Biology subjects

Ori, A.

Publications and source records attributed to Ori, A..

At least 19 recordsLinked to original sources

bioRxiv2026

A scalable human neuron model of Alzheimer's disease relevant tauopathy reveals mechanisms linking Tau fibrillization to synaptic dysfunction

Tauopathies, including Alzheimer's disease, are driven by pathological aggregation of hyperphosphorylated Tau, which disrupts synaptic integrity, impairs neuronal communication, and contributes to cognitive decline. To dissect tauopathy pathogenesis and enable therapeutic discovery, reliable and scalable human iPSC-neuron models are essential. Here, we developed two complementary iPSC-derived neuron models: an endogenous Tau seeding model, in which neurons are challenged with pre-formed Tau fragments that form paired helical filament (PHF)-consistent structures, and a Tau-0N3R overexpression seeding model to accelerate pathology. Both models recapitulate hallmark features of tauopathy, including the progressive formation of intracellular, hyperphosphorylated, sarkosyl-insoluble, and conformationally altered Tau aggregates (AT8, MC1 positive), along with synaptic and neuronal dysfunction. Cryogenic electron tomography (cryo-ET) further revealed the morphology of Tau fibrils within cells, as well as the ultrastructure of Tau fibrils trapping synaptic vesicles in situ. Using this platform, we performed integrated phosphoproteomics, high-content screening, and functional validation to identify key pathways driving Tau aggregation. MARK2-mediated phosphorylation within Tau's microtubule-binding domain emerged as an early trigger of aggregation, confirmed by site-specific mutagenesis. In parallel, small molecules targeting the PI3K/mTOR/GSK3 pathway reduced aggregation and restored synaptic function, with GSK3 inhibition lowering phosphorylation at critical aggregation-driving sites on Tau. Together, these findings establish a physiologically relevant, scalable platform for therapeutic screening that connects Tau seed uptake, site-specific phosphorylation, fibril formation, and synaptic disruption, ultimately identifying mechanistically separable intervention points along the aggregation cascade.

cell biology↗

bioRxiv2026

The PSD-95 interactome of hippocampal synapses potentiated in response to an in vivo learning task

Changes in the molecular composition of synapses underlie the functional changes in synaptic strength after learning. These phenomena have been studied either via imaging of individual candidate proteins, or via the global analysis of the synaptic proteome, lumping together all synapses, regardless of their activation or potentiation status. As a step towards the characterization of the proteome of synapses that have undergone potentiation following a learning paradigm, we have designed SynActive-PSD95-FLAG, a genetically encoded probe consisting of a FLAG-tagged PSD-95 endowed with activity-dependent local translation and expression enrichment at potentiating dendritic spines. SynActive-PSD95-FLAG was expressed via Adeno Associated Viral vectors (AAVs) in the hippocampus of mice which were exposed to contextual fear conditioning. Then, PSD95-FLAG immunoprecipitation from hippocampal extracts, followed by mass spectrometry, was used to characterize the changes in the PSD-95 interactome caused by in vivo synaptic potentiation. This revealed extensive rearrangements in the stoichiometry between PSD-95 and its direct and indirect interactors, highlighting proteins involved in peptide translation, including ribosomal components, and vesicle trafficking. Our results, obtained with a selectively expressed proteomic handle, provide new insight into the molecular substrates of hippocampal learning and a novel tool to study the proteome of potentiated synapses.

neuroscience↗

bioRxiv2026

Mitochondrial degradation of metallothionein enables local zinc mobilization during zinc limitation

Zinc is an essential structural and enzymatic cofactor for roughly 10% of proteins, including transcription factors, metabolic enzymes, and cytoskeletal components. It also supports critical functions across organelles such as gene regulation in the nucleus, protein folding in the endoplasmic reticulum, and energy production and antioxidant defense in mitochondria. Despite these indispensable roles, the cellular mechanism that recycles zinc to maintain homeostasis during zinc deficiency remains poorly understood. Here, we identify a biphasic response to zinc limitation, which involves the rapid degradation of the zinc-storing metallothionein followed by the degradation, in an autophagy-dependent manner, of other zinc-binding proteins. We show that metallothionein is rapidly imported into the mitochondria to be degraded by the mitoprotease LONP1. Zinc starvation leads to severe mitochondrial dysfunction and metallothionein degradation allows local zinc release to alleviate nutrient stress. Our results reveal a non-canonical, mitochondria-mediated degradation pathway for a nutrient-storing protein that mobilizes zinc locally to maintain metabolic homeostasis and establish mitochondria as active hubs for nutrient recycling.

cell biology↗

bioRxiv2026

Direct RNA Sequencing reveals epitranscriptomic regulation of brain cells and Alzheimer's Disease pathology

Alternative mRNA splicing and post-transcriptional RNA modification are key mechanisms that regulate transcript function; however, their role in neuronal activity and neurodegenerative disease remains poorly defined. In this study, we evaluated two nanopore-based long-read sequencing (LR-seq) formats: cDNA-PCR sequencing (CPS) and direct RNA sequencing (DRS). We then applied DRS to profile both full-length isoforms and RNA modifications in major brain cell types derived from induced pluripotent stem cells (iPSCs) and post-mortem Alzheimers disease (AD) brains. Relative to CPS, DRS achieved higher accuracy and sensitivity for transcript quantification, de novo transcript model construction, and open reading frame (ORF) annotation across neuropathological gene sets. Focusing on iPSC-derived neurons, we built a multi-omic atlas to connect transcriptional output with translational engagement and protein abundance, by integrating DRS-based mRNA abundance, N6-methyladenosine (m6A) status and poly(A) tail length with ribosome profiling (Ribo-seq) and mass spectrometry (MS). The combination of DRS and Ribo-seq data demonstrated synergism in predicting protein abundance. This analysis also uncovered a significant inverse relationship between m6A modification and mRNA abundance, which was dependent on the engagement of the ribosomal A-site. Lastly, we applied DRS to the epitranscriptomic analysis of AD brain samples, demonstrating that m6A profiles can be used to distinguish early-versus late-stage disease.

neuroscience↗

bioRxiv2026

Gene function and expression profiling in yeast spores, killifish diapause embryos, and their post-dormant offspring cells

Dormancy is a reversible cellular state characterised by suspended proliferation and increased stress resilience, enabling long-term viability under adverse conditions. Although dormant cells are critical for the life cycle of diverse organisms, from microbes to humans, they are understudied compared to proliferating cells. We present a comparative investigation of dormant cells in two divergent species: spores of fission yeast and diapause embryos of turquoise killifish. A genome-wide screen for genes affecting the lifespan and heat-shock resilience of spores uncovered a trade-off between longevity and heat resistance, and considerable differences in the genetic basis for lifespan between spores and chronologically aging yeast cells. RNA-seq and mass-spectrometry analyses revealed substantial transcriptomic and proteomic changes in spores and diapause embryos, with ribosomal proteins induced as transcripts but repressed as proteins. Transcriptomic regulation of biological processes, but less so of specific genes, is broadly conserved across yeast spores, killifish diapause, and human dormant cancer cells, including the induction of autophagy- and translation-related processes and the repression of cell cycle-related processes. Spores and diapause embryos modulate their transcriptomes and proteomes in response to heat stress and prolonged time. These RNA and protein expression changes are uncoupled and differ from aging-related expression signatures in yeast cells and adult fish. Cells derived from older or stressed spores retain phenotypic differences for several cell divisions, reflected in altered expression signatures, lifespan and stress resilience. Similarly, diapause duration and heat exposure are associated with long-term expression signatures in post-diapause embryos before hatching. This study highlights core biological processes and principles that are remarkably conserved in distinct types of dormant cells.

genetics↗

bioRxiv2026

Convergent molecular signatures of ageing and injury in the peripheral nervous system

Peripheral nervous system (PNS) ageing is marked by structural and functional decline, yet it remains unclear whether ageing constitutes a distinct biological programme or reflects a chronic injury-like state. To address this, we performed an unbiased, comparative molecular analysis of PNS ageing, neuroprotective dietary restriction (DR), and nerve injury. We conducted transcriptomic and proteomic profiling of peripheral nerves from young, old and geriatric mice fed ad libitum or subjected to long-term DR, and proteomics of nerves collected at multiple time points following injury. Age-associated molecular changes followed both linear and non-linear trajectories, and DR partially attenuated these ageing-related alterations. Notably, ageing-and injury-induced proteomic signatures showed considerable similarities, supporting the concept that an aged nerve resembles an injured nerve. Together, our study provides the most comprehensive molecular resource of PNS changes during ageing, DR, and injury, enabling the definition of key molecular signatures underlying PNS physiology. All datasets are integrated into the "PNS-omics Viewer", a Shiny web application designed to facilitate data mining of the herein presented datasets (tba).

neuroscience↗

bioRxiv2025

Oxidative stress causes a reversible decrease of deubiquitylases activity in old vertebrate brains

The ubiquitin-proteasome system is essential for neuronal proteostasis, and its activity declines with age. How deubiquitylating enzymes (DUBs) are affected by aging in the vertebrate brain remains unclear. Here, we profiled cysteine protease DUBs using activity-based proteomics in aging mouse and killifish brains. Despite stable protein levels, we identified a subset of DUBs that progressively lose catalytic activity with age. We demonstrated that oxidative stress impairs DUB function through thiol oxidation and that antioxidant treatment restores their activity in vitro and in vivo. Further, inhibition of DUBs in human iPSC-derived neurons significantly recapitulated ubiquitylation changes observed in aged brains, and temporal analysis in mice revealed that DUB inhibition precedes proteasome decline in the brain during aging. Together, these findings indicate a redox-sensitive subset of DUBs that undergo an age-associated decline in activity and suggest that impaired deubiquitylation is an early, yet potentially reversible, driver of proteostasis decline in the aging brain.

biochemistry↗

bioRxiv2025

Lonafarnib Partially Reverses Cardiac Senescence in Human and Mouse Progeria Models via Autophagy Activation

Hutchinson-Gilford Progeria Syndrome (HGPS), characterised by accelerated ageing, causes cardiovascular defects resembling aspects of cardiovascular ageing. We used human left ventricle cardiomyocytes (CMs) derived from HGPS-induced pluripotent stem cells (iPSCs), and their isogenic-corrected controls, to investigate HGPS-CM dysfunction and identify potential therapies. Our results revealed that HGPS-iPSC-CMs exhibit greater maturity and associated elevated oxidative stress compared to controls, which they could not contend with, leading to cellular senescence. Increased senescence was also observed in cardiac tissue from mouse and human physiologically-aged and HGPS individuals. Functionally, HGPS-iPSC-CMs showed dysregulated mitochondrial respiration and calcium handling. Amongst the six drugs tested, rapamycin and lonafarnib were the most effective against HGPS-cardiac phenotypes. Although lonafarnib raised safety concerns, it partially reverted the cardiac senescent phenotype by inducing cellular autophagy and decreasing progerin expression in progeroid mice. Our study supports the use of HGPS-iPSC-CMs to identify novel biomarkers and therapies for HGPS, and potentially cardiac physiological-ageing.

cell biology↗

bioRxiv2025

The microcephaly-associated protein YIPF5 differentially regulates ER-export

YIPF5 is a small ER-membrane protein implicated in ER-Golgi transport. Mutations in YIPF5 cause MEDS2 (microcephaly with simplified gyral pattern, epilepsy, and neonatal diabetes syndrome), a fatal disorder manifesting in early childhood. We demonstrate that YIPF5 is involved in ER export of a subset of proteins, including cargoes of the ER-export receptor SURF4, with which it directly interacts. In YIPF5 knockout cells, we observe a shift in the cell surface and secretome composition, marked by reduced neuronal adhesion molecules and increased secretion of ER chaperones influencing cell migration. YIPF5 depletion enhances cell migration in a wound-healing assay and alters SURF4 localization, causing elongated ERGIC53- and Rab1-positive tubules from COPII-labeled ER exit sites. Kinetic analysis suggests that YIPF5 negatively regulates SURF4-mediated ER export. In utero knockdown of Yipf5 in embryonic mouse brains induces premature neuronal migration and abnormal neuronal morphology. These findings suggest that YIPF5 and SURF4 coordinate ER export, and disruption may underlie cortical development defects leading to microcephaly.

cell biology↗

bioRxiv2024

The Aging Synapse: An Integrated Proteomic And Transcriptomic Analysis

An important hallmark of aging is the loss of proteostasis, which can lead to the formation of protein aggregates and mitochondrial dysfunction in neurons. Although it is well known that protein synthesis is finely regulated in the brain, especially at synapses, where mRNAs are locally translated in activity-dependent manner, little is known as to the changes in the synaptic proteome and transcriptome during aging. Therefore, this work aims to elucidate the relationship between transcriptome and proteome at soma and synaptic level during aging. Proteomic and transcriptomic data analysis reveal that, in young animals, proteins and transcripts are correlated and synaptic regulation is driven by changes in the soma. During aging, there is a decoupling between transcripts and proteins and between somatic and synaptic compartments. Furthermore, soma-synapse gradient of ribosomal genes changes upon aging, i.e. ribosomal transcripts are less abundant and ribosomal proteins are more abundant in synaptic compartment of old mice with respect to younglings. Additionally, transcriptomics data highlight a difference in the splicing of certain synaptic mRNA with aging. Taken together, our data provide a valuable resource for the study of the aging synapse.

neuroscience↗

bioRxiv2024

Cell-Type Resolved Protein Atlas of Brain Lysosomes Identifies SLC45A1-Associated Disease as a Lysosomal Disorder

Mutations in lysosomal genes cause neurodegeneration and neurological lysosomal storage disorders (LSDs). Despite their essential role in brain homeostasis, the cell-type-specific composition and function of lysosomes remain poorly understood. Here, we report a quantitative protein atlas of the lysosome from mouse neurons, astrocytes, oligodendrocytes, and microglia. We identify dozens of novel lysosomal proteins and reveal the diversity of the lysosomal composition across brain cell types. Notably, we discovered SLC45A1, mutations in which cause a monogenic neurological disease, as a neuron-specific lysosomal protein. Loss of SLC45A1 causes lysosomal dysfunction in vitro and in vivo. Mechanistically, SLC45A1 plays a dual role in lysosomal sugar transport and stabilization of V1 subunits of the V-ATPase. SLC45A1 deficiency depletes the V1 subunits, elevates lysosomal pH, and disrupts iron homeostasis causing mitochondrial dysfunction. Altogether, our work redefines SLC45A1-associated disease as a LSD and establishes a comprehensive map to study lysosome biology at cell-type resolution in the brain and its implications for neurodegeneration.

cell biology↗

bioRxiv2024

iSODA: A Comprehensive Tool for Integrative Omics Data Analysis in Single- and Multi-Omics Experiments

Omics technologies including genomics, proteomics, metabolomics, and lipidomics allow profound insights into health and disease. Thanks to plummeting costs of continuously evolving omics analytical platforms, research centers collect multi-omics data more routinely. They are, however, confronted with the lack of a versatile software solution to harmoniously analyze single-omics data and merge and interpret multi-omics data. We have developed iSODA, an interactive web-based application for the analysis of single-as well as multi-omics omics data. The software tool emphasizes intuitive, interactive visualizations designed for user-driven data exploration. Researchers can filter and normalize their datasets and access a variety of functions ranging from simple data visualization like volcano plots and PCA, to advanced functional analyses like enrichment analysis for proteomics and saturation analysis for lipidomics. For insights from integrated multi-omics, iSODA incorporates Multi-Omics Factor Analysis - MOFA, and Similarity Network Fusion - SNF. All results are presented in interactive plots with the possibility of downloading plots and associated data. The ability to adapt the imported data on-the-fly allows for tasks such as removal of outlier samples or failed features, various imputation strategies, or data normalization. The modular design allows for extensions with new analyses and plots. The software is accessible under http://isoda.online/. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/605811v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@1eb61c8org.highwire.dtl.DTLVardef@60a7feorg.highwire.dtl.DTLVardef@1b54b41org.highwire.dtl.DTLVardef@15fd1b2_HPS_FORMAT_FIGEXP M_FIG C_FIG Graphical summary for iSODA showcasing some application examples, the data import, the single-omics and multi-omics modules.

bioinformatics↗

bioRxiv2024

Polyamines sustain epithelial regeneration in aged intestines by modulating protein homeostasis

Aging hampers the regenerative potential of intestinal epithelium across species including humans, yet the underlying causes remain elusive. Here, using proteomic and metabolomic profiling of intestinal tissues together with functional assays, we characterized the temporal dynamics of regeneration following injury induced by 5-fluorouracil, a commonly used chemotherapeutic agent. Comparison of regeneration dynamics in mice of different ages revealed the emergence of a proteostasis stress signature and increased levels of polyamines following injury exclusively in old epithelia. Mechanistically, we show that delayed regeneration is an intrinsic feature of aged epithelial cells that display reduced protein synthesis and accumulation of ubiquitylated proteins. Notably, dietary restriction followed by re-feeding prior to injury increases polyamine pathway activation, enhances protein synthesis, and restores the regenerative capacity of aged intestines. Our findings highlight promising epithelial targets for interventions aimed at tackling the decline in tissue repair mechanisms associated with aging.

molecular biology↗

bioRxiv2024

The Neurolipid Atlas: a lipidomics resource for neurodegenerative diseases uncovers cholesterol as a regulator of astrocyte reactivity impaired by ApoE4

Lipid changes in the brain have been implicated in many neurodegenerative diseases including Alzheimers Disease (AD), Parkinsons disease and Amyotrophic Lateral Sclerosis. To facilitate comparative lipidomic research across brain-diseases we established a data commons named the Neurolipid Atlas, that we have pre-populated with novel human, mouse and isogenic induced pluripotent stem cell (iPSC)-derived lipidomics data for different brain diseases. We show that iPSC-derived neurons, microglia and astrocytes display distinct lipid profiles that recapitulate in vivo lipotypes. Leveraging multiple datasets, we show that the AD risk gene ApoE4 drives cholesterol ester (CE) accumulation in human astrocytes recapitulating CE accumulation measured in the human AD brain. Multi-omic interrogation of iPSC-derived astrocytes revealed that cholesterol plays a major role in astrocyte interferon-dependent pathways such as the immunoproteasome and major histocompatibility complex (MHC) class I antigen presentation. We show that through enhanced cholesterol esterification ApoE4 suppresses immune activation of astrocytes. Our novel data commons, available at neurolipidatlas.com, provides a user-friendly tool and knowledge base for a better understanding of lipid dyshomeostasis in neurodegenerative diseases.

neuroscience↗

bioRxiv2024

Leiomodin 1 promotes myogenic differentiation by modulating Sirtuin 1

During myogenic differentiation the cellular architecture and proteome of muscle stem cells and myoblasts undergo extensive remodeling. These molecular processes are only partially understood and display alterations in disease conditions as well as during aging resulting in impaired regeneration. Here, we used mass spectrometry to quantify the temporal dynamics of more than 6000 proteins during myogenic differentiation. We identified the actin nucleator leiomodin 1 (LMOD1) among a restricted subset of cytoskeletal proteins increasing in abundance in early phases of myogenic differentiation. We show that LMOD1 is already expressed by muscle stem cells in vivo and displays increased abundance during skeletal muscle regeneration, especially during early regeneration suggesting that LMOD1 is important for induction of myotube formation. Of note, knockdown of LMOD1 in primary myoblasts and during skeletal muscle regeneration severely affects myogenic differentiation, while overexpression accelerates and improves the initiation of myotube formation suggesting that LMOD1 is a critical component regulating myogenic differentiation. Mechanistically, we show that LMOD1 physically and functionally interacts with the deacetylase sirtuin1 (SIRT1), a regulator of myogenic differentiation, especially at the onset of myogenic differentiation. We demonstrate that LMOD1 influences SIRT1 localization and the expression of a subset of its target genes. Consistently, depletion or pharmacological inhibition of SIRT1 partially rescues the impairment of myogenic differentiation observed after knockdown of LMOD1. Our work identifies a new regulator of myogenic differentiation that might be targeted to improve muscle regeneration in aging and disease.

cell biology↗

bioRxiv2023

Remodeling of the protein ubiquitylation landscape in the aging vertebrate brain

Post-translational modifications (PTMs) regulate protein homeostasis and function. How aging affects the landscape of PTMs remains largely elusive. Here, we reveal changes in hundreds of protein ubiquitylation, acetylation, and phosphorylation sites in the aging brain of mice. We show that aging has a major impact on protein ubiquitylation and that 29% of the ubiquitylation sites are affected independently of protein abundance, indicating altered PTM stoichiometry. We found a subset of these sites to be also affected in the brain of the short-lived killifish Nothobranchius furzeri, highlighting a conserved aging phenotype. Furthermore, we estimated that over 35% of ubiquitylation changes observed in old mouse brains derive from partial proteasome inhibition, a well-established hallmark of brain aging. Our findings provide evidence of an evolutionarily conserved ubiquitylation signature of the aging brain and establish a causal link between proteasome inhibition and age-related remodeling of the ubiquitylome.

systems biology↗

bioRxiv2023

Transcriptome analysis of different strains of the turquoise killifish Nothobranchius furzeri identifies signatures associated with heritable lifespan differences

The killifish Nothobranchius furzeri is the shortest-lived vertebrate model organism and is characterized by laboratory strains differing in lifespan by more than a factor of two. The genetic architecture underlying this difference is complex and the pathways responsive remain elusive. We performed an analysis in N. furzeri of public transcritomic datasets comprehensive of four different tissues and one embryonic stage in two strains: the shorter-lived GRZ and the longer-lived MZM0410. Remarkably, the two strains differ in their transcriptome profile already at the embryonic stage. The short-lived strain GRZ shows an anticipated aging profile consistently in all tissues investigated, with differences in expression of aging-related genes detected already at sexual maturity. In addition, analysis of a longitudinal dataset revealed that genes whose expression is prognostic of longer lifespan at the individual level are also differentially expressed between strains at an early adult age, suggesting antagonistic pleiotropism.

genomics↗

bioRxiv2023

Automated workflow for BioID improves reproducibility and identification of protein-protein interactions

Proximity dependent biotinylation is an important method to study protein-protein interactions in cells, for which an expanding number of applications has been proposed. The laborious and time consuming sample processing has limited project sizes so far. Here, we introduce an automated workflow on a liquid handler to process up to 96 samples at a time. The automation does not only allow higher sample numbers to be processed in parallel, but also improves reproducibility and lowers the minimal sample input. Furthermore, we combined automated sample processing with shorter liquid chromatography gradients and data-independent acquisition to increase analysis throughput and enable reproducible protein quantitation across a large number of samples. We successfully applied this workflow to optimise the detection of proteasome substrates by proximity-dependent labelling.

biochemistry↗