Biology subjects

Biasetti, L.

Publications and source records attributed to Biasetti, L..

4 recordsLinked to original sources

bioRxiv2025

A validated panel of commercial antibodies for reliable detection of FET proteins.

The FET protein family comprises the highly conserved RNA-binding proteins FUS, EWS and TAF15 which are implicated in RNA metabolism and neurodegenerative diseases such as ALS and FTLD. Despite their structural similarity, reliable detection of individual FET proteins remains challenging due to antibody cross-reactivity and inconsistent localisation patterns reported in the literature. To address this, we systematically evaluated 23 commercially available antibodies for specificity and performance across western blotting, immunocytochemistry, and immunohistochemistry. Using single and double shRNA knockdowns in HeLa cells, we confirmed target engagement and identified antibodies which were specific as well as those with significant cross-reactivity. Immunofluorescence in rat primary neurons and HeLa cells showed antibody and cell type dependent variations in nuclear and cytoplasmic localisation; from these, we identified a subset that demonstrated high-quality, region-specific staining in postmortem human brain tissue. Our findings highlight substantial variability in antibody performance and underscore the need for rigorous validation to ensure reproducibility in FET protein research. We present a validated panel of antibodies suitable for diverse applications, providing a critical resource for studies of FET protein biology and pathology.

neuroscience↗

bioRxiv2022

Dityrosine cross-links are present in Alzheimer's disease-derived tau oligomers and paired helical filaments (PHF) which promotes the stability of PHF-core tau (297-391) in vitro.

A characteristic hallmark of Alzheimers Disease (AD) is the pathological aggregation and deposition of tau into paired helical filaments (PHF) in neurofibrillary tangles (NFTs). Oxidative stress is an early event during AD pathogenesis and is associated with tau-mediated AD pathology. Oxidative environments can result in the formation of covalent dityrosine crosslinks that can increase protein stability and insolubility. Dityrosine cross-linking has been shown to occur in vivo in A{beta} plaques and -synuclein aggregates in Lewy bodies, and this modification may increase the insolubility of these aggregates and their resistance to degradation. Using the PHF-core tau fragment (residues 297 - 391) as a model, we have previously demonstrated that dityrosine formation traps tau assemblies to reduce further elongation. However, it is unknown whether dityrosine crosslinks are found in tau deposits in vivo in AD and its relevance to disease mechanism is unclear. Here, using transmission electron microscope (TEM) double immunogold-labelling, we reveal that neurofibrillary NFTs in AD are heavily decorated with dityrosine crosslinks alongside tau. Single immunogold-labelling TEM and fluorescence spectroscopy revealed the presence of dityrosine on AD brain-derived tau oligomers and fibrils. Using the tau (297-391) PHF-core fragment as a model, we further showed that prefibrillar tau species are more amenable to dityrosine crosslinking than tau fibrils. Dityrosine formation results in heat and SDS stability of oxidised prefibrillar and fibrillar tau assemblies. This finding has implications for understanding the mechanism governing the insolubility and toxicity of tau assemblies in vivo.

biochemistry↗

bioRxiv2021

SARS-CoV-2 variants of concern Alpha, Beta, Gamma and Delta have extended ACE2 receptor host-ranges

Following the emergence of SARS-CoV-2 in China in late 2019 a number of variants have emerged, with two of these - Alpha and Delta - subsequently growing to global prevalence. One characteristic of these variants are changes within the Spike protein, in particular the receptor binding domain (RBD). From a public health perspective these changes have important implications for increased transmissibility and immune escape; however, their presence could also modify the intrinsic host-range of the virus. Using viral pseudotyping we examined whether the variants of concern (VOCs) Alpha, Beta, Gamma and Delta have differing host ACE2 receptor usage patterns, focusing on a range of relevant mammalian ACE2 proteins. All four VOCs were able to overcome a previous restriction for mouse ACE2, with demonstrable differences also seen for individual VOCs with rat, ferret or civet ACE2 receptors, changes which we subsequently attribute to N501Y and E484K substitutions within the Spike RBD.

microbiology↗

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↗