Tauopathies are a group of neurodegenerative diseases categorised into three types, 3R, 4R, or 3R+4R (mixed) tauopathies, based on the tau isoforms that make up the aberrant filaments. It is supposed that all six tau isoforms share functional characteristics. However, differences in the neuropathological features associated with different tauopathies offer the possibility that disease progression and tau accumulation may vary depending on the isoform composition. The presence or absence of repeat 2 (R2) in the microtubule-binding domain defines the type of isoform, which might influence tau pathology associated with a particular tau isoform. Therefore, our study aimed to identify the differences in the seeding propensities of R2 and repeat 3 (R3) aggregates using HEK293T biosensor cells. We show that the seeding induced by R2 was generally higher than by R3 aggregates, and lower concentrations of R2 aggregates are sufficient to induce seeding. Next, we found that both R2 and R3 aggregates dose-dependently increased triton-insoluble Ser262 phosphorylation of native tau, which is only visible in cells seeded with higher concentrations (12.5 nM or 100 nM) of R2 and R3 aggregates, despite the seeding by the lower concentrations of R2 aggregates after 72 h. However, the accumulation of triton-insoluble pSer262 tau was visible earlier in cells induced with R2 than in R3 aggregates. Our findings suggest that the R2 region may contribute to the early and enhanced induction of tau aggregation and define the difference in disease progression and neuropathology of 4R tauopathies.
Emerging experimental evidence suggests tau pathology spreads between neuroanatomically connected brain regions in a prion-like manner in Alzheimer's disease (AD). Tau seeding, the ability of prion-like tau to recruit and misfold naïve tau to generate new seeds, is detected early in human AD brains before the development of major tau pathology. Many antitumour drugs have been reported to confer protection against neurodegeneration, supporting the repurposing of approved and experimental or investigational oncology drugs for AD therapy. In this study, we evaluated whether antitumour drugs that abrogate the generation of seed-competent aggregates of tau Repeat 3 (R3) domain peptides can prevent tau seeding and toxicity in Tau-RD P301S FRET Biosensor cells and Caenorhabditis elegans. We demonstrate that drugs that interact with the N-terminal VQIVYK or the C-terminal region housing the Cys322 prevent R3 dimerisation, abolishing the generation of prion-like R3 seeds. Preformed R3 seeds (fibrils) capped with, or R3 seeds formed in the presence of VQIVYK- or Cys322-targeting drugs have a reduced potency to cause aggregation of naïve tau in biosensor cells and protect worms from aggregate toxicity. These findings indicate that VQIVYK- or Cys322-targeting drugs may act as prophylactic agents against tau seeding.
Tauopathies are characterised by intracellular deposits of fibrillar tau tangles. However, the interneuronal spread of pathological tau species precedes the development of major tau burdens. Two amyloid motifs, VQIINK in repeat 2 and VQIVYK in repeat 3, of tau repeat domain, assemble into β-sheet-rich fibrils on their own but alone do not form seed-competent fibrils. In contrast, the entire R3 region self-aggregates and forms seed-competent fibrils. Our study aimed to identify the minimal regions in the tau repeat domain that define seeding and its impact on intracellular tau phosphorylation and aggregation. Using peptides of individual repeats, we show that R2, like R3, forms seed-competent fibrils when assembled in the presence of heparin. However, R3, but not R2, forms seed-competent fibrils when assembled without heparin, even though both R2 and R3 have identical N-terminal hexapeptide and cysteine residue sequences. Moreover, cysteine to alanine substitution in R3 abrogates its self-aggregation and seeding potency. Tau RD P301S biosensor cells and Tau P301L (0N4R)-expressing HEK293 cells seeded with R2 and R3 fibrils show the induction of pathological phosphorylation of tau at Ser262/Ser396/Ser404 positions and oligomerisation of native tau. Protein fractions of biosensor cells seeded with R2 and R3 fibrils reseed endogenous tau aggregation when introduced into a fresh set of biosensor cells. Our findings suggest that R3 may be the minimal region for pathological seed generation under physiological conditions, whereas R2 might need polyanionic cofactors to generate pathogenic seeds. Lastly, R2 and R3 fibrils induce template-induced misfolding and pathological hyperphosphorylation of intracellular tau, making intracellular tau seed-competent.
- MeSH
- Alzheimerova nemoc * metabolismus MeSH
- cystein MeSH
- HEK293 buňky MeSH
- heparin MeSH
- lidé MeSH
- proteiny tau genetika metabolismus MeSH
- tauopatie * metabolismus patologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Photodynamic therapy (PDT) has been approved as proper and effective kind of treatment for certain types of cancer and non-malignant diseases. We tested photodynamic effects on G361 human melanoma cells sensitized by zinc-5,10,15,20-tetrakis(4-sulphonatophenyl) porphyrine (ZnTPPS(4)), chloraluminium phtalocyanine disulfonate (ClAlPcS(2)) and 5-aminolevulinic acid (ALA). In particular, we examined the PDT efficiency depending on applied light dose (0.8; 1.7; 3.3; 6.6; 13.2; 26.4Jcm(-2)). The DNA gel electrophoresis, methylthiazol tetrazolium bromide (MTT) viability test, fluorescent microscopy using calcein AM and propidium iodide (PI) staining, and rhodamine 123 mitochondrial membrane potential assay were performed to detect and evaluate the cell death process. We also measured the time course of reactive oxygen species (ROS) production and its dependence on sensitizer concentration within continuously irradiated sensitized cells. In conclusion, these results demonstrate most significant phototoxic effect of ClAlPcS(2)-PDT in spite of significantly higher ROS production induced by ZnTPPS(4)-PDT on G361 cells. On the other hand, ALA-PDT has a minimal photoeffect and induces negligible ROS formation in G361 cells at the conditions described below.
- MeSH
- barvicí látky MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- fluorescenční barviva MeSH
- fluorescenční mikroskopie MeSH
- fotochemoterapie MeSH
- fotosenzibilizující látky farmakologie MeSH
- fragmentace DNA účinky léků účinky záření MeSH
- indoly farmakologie MeSH
- kyselina aminolevulová farmakologie MeSH
- lidé MeSH
- melanom experimentální terapie MeSH
- membránové potenciály účinky léků MeSH
- metaloporfyriny farmakologie MeSH
- mitochondriální membrány účinky léků MeSH
- nádorové buněčné linie MeSH
- organokovové sloučeniny farmakologie MeSH
- propidium MeSH
- reaktivní formy kyslíku metabolismus MeSH
- rhodamin 123 MeSH
- světlo MeSH
- tetrazoliové soli MeSH
- thiazoly MeSH
- vztah dávky záření a odpovědi MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
