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Proteopathic tau seeding predicts tauopathy in vivo.

ABSTRACT: Transcellular propagation of protein aggregates, or proteopathic seeds, may drive the progression of neurodegenerative diseases in a prion-like manner. In tauopathies such as Alzheimer's disease, this model predicts that tau seeds propagate pathology through the brain via cell-cell transfer in neural networks. The critical role of tau seeding activity is untested, however. It is unknown whether seeding anticipates and correlates with subsequent development of pathology as predicted for a causal agent. One major limitation has been the lack of a robust assay to measure proteopathic seeding activity in biological specimens. We engineered an ultrasensitive, specific, and facile FRET-based flow cytometry biosensor assay based on expression of tau or synuclein fusions to CFP and YFP, and confirmed its sensitivity and specificity to tau (? 300 fM) and synuclein (? 300 pM) fibrils. This assay readily discriminates Alzheimer's disease vs. Huntington's disease and aged control brains. We then carried out a detailed time-course study in P301S tauopathy mice, comparing seeding activity versus histological markers of tau pathology, including MC1, AT8, PG5, and Thioflavin S. We detected robust seeding activity at 1.5 mo, >1 mo before the earliest histopathological stain. Proteopathic tau seeding is thus an early and robust marker of tauopathy, suggesting a proximal role for tau seeds in neurodegeneration.

SUBMITTER: Holmes BB

PROVIDER: S-EPMC4205609 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature

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Proteopathic tau seeding predicts tauopathy in vivo.

Holmes Brandon B BB Furman Jennifer L JL Mahan Thomas E TE Yamasaki Tritia R TR Mirbaha Hilda H Eades William C WC Belaygorod Larisa L Cairns Nigel J NJ Holtzman David M DM Diamond Marc I MI

Proceedings of the National Academy of Sciences of the United States of America 20140926 41

Transcellular propagation of protein aggregates, or proteopathic seeds, may drive the progression of neurodegenerative diseases in a prion-like manner. In tauopathies such as Alzheimer's disease, this model predicts that tau seeds propagate pathology through the brain via cell-cell transfer in neural networks. The critical role of tau seeding activity is untested, however. It is unknown whether seeding anticipates and correlates with subsequent development of pathology as predicted for a causal ...[more]

PMID: 25261551

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Project description:Tauopathies are neurodegenerative disorders characterized by aggregation of microtubule associated tau protein in neurons and glia. They are clinically and pathologically heterogeneous depending on the isoform of tau protein that accumulates (three or four 31-to-32-amino-acid repeats [3R or 4R] in the microtubule binding domain), as well as the cellular and neuroanatomical distribution of tau pathology. Growing evidence suggests that distinct tau conformers may contribute to the characteristic features of various tauopathies. Globular glial tauopathy (GGT) is a rare 4R tauopathy with globular cytoplasmic inclusions within neurons and glial cells. Given the unique cellular distribution and morphology of tau pathology in GGT, we sought to determine if tau species in GGT had distinctive biological properties. To address this question, we performed seeding analyses with postmortem brain tissues using a commercial tau biosensor cell line. We found that brain lysates from GGT cases had significantly higher seeding competency than other tauopathies, including corticobasal degeneration (CBD), progressive supranuclear palsy (PSP), and Alzheimer's disease (AD). The robust seeding activity of GGT brain lysates was independent of phosphorylated tau burden and diminished upon removal of tau from samples, suggesting that seeding properties were indeed mediated by tau in the lysates. In addition, cellular inclusions in the tau biosensor cell line induced by GGT had a distinct, globular morphology that was markedly different from inclusions induced by other tauopathies, further highlighting the unique nature of tau species in GGT. Characterization of different tau species in GGT showed that detergent-insoluble, fibril-like tau contained the highest seeding activity, as reflected in its ability to increase tau aggregation in primary glial cultures. Taken together, our data suggest that unique seeding properties differentiate GGT-tau from other tauopathies, which provides new insight into pathogenic heterogeneity of primary neurodegenerative tauopathies.

Project description:Co-occurrence of tau and α-synuclein pathologies in a subset of Alzheimer's disease patients has led to the idea that mixed pathologies may play a unique characteristic role in the Alzheimer's disease neurodegenerative cascade. To understand the aetiology of such mixed pathologies, we investigated cross-seeding by human recombinant tau and human recombinant α-synuclein fibrillar species in a mouse model of tauopathy (Line PS19) or synucleinopathy (Line M20). Unilateral hippocampal injection of tau fibrils or α-synuclein fibrils, and to a lesser extent tau + α-synuclein copolymer fibrils prepared from co-incubating individual recombinant monomers, induced robust phosphorylated tau pathology in PS19 mice relative to control mice. Though the tau + α-synuclein copolymer fibrils did not modulate induction of pathologies at the site of injection, examination of the whole brain showed that these copolymers exacerbated neuroanatomic transmission of seeded tau pathology compared to tau fibril-injected mice. Only α-synuclein fibrils, but not tau alone or tau + α-synuclein copolymers, triggered modest levels of endogenous phosphorylated α-synuclein pathology. Overall, data from the PS19 mice suggest that human α-synuclein fibrils can efficiently cross-seed human tau and have a modest priming effect on mouse α-synuclein, and the presence of tau fibrils does not exacerbate the priming process. In M20 mice, unilateral hippocampal injection of α-synuclein fibrils or tau fibrils induced robust bilateral phosphorylated α-synuclein pathology, while tau + α-synuclein copolymer injection resulted in restricted phosphorylated α-synuclein pathology predominantly in the ipsilateral cortex. This suggests that human tau fibrils can also induce human α-synuclein pathogenesis, and the presence of combinatorial seeds is not synergistic. None of these aggregates induced phosphorylated tau pathology in M20 mice, showing that mouse tau cannot be primed efficiently by human tau fibrils or human α-synuclein fibrils. Neuropathological analysis of the whole brain of M20 mice showed that tau + α-synuclein copolymer-injected mice had lower abundance of bilaterally transmitted α-synuclein pathologies relative to α-synuclein fibril-injected mice. Thus, the tau + α-synuclein copolymer fibrils show robust transmission properties preferentially in rodent model of tauopathies but not in synucleinopathy, probably signifying an enhanced cooperative relationship between tau and α-synuclein in the tau seeding process. Together, our data highlight the unique cross-seeding properties of tau and αSyn in neurodegenerative proteinopathies.

Project description:BackgroundMAPT mutations cause neurodegenerative diseases such as frontotemporal dementia but, strikingly, patients with the same mutation may have different clinical phenotypes.MethodsGiven heterogeneities observed in a transgenic (Tg) mouse line expressing low levels of human (2 N, 4R) P301L Tau, we backcrossed founder stocks of mice to C57BL/6Tac, 129/SvEvTac and FVB/NJ inbred backgrounds to discern the role of genetic versus environmental effects on disease-related phenotypes.ResultsThree inbred derivatives of a TgTauP301L founder line had similar quality and steady-state quantity of Tau production, accumulation of abnormally phosphorylated 64-68 kDa Tau species from 90 days of age onwards and neuronal loss in aged Tg mice. Variegation was not seen in the pattern of transgene expression and seeding properties in a fluorescence-based cellular assay indicated a single "strain" of misfolded Tau. However, in other regards, the aged Tg mice were heterogeneous; there was incomplete penetrance for Tau deposition despite maintained transgene expression in aged animals and, for animals with Tau deposits, distinctions were noted even within each subline. Three classes of rostral deposition in the cortex, hippocampus and striatum accounted for 75% of pathology-positive mice yet the mean ages of mice scored as class I, II or III were not significantly different and, hence, did not fit with a predictable progression from one class to another defined by chronological age. Two other patterns of Tau deposition designated as classes IV and V, occurred in caudal structures. Other pathology-positive Tg mice of similar age not falling within classes I-V presented with focal accumulations in additional caudal neuroanatomical areas including the locus coeruleus. Electron microscopy revealed that brains of Classes I, II and IV animals all exhibit straight filaments, but with coiled filaments and occasional twisted filaments apparent in Class I. Most strikingly, Class I, II and IV animals presented with distinct western blot signatures after trypsin digestion of sarkosyl-insoluble Tau.ConclusionsQualitative variations in the neuroanatomy of Tau deposition in genetically constrained slow models of primary Tauopathy establish that non-synchronous, focal events contribute to the pathogenic process. Phenotypic diversity in these models suggests a potential parallel to the phenotypic variation seen in P301L patients.

Project description:BACKGROUND:The accumulation of pathological tau is the main component of neurofibrillary tangles and other tau aggregates in several neurodegenerative diseases, referred to as tauopathies. Recently, immunotherapeutic approaches targeting tau have been demonstrated to be beneficial in decreasing tauopathy in animal models. We previously found that passive immunotherapy with anti-tau antibody to human tau or expression of an anti-tau secreted single-chain variable fragment (scFv) in the central nervous system of a mouse model of tauopathy decreased but did not remove all tau-associated pathology. Although these and other studies demonstrate that conventional immunotherapeutic approaches targeting tau can influence tau pathogenesis, the majority of pathological tau remains in the cytosol of cells, not typically accessible to an extracellular antibody. Therefore, we reasoned targeting intracellular tau might be more efficacious in preventing or decreasing tauopathy. METHODS:By utilizing our anti-tau scFv, we generated anti-tau intrabodies for the expression in the cytosol of neurons. To enhance the degradation capacity of conventional intrabodies, we engineered chimeric anti-tau intrabodies fused to ubiquitin harboring distinct mutations that shuttle intracellular tau for either the proteasome or lysosomal mediated degradation. To evaluate the efficacy in delaying or eliminating tauopathy, we expressed our tau degrading intrabodies or controls in human tau transgenic mice by adeno-associated virus prior to overt tau pathology and after tau deposition. RESULTS:Our results demonstrate, the expression of chimeric anti-tau intrabodies significantly reduce tau protein levels in primary neuronal cultures expression human tau relative to a non-modified anti-tau intrabody. We found the expression of engineered tau-degrading intrabodies destined for proteasomal-mediated degradation are more effective in delaying or eliminating tauopathy than a conventional intrabody in aged human tau transgenic mice. CONCLUSION:This study, harnesses the strength of intrabodies that are amendable for targeting specific domains or modifications with the cell-intrinsic mechanisms that regulate protein degradation providing a new immunotherapeutic approach with potentially improved efficacy.

Project description:Alzheimer's disease (AD) is defined by the presence of intraneuronal neurofibrillary tangles (NFTs) composed of hyperphosphorylated tau aggregates as well as extracellular amyloid-beta plaques. The presence and spread of tau pathology through the brain is classified by Braak stages and thought to correlate with the progression of AD. Several in vitro and in vivo studies have examined the ability of tau pathology to move from one neuron to the next, suggesting a "prion-like" spread of tau aggregates may be an underlying cause of Braak tau staging in AD. Using the HEK293 TauRD-P301S-CFP/YFP expressing biosensor cells as a highly sensitive and specific tool to identify the presence of seed competent aggregated tau in brain lysate-i.e., tau aggregates that are capable of recruiting and misfolding monomeric tau-, we detected substantial tau seeding levels in the entorhinal cortex from human cases with only very rare NFTs, suggesting that soluble tau aggregates can exist prior to the development of overt tau pathology. We next looked at tau seeding levels in human brains of varying Braak stages along six regions of the Braak Tau Pathway. Tau seeding levels were detected not only in the brain regions impacted by pathology, but also in the subsequent non-pathology containing region along the Braak pathway. These data imply that pathogenic tau aggregates precede overt tau pathology in a manner that is consistent with transneuronal spread of tau aggregates. We then detected tau seeding in frontal white matter tracts and the optic nerve, two brain regions comprised of axons that contain little to no neuronal cell bodies, implying that tau aggregates can indeed traverse along axons. Finally, we isolated cytosolic and synaptosome fractions along the Braak Tau Pathway from brains of varying Braak stages. Phosphorylated and seed competent tau was significantly enriched in the synaptic fraction of brain regions that did not have extensive cellular tau pathology, further suggesting that aggregated tau seeds move through the human brain along synaptically connected neurons. Together, these data provide further evidence that the spread of tau aggregates through the human brain along synaptically connected networks results in the pathogenesis of human Alzheimer's disease.

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Proteopathic tau seeding predicts tauopathy in vivo.

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Proteopathic tau seeding predicts tauopathy in vivo.

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