Biology subjects

Serpell, L.

Publications and source records attributed to Serpell, L..

13 recordsLinked to original sources

bioRxiv2025

A proteomic analysis of the PHF-forming tau fragment (tau297-391) following uptake into differentiated human neuronal SHSY5Y cells

Tau self-assembly and intracellular deposition is associated with a group of neurodegenerative diseases called tauopathies, which include Alzheimers disease (AD) and Picks disease. Here, we measured the proteome response in human neuronal cells (differentiated SH-SY5Y) following addition of a spontaneously amyloidogenic region of tau known as dGAE (tau297-391) which forms AD-like paired helical filaments in vitro and proteomic analysis showed increased endogenous tau expression. Further interactome analysis uncovered increased association between tau and proteins associated with nuclear chromatin, the nucleolus, and the spliceosome as well as the thiol-peroxidase, PRDX6 alongside an increase in reactive oxygen species (ROS). This work highlights a method to identify proteome pathways that may play an important role in the development of tau pathology.

cell biology↗

bioRxiv2025

The architecture of amyloid fibrils formed by a human tau-derived hexapeptide VQIVYK

The sequence 306VQIVYK311 is an aggregation prone region of the tau protein implicated in driving assembly of tau into paired helical filaments. These filaments accumulate as intraneuronal neurofibrillary tangles in Alzheimers disease and a range of tauopathies. Here, we demonstrate that VQIVYK forms highly ordered fibrillar samples after prolonged incubation at room temperature. Remarkably, aligned fibre bundles give rise to unusually detailed and highly oriented X-ray fibre diffraction patterns. Analysis of these patterns provide a model of the core protofilament structure that satisfies the experimental diffraction data. This structural model is consistent with data from X-ray crystallography of microcrystals and conforms to the cross-beta architecture that defines amyloid, but diffraction data analysis shows a highly twisted filamentous protofilament architecture. Analysis of individual fibril envelopes by 3D contact point reconstruction atomic force microscopy reveals a diverse polymorphous population with a major fibril morphology of apparent cylindrical fibrils, and morphological subpopulations of fibrils with clear left-hand twisting patterns, despite the commonality of the core structure indicated by the detailed diffraction pattern. Together, these data suggest that VQIVYK amyloid fibrils form a polymorphous amyloid population by assembly of highly ordered protofilaments and provides molecular information regarding amyloid fibril twist.

biophysics↗

bioRxiv2025

Inhibiting disulphide bonding in truncated tau297-391 results in enhanced self-assembly of tau into seed-competent assemblies.

Tau undergoes fibrillogenesis in a group of neurodegenerative diseases termed tauopathies. Each tauopathy is characterized by tau fibrils with disease-specific conformations, highlighting the complexity of tau self-assembly. This has led to debate surrounding the precise mechanisms that govern the self-assembly of tau in disease, especially the involvement of disulphide bonding (DSB) between cysteine residues. In this study, we use a truncated form of tau, dGAE, capable of forming filaments identical to those in disease. We reveal the impact of DSB in dGAE assembly and propagation by resolving the global mechanisms that dominate its assembly. We found evidence for surface-mediated secondary nucleation and fragmentation being active in dGAE assembly. The inhibition of DSB during dGAE assembly leads to an enhanced aggregation rate through a reduced lag phase, but with no effect on the global assembly mechanisms. We suggest this is due to the formation a dominant, seed-competent species in the absence of DSB that facilitates elongation and secondary nucleation resulting in enhanced assembly. In vitro seeding assays reveal the recruitment of endogenous tau in a cell model only when using dGAE species formed under conditions that inhibit DSB. Our results further support the use of the in vitro dGAE tau aggregation model for investigating the mechanism of tau assembly, the effect of varying conditions on tau assembly and how these conditions affect the resultant species. Further studies may utilise dGAE and its aggregates to investigate tau seeding, propagation and to highlight or test potential targets for therapies that reduce the spread of pathologic tau throughout the brain.

biochemistry↗

bioRxiv2023

Generation of an induced pluripotent stem cell line from a healthy adult indigenous Nigerian participant

Genetic backgrounds contribute to cellular phenotypes, drug responsiveness, and health outcomes. However, the majority of human induced pluripotent stem cell (iPSC) lines are derived from individuals of European descent. Thus, there is a major, unmet need in the generation, characterisation, and distribution of iPSCs from diverse ancestries. To begin to address this need, we have generated iPSCs from dermal fibroblasts isolated from a healthy 60-year-old indigenous Nigerian male belonging to the Babur ethnic group. The iPSCs were generated using Sendai virus, and copy number variation (CNV) analysis revealed no new major abnormalities compared to the parental fibroblasts. The iPSCs have been characterised for pluripotency markers and morphology and successfully differentiated into neural progenitor cells and astrocytes. This iPSC line could serve as a healthy control in comparative studies and can be used in disease modelling, toxicity assessments, genetic analyses, and drug discovery processes within an African genetic background. To bolster the inclusion of African models in biomedical research, this iPSC line will be made available to the broader scientific community. Ongoing efforts focus on generating more lines from diverse indigenous populations towards creating a dedicated open-access African iPSC biobank.

cell biology↗

bioRxiv2021

The disease associated Tau35 fragment has an increased propensity to aggregate compared to full-length tau

Tau35 is a truncated form of tau found in human brain in a subset of tauopathies. Tau35 expression in mice recapitulates key features of human disease, including progressive increase in tau phosphorylation, along with cognitive and motor dysfunction. The appearance of aggregated tau suggests that Tau35 may have structural properties distinct from those of other tau species that could account for its pathological role in disease. To address this hypothesis, we performed a structural characterization of monomeric and aggregated Tau35 and compared the results to those of two longer isoforms, 2N3R and 2N4R tau. We used small angle X-ray scattering to show that Tau35, 2N3R and 2N4R tau all behave as disordered monomeric species but Tau35 exhibits higher rigidity. In the presence of the poly-anion heparin, Tau35 increases thioflavin T fluorescence significantly faster and to a greater extent than full-length tau, demonstrating a higher propensity to aggregate. We used atomic force microscopy, transmission electron microscopy and X-ray fiber diffraction to demonstrate that Tau35 aggregates are morphologically similar to previously reported tau fibrils but they are more densely packed. These data increase our understanding of the aggregation inducing properties of clinically relevant tau fragments and their potentially damaging role in the pathogenesis of human tauopathies.

biophysics↗

bioRxiv2020

Oxidative stress conditions result in trapping of PHF-core tau (297-391) intermediates.

The self-assembly of tau into paired helical filaments (PHF) in neurofibrillary tangles (NFTs) is a significant event in Alzheimers disease (AD) pathogenesis. Oxidative stress, which accompanies AD, induces multiple post-translational modifications in proteins including the formation of dityrosine (DiY) cross-links, previously observed for A{beta}. Here, metal catalysed- and ultra-violet oxidation were used to study the influence of DiY cross-linking on the self-assembly of the PHF-core tau fragment. We report that DiY cross-linking facilitates tau assembly into tau oligomers and prevents their elongation into filaments. The DiY cross-linked tau oligomers were not toxic to cells. Our findings suggest that DiY cross-linking of pre-assembled tau, promotes the formation of soluble tau oligomers that show no acute toxicity.

biochemistry↗

bioRxiv2020

Synaptic dysfunction caused by truncated tau is associated with hyperpolarization-activated cyclic nucleotide-gated channelopathy

Progressive neurodegeneration in tauopathies is mediated through an elusive mechanism. Here, we show that hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are functionally linked to disease-associated abnormalities in tau. Selective rises in the proportion of HCN-positive neurons are detected both in post-mortem human brain from Alzheimers disease and in the Tau35 mouse model of tauopathy. Tau35 mice develop progressive abnormalities including increased phosphorylated tau, enhanced HCN channel expression and decreased dendritic branching, as well as reduced synapse density that is accompanied by vesicle clustering defects. Notably, altered spine density and increased HCN channel expression in Tau35 neurons correlates with functional abnormalities in network properties, including enhanced hyperpolarization-induced membrane voltage sag and changes in the frequency and kinetics of spontaneous excitatory postsynaptic currents. Our findings are consistent with pathological changes in tauopathies impacting on HCN channels to drive network-wide structural and functional synaptic deficits, providing new targets for therapeutic intervention.

neuroscience↗

bioRxiv2020

Dityrosine cross-link trapping of amyloid-β intermediates reveals that self-assembly is required for Aβ-induced cytotoxicity

Multiple chemical reactions, such as the production of reactive oxygen species (ROS) can lead to dityrosine (DiY) formation via the cross-linking of closely spaced tyrosine residues and this can serve as a marker for aging. Amyloid-{beta} (A{beta}) has been found to be DiY cross-linked in the brains of AD patients. In vitro, A{beta} forms DiY cross-links via metal-catalysed oxidation (Cu2+ and H202) (MCO) leading to the formation of fibrils that are resistant to formic acid denaturation. However, copper is well known to influence and enhance self-assembly. Here, to investigate the interplay between self-assembly and DiY cross-linking we have utilised a non-assembly competent variant of A{beta} (vA{beta}). MCO and UV oxidation experiments using vA{beta} and wild-type A{beta}, revealed that DiY cross-linking stabilises, but does not induce or promote A{beta} assembly. Cu2+ alone, without H202, facilitates the formation and DiY cross-linking of wild-type A{beta} into long-lived oligomers. Our work reveals DiY formation halts further A{beta} self-assembly. DiY cross-linked A{beta} is non-toxic to neuroblastoma cells at all stages of self-assembly in contrast to oligomeric non-cross-linked A{beta}. These findings point to a mechanism of toxicity that necessitates continuing self-assembly of the A{beta} peptide, whereby trapped DiY A{beta} assemblies and assembly incompetent variant A{beta} are unable to result in cell death.

biochemistry↗

bioRxiv2020

Three-dimensional reconstruction of individual helical nano-filament structures from atomic force microscopy topographs

Atomic force microscopy, AFM, is a powerful tool that can produce detailed topographical images of individual nano-structures with a high signal-to-noise ratio without the need for ensemble averaging. However, the application of AFM in structural biology has been hampered by the tip-sample convolution effect, which distorts images of nano-structures, particularly those that are of similar dimensions to the cantilever probe tips used in AFM. Here we show that the tip-sample convolution results in a feature-dependent and non-uniform distribution of image resolution on AFM topographs. We show how this effect can be utilised in structural studies of nano-sized upward convex objects such as spherical or filamentous molecular assemblies deposited on a flat surface, because it causes magnification of such objects in AFM topographs. Subsequently, this enhancement effect is harnessed through contact-point based deconvolution of AFM topographs. Here, the application of this approach is demonstrated through the 3D reconstruction of the surface envelope of individual helical amyloid filaments without the need of cross-particle averaging using the contact-deconvoluted AFM topographs. Resolving the structural variations of individual macromolecular assemblies within inherently heterogeneous populations is paramount for mechanistic understanding of many biological phenomena such as amyloid toxicity and prion strains. The approach presented here will also facilitate the use of AFM for high-resolution structural studies and integrative structural biology analysis of single molecular assemblies.

biophysics↗

bioRxiv2020

Quantification of amyloid fibril polymorphism by nano-morphometry reveals the individuality of filament assembly

Amyloid fibrils are highly polymorphic structures formed by many different proteins. They provide biological function but also abnormally accumulate in numerous human diseases. The physicochemical principles of amyloid polymorphism are not understood due to lack of structural insights at the single-fibril level. To identify and classify different fibril polymorphs and to quantify the level of heterogeneity is essential to decipher the precise links between amyloid structures and their functional and disease associated properties such as toxicity, strains, propagation and spreading. Employing gentle, force-curve based AFM, we produce detailed images, from which the 3D reconstruction of individual filaments in heterogeneous amyloid samples is achieved. Distinctive fibril polymorphs are then classified by hierarchical clustering, and sample heterogeneity is objectively quantified. These data demonstrate the polymorphic nature of fibril populations, provide important information regarding the energy landscape of amyloid self-assembly, and offer quantitative insights into the structural basis of polymorphism in amyloid populations.

biophysics↗

bioRxiv2019

Misfolded Amyloid β-42 induced impairment of the endo-lysosomal pathway revealed by real-time optical monitoring

Misfolding and aggregation of proteins is strongly linked to several neurodegenerative diseases but how such species bring about their cytotoxic actions remains poorly understood. Here we used specifically-designed optical reporter probes and live fluorescence imaging in primary hippocampal neurons to characterise the mechanism by which prefibrillar, oligomeric forms of the Alzheimers-associated peptide, A{beta}42, exert their detrimental effects. We used a pH-sensitive reporter, A{beta}42-CypHer, to track A{beta} internalisation in real time, demonstrating that oligomers are rapidly taken up into cells in a dynamin-dependent manner, and trafficked via the endo-lysosomal pathway resulting in accumulation in lysosomes. In contrast, a non-assembling variant of A{beta}42 (vA{beta}42) assayed in the same way is not internalised. Tracking ovalbumin uptake into cells using CypHer or Alexa Fluor tags shows that preincubation with A{beta}42 significantly disrupts protein uptake. Our results identify a potential mechanism by which amyloidogenic aggregates directly impair cellular function through disruption of the endosomal-lysosomal pathway.

neuroscience↗

bioRxiv2018

Short-term fasting selectively influences impulsivity in healthy individuals

Previous research has shown that short-term fasting in healthy individuals (HIs) is associated with changes in risky decision-making. The current experiment was designed to examine the influence of short-term fasting in HIs on four types of impulsivity: reflection impulsivity, risky decision-making, delay aversion, and action inhibition. HIs were tested twice, once when fasted for 20 hours, and once when satiated. Participants demonstrated impaired action inhibition when fasted; committing significantly more errors of commission during a food-related Affective Shifting Task. Participants also displayed decreased reflection impulsivity when fasted, opening significantly more boxes during the Information Sampling Task (IST). There were no significant differences in performance between fasted and satiated sessions for risky decision-making or delay aversion. These findings may have implications for understanding eating disorders such as Bulimia Nervosa (BN). Although BN has been characterised as a disorder of poor impulse control, inconsistent findings when comparing individuals with BN and HIs on behavioural measures of impulsivity question this characterisation. Since individuals with BN undergo periods of short-term fasting, the inconsistent findings could be due to differences in the levels of satiation of participants. The current results indicate that fasting can selectively influence performance on the IST, a measure of impulsivity previously studied in BN. However, the results from the IST were contrary to the original hypothesis and should be replicated before specific conclusions can be made.

neuroscience↗

bioRxiv2017

Formation of functional, non-amyloidogenic fibres by recombinant Bacillus subtilis TasA

Bacterial biofilms are communities of microbial cells encased within a self-produced polymeric matrix. In the Bacillus subtilis biofilm matrix the extracellular fibres of TasA are essential. Here a recombinant expression system allows interrogation of TasA, revealing that monomeric and fibre forms of TasA have identical secondary structure, suggesting that fibrous TasA is a linear assembly of globular units. Recombinant TasA fibres form spontaneously, and share the biological activity of TasA fibres extracted from B. subtilis, whereas a TasA variant restricted to a monomeric form is inactive and subjected to extracellular proteolysis. The biophysical properties of both native and recombinant TasA fibres indicate that they are not functional amyloid-like fibres. A gel formed by TasA fibres can recover after physical shear force, suggesting that the biofilm matrix is not static and that these properties may enable B. subtilis to remodel its local environment in response to external cues. Using recombinant fibres formed by TasA orthologues we uncover species variability in the ability of heterologous fibres to cross-complement the B. subtilis tasA deletion. These findings are indicative of specificity in the biophysical requirements of the TasA fibres across different species and/or reflect the precise molecular interactions needed for biofilm matrix assembly.\n\nContributionsConceived and designed the experiments: CE, EE, RG, CEM, RJM, MS, NSW; Performed the experiments: KB, LC, CE, EE, PKF, RG, CEM, RJM, MS, TS; Contributed new analytical tools: CE, EE, RG, TS; Analysed the data: CE, EE, CEM, RJM, MS, LCS, NSW; Wrote the paper: EE, RJM, CEM, MS, NSW.

microbiology↗