolded proteins in the brain called Lewy bodies. An important protein found in Lewy bodies is α-synuclein, which aggregates into large fibrils that are thought to have toxic effects on neurons. To study the effect of α-synuclein fibrils, many scientists use pre-formed fibrils (PFFs), which are α-synuclein fibrils made in the lab under a variety of conditions. In this article, two researchers will debate the use of PFFs as a model of Parkinson's disease. Amanda Woerman discusses the data suggesting why PFFs are an inadequate tool for understanding Parkinson's disease. Kelvin Luk discusses the data supporting the use of PFFs as a Parkinson's disease model.
RESUMEN
Multiple system atrophy (MSA) and Parkinson's disease (PD) are caused by misfolded α -synuclein spreading throughout the central nervous system. While familial PD is linked to several point mutations in α -synuclein, there are no known mutations associated with MSA. Our previous work investigating differences in α -synuclein misfolding between the two disorders showed that the familial PD mutation E46K inhibits replication of MSA prions both in vitro and in vivo, providing key evidence to support the hypothesis that α -synuclein adopts unique strains in patients. Here, to further interrogate α -synuclein misfolding, we engineered a panel of cell lines harboring both PD-linked and novel mutations designed to identify key residues that facilitate α -synuclein misfolding in MSA. These data were paired with in silico analyses using Maestro software to predict the effect of each mutation on the ability of α -synuclein to misfold into one of the reported MSA cryo-electron microscopy conformations. In many cases, our modeling accurately identified mutations that facilitated or inhibited MSA replication. However, Maestro was occasionally unable to predict the effect of a mutation on MSA propagation in vitro, demonstrating the challenge of using computational tools to investigate intrinsically disordered proteins. Finally, we used our cellular models to determine the mechanism underlying the E46K-driven inhibition of MSA replication, finding that the E46/K80 salt bridge is necessary to support α -synuclein misfolding. Overall, our studies use a structure-based approach to investigate α -synuclein misfolding, resulting in the creation of a powerful panel of cell lines that can be used to interrogate MSA strain biology.
RESUMEN
The recent discovery by cryo-electron microscopy that the neuropatho-logical hallmarks of different tauopathies, including Alzheimer's disease, corticobasal degeneration (CBD), and progressive supranuclear palsy (PSP), are caused by unique misfolded conformations of the protein tau is among the most profound developments in neurodegenerative disease research. To capitalize on these discoveries for therapeutic development, one must achieve in vitro replication of tau fibrils that adopt the rep-resentative tauopathy disease folds - a grand challenge. To understand whether the commonly used, but imperfect, fragment of the tau pro-tein, K18, is capable of inducing specific protein folds, fibril seeds derived from CBD- and PSP-infected biosensor cells expressing K18, were used to achieve cell-free assembly of naïve, recombinant 4R tau into fibrils without the addition of any cofactors. Using Double Electron Electron Resonance (DEER) spectroscopy, we discovered that cell-passaged patho-logical seeds generate heterogeneous fibrils that are distinct between the CBD and PSP lysate-seeded fibrils, and are also unique from heparin-induced tau fibril populations. Moreover, the lysate-seeded fibrils contain a characteristic sub-population that resembles either the CBD or PSP disease fold, corresponding with the respective starting patient sam-ple. These findings indicate that CBD and PSP patient-derived fibrils retain strain properties after passaging through K18 reporter cells.