RESUMEN
Microglia are crucial for maintaining brain health and neuron function. Here, we report that microglia establish connections with neurons using tunneling nanotubes (TNTs) in both physiological and pathological conditions. These TNTs facilitate the rapid exchange of organelles, vesicles, and proteins. In neurodegenerative diseases like Parkinson's and Alzheimer's disease, toxic aggregates of alpha-synuclein (α-syn) and tau accumulate within neurons. Our research demonstrates that microglia use TNTs to extract neurons from these aggregates, restoring neuronal health. Additionally, microglia share their healthy mitochondria with burdened neurons, reducing oxidative stress and normalizing gene expression. Disrupting mitochondrial function with antimycin A before TNT formation eliminates this neuroprotection. Moreover, co-culturing neurons with microglia and promoting TNT formation rescues suppressed neuronal activity caused by α-syn or tau aggregates. Notably, TNT-mediated aggregate transfer is compromised in microglia carrying Lrrk22(Gly2019Ser) or Trem2(T66M) and (R47H) mutations, suggesting a role in the pathology of these gene variants in neurodegenerative diseases.
RESUMEN
Huntington's disease neurodegeneration occurs when the number of consecutive glutamines in the huntingtin exon-1 (HTTExon1) exceeds a pathological threshold of 35. The sequence homogeneity of HTTExon1 reduces the signal dispersion in NMR spectra, hampering its structural characterization. By simultaneously introducing three isotopically labeled glutamines in a site-specific manner in multiple concatenated samples, 18 glutamines of a pathogenic HTTExon1 with 36 glutamines were unambiguously assigned. Chemical shift analyses indicate the α-helical persistence in the homorepeat and the absence of an emerging toxic conformation around the pathological threshold. Using the same type of samples, the recognition mechanism of Hsc70 molecular chaperone has been investigated, indicating that it binds to the N17 region of HTTExon1, inducing the partial unfolding of the poly-Q. The proposed strategy facilitates high-resolution structural and functional studies in low-complexity regions.
Asunto(s)
Péptidos , Péptidos/química , Exones , Conformación Proteica en Hélice alfa , Espectroscopía de Resonancia Magnética , Proteína Huntingtina/químicaRESUMEN
The role of alpha-synuclein in Parkinson's disease has been heavily investigated since its discovery as a component of Lewy bodies. Recent rodent data demonstrate that alpha-synuclein strain structure is critical for differential propagation and toxicity. Based on these findings, we have compared, for the first time, in this pilot study, the capacity of two alpha-synuclein strains and patient-derived Lewy body extracts to model synucleinopathies after intra-putaminal injection in the non-human primate brain. Functional alterations triggered by these injections were evaluated in vivo using glucose positron emission tomography imaging. Post-mortem immunohistochemical and biochemical analyses were used to detect neuropathological alterations in the dopaminergic system and alpha-synuclein pathology propagation. In vivo results revealed a decrease in glucose metabolism more pronounced in alpha-synuclein strain-injected animals. Histology showed a decreased number of dopaminergic tyrosine hydroxylase-positive cells in the substantia nigra to different extents according to the inoculum used. Biochemistry revealed that alpha-synuclein-induced aggregation, phosphorylation, and propagation in different brain regions are strain-specific. Our findings show that distinct alpha-synuclein strains can induce specific patterns of synucleinopathy in the non-human primate, changes in the nigrostriatal pathway, and functional alterations that resemble early-stage Parkinson's disease.
Asunto(s)
Enfermedad de Parkinson , Sinucleinopatías , Animales , alfa-Sinucleína/metabolismo , Enfermedad de Parkinson/metabolismo , Proyectos Piloto , Cuerpos de Lewy/metabolismo , Sinucleinopatías/patología , Sustancia Negra/metabolismo , Dopamina/metabolismo , Primates/metabolismoRESUMEN
Parkinson's disease (PD) is a progressive, neurodegenerative disorder characterised by the abnormal accumulation of α-synuclein (α-syn) aggregates. Central to disease progression is the gradual spread of pathological α-syn. α-syn aggregation is closely linked to progressive neuron loss. As such, clearance of α-syn aggregates may slow the progression of PD and lead to less severe symptoms. Evidence is increasing that non-neuronal cells play a role in PD and other synucleinopathies such as Lewy body dementia and multiple system atrophy. Our previous work has shown that pericytes-vascular mural cells that regulate the blood-brain barrier-contain α-syn aggregates in human PD brains. Here, we demonstrate that pericytes efficiently internalise fibrillar α-syn irrespective of being in a monoculture or mixed neuronal cell culture. Pericytes cleave fibrillar α-syn aggregates (Fibrils, Ribbons, fibrils65, fibrils91 and fibrils110), with cleaved α-syn remaining present for up to 21 days. The number of α-syn aggregates/cell and average aggregate size depends on the type of strain, but differences disappear within 5 five hours of treatment. Our results highlight the role brain vasculature may play in reducing α-syn aggregate burden in PD.
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Enfermedad por Cuerpos de Lewy , Enfermedad de Parkinson , Humanos , alfa-Sinucleína/metabolismo , Pericitos/metabolismo , Enfermedad por Cuerpos de Lewy/metabolismo , Enfermedad de Parkinson/patología , Neuronas/metabolismoRESUMEN
Synucleinopathies are neurodegenerative diseases characterized by the presence of intracellular deposits containing the protein alpha-synuclein (aSYN) within patients' brains. It has been shown that aSYN can form structurally distinct fibrillar assemblies, also termed polymorphs. We previously showed that distinct aSYN polymorphs assembled in vitro, named fibrils, ribbons, and fibrils 91, differentially bind to and seed the aggregation of endogenous aSYN in neuronal cells, which suggests that distinct synucleinopathies may arise from aSYN polymorphs. In order to better understand the differential interactions of aSYN polymorphs with their partner proteins, we mapped aSYN polymorphs surfaces. We used limited proteolysis, hydrogen-deuterium exchange, and differential antibody accessibility to identify amino acids on their surfaces. We showed that the aSYN C-terminal region spanning residues 94 to 140 exhibited similarly high solvent accessibility in these three polymorphs. However, the N-terminal amino acid residues 1 to 38 of fibrils were exposed to the solvent, while only residues 1 to 18 within fibrils 91 were exposed, and no N-terminal residues within ribbons were solvent-exposed. It is likely that these differences in surface accessibility contribute to the differential binding of distinct aSYN polymorphs to partner proteins. We thus posit that the polypeptides exposed on the surface of distinct aSYN fibrillar polymorphs are comparable to fingerprints. Our findings have diagnostic and therapeutic potential, particularly in the prion-like propagation of fibrillar aSYN, as they can facilitate the design of ligands that specifically bind and distinguish between fibrillar polymorphs.
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Sinucleinopatías/metabolismo , alfa-Sinucleína/metabolismo , Humanos , Conformación Proteica , Proteolisis , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/ultraestructura , Solventes/metabolismo , Sinucleinopatías/patología , alfa-Sinucleína/química , alfa-Sinucleína/ultraestructuraRESUMEN
The spread of fibrillar alpha-synuclein from affected to naïve neuronal cells is thought to contribute to the progression of synucleinopathies. The binding of fibrillar alpha-synuclein to the plasma membrane is key in this process. We and others previously showed that coating fibrillar alpha-synuclein by the molecular chaperone Hsc70 affects fibrils properties. Here we assessed the effect of the two molecular chaperones alpha B-crystallin and CHIP on alpha-synuclein fibrils uptake by Neuro-2a cells. We demonstrate that both chaperones diminish fibrils take up by cells. We identify through a cross-linking and mass spectrometry strategy the interaction interfaces between alpha-synuclein fibrils and alpha B-crystallin or CHIP. Our results open the way for designing chaperone-derived polypeptide binders that interfere with the propagation of pathogenic alpha-synuclein assemblies.
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Amiloide/metabolismo , Mapas de Interacción de Proteínas , Ubiquitina-Proteína Ligasas/metabolismo , Cadena B de alfa-Cristalina/metabolismo , alfa-Sinucleína/metabolismo , Amiloide/ultraestructura , Animales , Línea Celular Tumoral , Humanos , Ratones , Modelos Moleculares , Neuronas/metabolismoRESUMEN
α-synuclein fibrillar polymorphs, Tau and Aß 1-42 fibrillar assemblies have been shown to propagate, amplify and trigger the formation of protein deposits reminiscent of those present within the central nervous system of patients developing synucleinopathies, tauopathies and amyloid plaques after injection intracerebrally, intramuscularly, intraperitoneally or within the blood stream of model animals. They are thus hazardous and there is need for decontamination and inactivation procedures for laboratory surfaces and non-disposable material. We assessed the effectiveness of different reagents to clean and disassemble potentially pathogenic assemblies adsorbed on non-disposable materials in laboratories. We show that commercial detergents and SDS are way more suited to detach α-synuclein fibrillar polymorphs, Tau and Aß 1-42 fibrillar assemblies from contaminated surfaces and disassemble the fibrils than methods designed to decrease PrP prion infectivity. Our observations reveal that the choice of the most adapted cleaning procedure for one given protein assembly or fibrillar polymorph should integrate detergent's cleaning efficiency, material compatibility and capacity to dismantle assemblies. We provide an integrated representation where desorption and neutralization efficacy and surface compatibility are combined to facilitate the choice of the most adapted decontamination procedure. This representation, together with good laboratory practices, contributes to reducing potential health hazards associated to manipulating protein assemblies with prion-like properties.