Formation of aberrant phosphotau fibrillar polymers in neural cultured cells

Pérez, Mar; Hernández, Félix; Gómez-Ramos, Alberto; Smith, Mark A.; Perry, George; Ávila, Jesús

doi:10.1046/j.1432-1033.2002.02794.x

Cited by 97 publications

(62 citation statements)

References 42 publications

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“…Hyperphosphorylation of tau at these sites is associated with increased propensity to form tangles (Buee et al, 2000). Treatment with OA, a phophatase inhibitor, caused an increase in levels of tau phosphorylation at serine 199/202, as previously reported (Perez et al, 2002), but no significant increase in phosphorylation of tau at serine 262 or serine 396. However, this is not surprising, insofar as large variability in OA-induced phosphorylation among different tau sites has been observed previously.…”

Section: Chemical Induction Of Upr In Sh-sy5y Cells Did Not Affect Tasupporting

confidence: 78%

“…To induce the UPR, medium containing 10 µg/ml tunicamycin or 500 nM thapsigargin was added to differentiated cells in six-well plates. As a positive control for tau phosphorylation, some cells were also treated with 75 nM okadaic acid, a phos-phatase inhibitor (Perez et al, 2002;Xu et al, 2004). At specified times after treatment, cell samples were taken and analyzed for both UPR activity and changes in tau phospho-rylation state and solubility.…”

Section: Chemical Induction Of the Uprmentioning

confidence: 99%

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P1‐367: Transgenic mouse and cell culture models demonstrate a lack of mechanistic connection between endoplasmic reticulum stress and tau dysfunction

Robinson

Spatara

2010

Alzheimer's & Dementia

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In vivo aggregation of tau protein is a hallmark of many neurodegenerative disorders, including Alzheimer's disease (AD). Recent evidence has also demonstrated activation of the unfolded protein response (UPR), a cellular response to endoplasmic reticulum (ER) stress, in AD, although the role of the UPR in disease pathoge-nesis is not known. Here, three model systems were used to determine whether a direct mechanistic link could be demonstrated between tau aggregation and the UPR. The first model system used was SH-SY5Y cells, a neuronal cultured cell line that endogenously expresses tau. In this system, the UPR was activated using chemical stressors, tunicamycin and thapsigargin, but no changes in tau expression levels, solubility, or phosphorylation were observed. In the second model system, wild-type 4R tau and P301L tau, a variant with increased aggregation propensity, were heterologously overexpressed in HEK 293 cells. This overexpression did not activate the UPR. The last model system examined here was the PS19 transgenic mouse model. Although PS19 mice, which express the P301S variant of tau, display severe neurodegeneration and formation of tau aggregates, brain tissue samples did not show any activation of the UPR. Taken together, the results from these three model systems suggest that a direct mechanistic link does not exist between tau aggregation and the UPR. Keywordsunfolded protein response; Alzheimer's disease; SH-SY5Y; PS19; neurofibrillary tangles; paired helical filaments Alzheimer's disease (AD) is the most prevalent form of dementia; approximately 5.3 million Americans currently have AD, and the number increases each year as the average age in our society increases (Alzheimer's Association, 2009). Pathologically, the disease involves neuronal death accompanied by accumulation of two distinct types of protein aggregates, extracellular aggregates of amyloid β (Aβ) peptide, often referred to as plaques, and intracellular aggregates of tau protein, often referred to as neurofibrillary tangles (NFTs) or paired helical filaments (PHFs;Selkoe, 2001). It is unclear how Aβ aggregates and tau aggregates are related and which is more important in disease pathology (Lee, 2001). *Correspondence to: A.S. Robinson, 227 Colburn Laboratory, Department of Chemical Engineering, University of Delaware, Newark, DE, 19716. asr@udel.edu. HHS Public Access Author Manuscript Author ManuscriptAuthor Manuscript Author ManuscriptTau protein is a natively unfolded protein, abundant in neurons, which binds to and stabilizes microtubules. Tau has 79 potential phosphorylation sites, and its phosphorylation is regulated by phosphatases and kinases, including glycogen synthase kinase-3β (GSK-3β). Hyperphosphorylated tau is associated with the disease state, insofar as it shows decreased microtubule binding and an increased propensity for tangle formation (Buee et al., 2000). In contrast to Aβ aggregation, which is specifically associated with AD, tau aggregates form in multiple neurodegenerative disorders, collective...

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Section: Chemical Induction Of Upr In Sh-sy5y Cells Did Not Affect Tasupporting

confidence: 78%

Section: Chemical Induction Of the Uprmentioning

confidence: 99%

P1‐367: Transgenic mouse and cell culture models demonstrate a lack of mechanistic connection between endoplasmic reticulum stress and tau dysfunction

Robinson

Spatara

2010

Alzheimer's & Dementia

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“…Fibrillar aggregates of tau were previously observed in Chinese hamster ovary cells that had been transfected with triple mutant tau expression constructs (21). In a related study (22), combined treatments of SH-SY5Y cells with okadaic acid and 4-hydroxynonenal induced 2-3-nm-wide fibrillar tau polymers. The PHF-like filaments in our tissue culture system clearly differed from both of the above in that they were much longer, about 10 times wider, and had readily identifiable twisted structures (Fig.…”

Section: Discussionmentioning

confidence: 81%

β-Amyloid Induces Paired Helical Filament-like Tau Filaments in Tissue Culture

Ferrari

Hoerndli

Baechi

et al. 2003

Journal of Biological Chemistry

170

135

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Paired helical filaments (PHF) are the principal pathologic components of neurofibrillary tangles in Alzheimer's disease (AD). To reproduce the formation of PHF in tissue culture, we stably expressed human tau with and without pathogenic mutations in human SH-SY5Y cells and exposed them for 5 days to aggregated synthetic ␤-amyloid peptide (A␤ 42 ). This caused a decreased solubility of tau along with the generation of PHF-like tau-containing filaments. These were 20 nm wide and had periodicities of 130 -140 nm in the presence of P301L mutant tau or 150 -160 nm in the presence of wild-type tau. Mutagenesis of the phosphoepitope serine 422 of tau prevented both the A␤ 42 -mediated decrease in solubility and the generation of PHF-like filaments, suggesting a role of serine 422 or its phosphorylation in tau filament formation. Together, our data underscore a role of A␤ 42 in the formation of PHF-like filaments. Our culture system will be useful to map phosphoepitopes of tau involved in PHF formation and to identify and characterize modifiers of the tau pathology. Further adaptation of the system may allow the screening and validation of compounds designed to prevent PHF formation. Neurofibrillary tangles (NFT)1 are abundant in many neurodegenerative diseases, including Alzheimer's disease (AD) and frontotemporal dementia with Parkinsonism linked to chromosome 17 (FTDP-17) caused by mutations in the tau gene (1-3). NFT are composed of filamentous aggregates that include paired helical filaments (PHF) and narrow twisted ribbons. These are made of the microtubule-associated protein tau (4, 5). Tau in these filaments is hyperphosphorylated both at physiological sites and at additional, pathological sites. Phosphorylation of tau is associated with the dissociation of tau from microtubules and with its relocalization from axons to cell bodies and dendrites. Pathologic increases in this pool of soluble tau, along with conformational changes of the natively unfolded tau, are initial critical steps in the assembly of the more insoluble pathological tau filaments.Transgenic mice expressing pathogenic mutations of human tau generate a few low abundant NFT (6, 7), and both intracerebral microinjections of ␤-amyloid into P301L mutant tau transgenic mice and co-expression of mutant amyloid precursor protein greatly accelerate the rate of NFT formation in mice (8,9). These data established a mechanistic relationship of ␤-amyloid toxicity with NFT formation.To circumvent the inherent limitations of the above in vivo experiments (including poor breeding of doubly transgenic mice as well as stereotaxic techniques) and to establish PHF formation in tissue culture, we expressed both wild-type and mutant forms of human tau in human SH-SY5Y cells and exposed them to preparations of aggregated synthetic ␤-amyloid. We were able to produce bona fide PHF-like tau filaments within 5 days of tissue culture. EXPERIMENTAL PROCEDURESSite-directed Mutagenesis-The longest human 4-repeat tau isoform, htau40, was cloned into the pRc/RSV expression ve...

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“…Tau phosphorylation is upregulated by oxidative stress [55] and tau and neurofilaments are modified by the products of oxidative stress, including 4-hydroxy-2-nonenal (HNE) [56] and other cytotoxic carbonyls [57], leading to protein aggregation as NFTs [58]. Moreover, following oxidative modification, phosphorylated tau, but not other tau forms, polymerizes more easily [58,59].…”

mentioning

confidence: 99%

“…Additionally, once phosphorylated, tau and neurofilaments are vulnerable to modification by carbonyl products of oxidative stress [54,58,59] and, consequently, aggregation into fibrils [58]. Because phosphorylation has a pivotal role in the redox balance, it is not surprising that the oxidative-stress-associated activation of MAPK pathways leads to the phosphorylation of tau and neurofilaments [60,61,65].…”

mentioning

confidence: 99%