Biology subjects

Katsinelos, T.

Publications and source records attributed to Katsinelos, T..

9 recordsLinked to original sources

bioRxiv2026

Oligodendrocytes may provide the extra density of α-synuclein filaments from multiple system atrophy

Prion-like templated seeding of a-synuclein filaments is thought to underlie the spreading of multiple system atrophy (MSA) and other synucleinopathies through the brain. Here, we used -synuclein filaments extracted from the brains of individuals with MSA to induce seeded aggregation in undifferentiated human neuroblastoma SH-SY5Y cells, human iNeurons, mixed primary murine cortical cells and murine oligodendrocyte-enriched primary cultures, all overexpressing human -synuclein, followed by determination of the cryo-EM structures of the seeded filaments. Whereas MSA seeds comprise two protofilaments with an unidentified non-proteinaceous density located between protofilaments, seeding resulted in the formation of filaments with a single protofilament. Only the seeded structures derived from oligodendrocytes contained an extra density that resembled that of the seeds. This suggests that oligodendrocytes may provide the chemical component that is responsible for the extra density at the protofilament interface of MSA filaments. In support, the injection of MSA seeds into the striatum of transgenic mice expressing human -synuclein under the control of the oligodendrocyte-specific proteolipid protein promoter, also yielded seeded filaments with a single protofilament and a similar extra density.

neuroscience↗

bioRxiv2026

Templated seeding induces the formation of Alzheimer paired helical tau filaments in human brain organoids

The prion concept postulates that seeded aggregation results in the recruitment of soluble protein monomers and their assembly into amyloid filaments with the same structures as those of the seeds. Here we show that this concept applies to tau filaments formed in cortical organoids obtained from human iPSCs of control subjects. Tau seeds were extracted from the frontal cortex of individuals with Alzheimer disease. When added to human cortical organoids expressing endogenous levels of three-repeat tau, they gave rise to filaments of three-repeat tau whose formation required endogenous tau expression. By cryo-EM, these seeded tau filaments were identical to the paired helical filaments of the seeds. This novel finding demonstrates the faithful templated seeding of tau in human nerve cells expressing endogenous levels of tau and opens the way to studying the mechanisms of paired helical filament formation and their downstream effects in a human system amenable to genetic modification.

neuroscience↗

bioRxiv2024

Twelve phosphomimetic mutations induce the assembly of recombinant full-length human tau into paired helical filaments

The assembly of tau into amyloid filaments is associated with more than twenty neurodegenerative diseases, collectively termed tauopathies. Cryo-EM structures of brain-derived tau filaments revealed that specific structures define different diseases, triggering a quest for the development of experimental model systems that replicate the structures of disease. Here, we describe twelve phosphomimetic, serine/threonine to aspartate, mutations in tau, which we termed PAD12, that collectively induce the in vitro assembly of full-length tau into filaments with the same structure as that of paired helical filaments extracted from the brains of individuals with Alzheimers disease. Solution-state nuclear magnetic resonance spectroscopy suggests that phosphomimetic mutations in the carboxy-terminal domain of tau may facilitate filament formation by disrupting an intramolecular interaction between two IVYK motifs. PAD12 tau can be used for both nucleation-dependent and multiple rounds of seeded assembly in vitro, as well as for the seeding of tau biosensor cells. PAD12 tau can be assembled into paired helical filaments under various shaking conditions, with the resulting filaments being stable for extended periods of time. They can be labelled with fluorophores and biotin. Tau filaments extracted from the brains of individuals with Alzheimers disease have been known to be made of hyperphosphorylated and abnormally phosphorylated full-length tau, but it was not known if the presence of this post-translational modification is more than a mere correlation. Our findings suggest that hyperphosphorylation of tau may be sufficient for the formation of the Alzheimer tau fold. PAD12 tau will be a useful tool for the study of molecular mechanisms of neurodegeneration.

neuroscience↗

bioRxiv2024

Drosophila appear resistant to trans-synaptic tau propagation

Alzheimers disease (AD) is the most common cause of dementia in the elderly, prompting extensive efforts to pinpoint novel therapeutic targets for effective intervention. Among the hallmark features of AD is the development of neurofibrillary tangles comprised of hyperphosphorylated tau protein, whose progressive spread throughout the brain is associated with neuronal death. Trans-synaptic propagation of tau has been observed in mouse models and indirect evidence for tau spread via synapses has been observed in human AD. Halting tau propagation is a promising therapeutic target for AD, thus a scalable model system to screen for modifiers of tau spread would be very useful for the field. To this end, we sought to emulate the trans-synaptic spread of human tau (hTau) in Drosophila melanogaster. Employing the trans-Tango circuit mapping technique, we investigated whether tau spreads between synaptically connected neurons. Immunohistochemistry and confocal imaging were used to look for tau propagation. Examination of hundreds of flies expressing 4 different human tau constructs in two distinct neuronal populations reveal a robust resistance in Drosophila to the trans-synaptic spread of hTau. This resistance persisted in lines with concurrent expression of amyloid-{beta}, in lines with global hTau knock-in to provide a template for human tau in downstream neurons, and with manipulations of temperature. These negative data are important for the field as we establish that Drosophila expressing human tau in subsets of neurons are unlikely to be useful to perform screens to find mechanisms to reduce the trans-synaptic spread of tau. The inherent resistance observed in Drosophila may serve as a valuable clue, offering insights into strategies for impeding tau spread in future studies.

neuroscience↗

bioRxiv2023

Characterization and super-resolution imaging of small tau aggregates in human samples.

Hyperphosphorylation and aggregation of the microtubule binding protein tau plays a key role in the development of Alzheimers disease. While the structure of the filamentous aggregates formed in humans has recently been determined to atomic resolution, there is far less information available about the smaller aggregate precursors, thought to be the most neurotoxic. To address this gap, we have developed a single molecule pull-down (SiMPull) able to detect tau aggregates in clinically relevant human samples. This method enables the detection and characterisation of individual tau aggregates, as opposed to averaged features obtained from traditional bulk techniques. We report the number, size and shape of individual aggregates measured via super-resolution microscopy, revealing disease-specific differences in tau aggregate morphology. By adapting the assay to simultaneously detect multiple phosphorylation sites in individual aggregates, we were also able to derive compositional profiles for pathological modifications present in individual aggregates. We demonstrate that tau aggregates in Alzheimers disease are significantly more likely to contain both the AT8 and T181 pathological phosphorylation markers, rather than only one. Together, tau SiMPull identified distinct subpopulations of large, modified tau aggregates that were invisible to traditional methodologies. These morphological and compositional differences distinguish samples taken from disease cohorts, offering to illuminate underlying disease mechanisms, and providing a foundation for novel diagnostic strategies.

biochemistry↗

bioRxiv2021

Tau assemblies enter the cytosol of neurons in a cholesterol sensitive manner

Accumulating evidence supports a prion-like mechanism in the spread of assembled tau in neurodegenerative diseases. Prion-like spread is proposed to require the transit of tau assemblies to the interior of neurons in order to seed aggregation of native, cytosolic tau. This process is poorly understood and remains largely hypothetical. Here, we develop sensitive techniques to quantify the cytosolic entry of tau in real-time. We find that tau does not promote its own entry but, rather, is wholly dependent on cellular machinery. We find that entry to the widely used reporter cell line HEK293 requires clathrin whereas entry to neurons does not. Cholesterol depletion or knockdown of cholesterol transport protein Niemann-Pick type C1 in neurons renders cells highly vulnerable to cytosolic entry and seeded aggregation. Our findings establish entry as the rate-limiting step in seeded aggregation and demonstrate that dysregulated cholesterol, a feature of several neurodegenerative diseases, potentiates tau aggregation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=170 SRC="FIGDIR/small/449238v3_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@1849023org.highwire.dtl.DTLVardef@13129eorg.highwire.dtl.DTLVardef@2b4736org.highwire.dtl.DTLVardef@15038ac_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG

cell biology↗

bioRxiv2021

Microglia become hypofunctional and release metalloproteases and tau seeds after phagocytosing live neurons with P301S tau aggregates

The microtubule-associated protein tau aggregates in multiple neurodegenerative diseases, causing inflammation and changing the inflammatory signature of microglia by unknown mechanisms. We have shown that microglia phagocytose live neurons containing tau aggregates cultured from P301S tau transgenic mice due to neuronal tau aggregate-induced exposure of the eat me signal phosphatidylserine. Here we show that after phagocytosis, microglia become hypophagocytic while releasing seed-competent insoluble tau aggregates. These microglia activate acidic {beta}-galactosidase, and release senescence-associated cytokines and matrix remodeling enzymes alongside tau, indicating a senescent phenotype. In particular, the marked NF{kappa}B-induced activation of matrix metalloprotease 3 (MMP3/stromelysinl) was replicated in the brains of P301S mutant tau transgenic mice, and in human brains from tauopathy patients. These data show that microglia that have been activated to ingest live neurons with tau aggregates behave hormetically, becoming hypofunctional while acting as vectors of tau aggregate spreading.

neuroscience↗

bioRxiv2021

Tau assemblies do not behave like independently acting prion-like particles in mouse neural tissue

A fundamental property of infectious agents is their particulate nature: infectivity arises from independently-acting particles rather than as a result of collective action. Assemblies of the protein tau can exhibit seeding behaviour, potentially underlying the apparent spread of tau aggregation in many neurodegenerative diseases. Here we ask whether tau assemblies share with classical pathogens the characteristic of particulate behaviour. We used organotypic hippocampal slice cultures from P301S tau transgenic mice in order to precisely control the concentration of extracellular tau assemblies. Whilst untreated slices displayed no overt signs of pathology, exposure to tau assemblies could result in the formation of intraneuronal, hyperphosphorylated tau structures. However, seeding ability of tau assemblies did not titrate in a one-hit manner in neural tissue. The results suggest that seeding behaviour of tau only arises at supra-physiological concentrations, with implications for the interpretation of high-dose intracranial challenge experiments and the possible contribution of seeded aggregation to human disease.

neuroscience↗

bioRxiv2019

Disassembly of Tau fibrils by the human Hsp70 disaggregation machinery generates small seeding-competent species

The accumulation of amyloid Tau aggregates is implicated in Alzheimers disease and other Tauopathies. Molecular chaperones are known for their function in maintaining protein homeostasis by preventing the formation or promoting the disaggregation of amorphous and amyloid protein aggregates. Here we show that an ATP-dependent human chaperone system disassembles Tau fibrils in vitro. This function is mediated by the core chaperone Hsc70, assisted by specific co-chaperones, in particular class B J-domain proteins and an Hsp110-type NEF. Recombinant fibrils assembled from all six Tau isoforms as well as Sarkosyl-resistant Tau aggregates extracted from cell culture were processed by the Hsp70 disaggregation machinery, demonstrating the ability of this machinery to recognize a broad range of Tau aggregates. Chaperone treatment released monomeric, and small oligomeric Tau species, which induced the aggregation of self-propagating Tau species in a Tau cell culture model. We infer from these results that the activity of the Hsp70 disaggregation machinery is a double-sided sword as it attempts to eliminate Tau amyloids but with the price of generating new seeds. The Hsp70 disaggregase therefore has a crucial function in the Tau propagation cycle, rendering it a potential drug target in Tauopathies.

molecular biology↗