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Biology subjects

Daniel, J.

Publications and source records attributed to Daniel, J..

13 recordsLinked to original sources

Learning from tandem mass spectra at scale with a self-supervised foundation model for proteomics

Mass spectrometry-based proteomics increasingly relies on machine learning, yet existing models are trained for defined supervised tasks such as peptide identification, de novo sequencing or fragment intensity prediction, limiting transfer across datasets, instruments and acquisition methods. Here we present InstaNovo-FM, a self-supervised foundation model for bottom-up proteomics trained to reconstruct masked regions of tandem mass spectra. We assemble a diverse training corpus spanning 1.63 billion MS/MS spectra and 184.6 million high-confidence annotations. We train an encoder-only transformer on the annotated tier using a physics-aware masked reconstruction objective. We demonstrate that the InstaNovo-FM embeddings encode fundamental experimental and biological properties, including fragmentation method, sequence properties and post-translational modifications, without requiring peptide labels. Furthermore, this foundation model directly enables diverse downstream applications, including de novo peptide sequencing, database-free identification and analytical run classification. InstaNovo-FM establishes a unified representation space for peptide fragmentation spectra, enabling robust transferability across the proteomics ecosystem.

bioinformatics↗

AlphaConformers: Structure-guided sampling enables prediction of multiple protein conformations

Proteins are dynamic molecules capable of adopting multiple conformations. However, AlphaFold2 predominantly generates models around a single conformation, usually representing a ligand-bound state. To address this limitation, we developed AlphaConformers, a structure-guided pipeline that steers AlphaFold2 toward alternative conformations. It is based on the idea that protein structure databases can capture the structural space accessible to members of a protein family. Given a target protein, AlphaConformers retrieves structures from structurally similar proteins. These structures are organized into structure-based alignments and template sets, which are supplied to AlphaFold2 as conformational hypotheses. The resulting models are clustered and filtered, facilitating their analysis. Evaluated on a curated benchmark of 88 proteins with known ligand-bound and unbound conformations, AlphaConformers expanded AlphaFold2 conformational sampling and recovered alternative states missed by AlphaFold2 and other state-of-the-art methods. AlphaConformers ranked first for modelling subtle conformational changes commonly observed between ligand-bound and unbound states. These results show that structural information from protein databases can be leveraged to steer AlphaFold2 toward alternative conformations.

bioinformatics↗

Glutamatergic synaptic resilience to overexpressed human alpha-synuclein

Alpha synuclein (aSyn) is an abundant protein that, in the brain, is concentrated in neuronal presynaptic boutons and associates with synaptic vesicles. aSyn is strongly linked with Parkinsons disease (PD) due to its accumulation in pathognomonic inclusions in neuronal cells, including in glutamatergic neurons. While increased expression of wild-type (WT) aSyn due to multiplications of the SNCA gene, or the expression of mutant forms of aSyn, can cause familial forms of PD, it is still unclear whether increased levels of aSyn alone are sufficient to impair synaptic function. Previous studies have used experimental systems that do not allow systematic characterisation of presynaptic physiology. To address this gap in research, we used lentiviral vectors to overexpress human aSyn (haSyn) in continental and autaptic glutamatergic neurons from rodent hippocampus and systematically analysed their presynaptic function. Virally-transduced neurons exhibited levels of expression of haSyn that mimic those associated with triplications of the SNCA gene in PD patients (2-fold increase in total aSyn), as determined using quantitative immunofluorescence imaging and immunoblots. Neuronal toxicity, neuronal morphology, and SNAP-25, a presynaptic protein, were not altered in continental cultures. Finally, a systematic characterization of autaptic neurons expressing haSyn exhibited no significant difference in any parameter of synaptic function, including basal properties of evoked and spontaneous neurotransmitter release, and synaptic plasticity compared to neurons infected with a control virus. These results indicate that rodent glutamatergic neurons are resilient to aSyn overexpression. In conclusion, our findings suggest neurotoxicity associated with aSyn overexpression is not universal, and that a deeper understanding of aSyn biology and pathobiology is necessary.

neuroscience↗

Rapid catecholamine trafficking regulates neutrophil functions and neutrophil-platelet interactions

Neutrophils are key effector cells of the innate immune system that respond to small signaling molecules regulating immune responses. For a long time, similarities between neuronal and immune cells have been discussed. Here, we show that human neutrophils rapidly take up, package and use catecholamine neurotransmitters such as dopamine or epinephrine via the machinery known from neurons. Uptake and release of catecholamines as well as trafficking and packaging into MPO/VMAT2-positive primary vesicles is visualized with false fluorescent neurotransmitters. We also directly image the fast (> 10 s) and transient release of catecholamines from neutrophils with near infrared fluorescent nanosensors. Serotonin or activated platelets trigger calcium (Ca2+) signaling and consequently exocytosis of catecholamines. They reduce NET-formation but increase platelet aggregation. Thus, we establish similarities between neurons and neutrophils and identify a paracrine neutrophil-platelet feedback loop relevant for inflammatory and coagulatory conditions.

immunology↗

Immunologic comparisons of strain and induction method in an improved mouse model of intrauterine fibrosis

Intrauterine adhesions are growths of fibrotic tissue within the uterine cavity and can arise from a variety of tissue-damaging stimuli. Immune cells are known to mediate fibrotic responses, but specific mechanisms require further elucidation. Here, we compared intrauterine fibrosis development and immune responses across different mouse strains and induction methods. We aimed to identify a consistent and more clinically relevant mouse model of intrauterine fibrosis, whether immune responses differ in response to different stimuli, and which potential key immune cell populations are responsible for intrauterine fibrosis susceptibility. Intrauterine fibrosis induction methods were compared using surgical curettage or transcervically administered chemical (quinacrine) models. Measurements of tissue morphology and collagen gene expression indicate BALB/c mice are more susceptible than C57BL/6 mice to intrauterine fibrosis. In chemically induced BALB/c uterine tissues, gene expression and flow cytometry data show greater pro-inflammatory macrophage responses, implicating a possible role in fibrogenesis consistent with human intrauterine adhesion data. Findings from this study demonstrate the importance of mouse strain selection in studies of intrauterine adhesions. Furthermore, we show that a new hormone-synchronized, chemically induced mouse model can more uniformly and reliably provoke fibrotic tissue response. This model may allow for greater elucidation of mechanisms involved in intrauterine adhesion development, and exploratory therapeutic studies for treatment intervention.

bioengineering↗

Dye-based Fluorescent Organic Nanoparticles, New Promising Tools for Optogenetics

Dye-based fluorescent organic nanoparticles are a specific class of nanoparticles obtained by nanoprecipitation in water of pure dyes only. While the photophysical and colloidal properties of the nanoparticles strongly depend on the nature of the aggregated dyes, their excellent brightness in the visible and in the near infrared make these nanoparticles a unique and versatile platform for in vivo application. This article examines the promising utilization of these nanoparticles for in vivo optogenetics applications. Their photophysical properties as well as their biocompatibility and their capacity to activate Chrimson opsin in vivo through fluorescence reabsorption process are demonstrated. Additionally, an illustrative example of employing these nanoparticles in fear reduction in mice through close-loop stimulation is presented. Through an optogenetic methodology, the nanoparticles demonstrate an ability to selectively manipulate neurons implicated in the fear response and diminish the latter. Dye-based fluorescent organic nanoparticles represent a promising and innovative strategy for optogenetic applications, holding substantial potential in the domain of translational neuroscience. This work paves the way for novel therapeutic modalities for neurological and neuropsychiatric disorders.

neuroscience↗

Air-liquid interface exposure of A549 human lung cells to characterize the hazard potential of a gaseous bio-hybrid fuel blend

Gaseous and semi-volatile organic compounds emitted by the transport sector contribute to air pollution and have adverse effects on human health. To reduce harmful effects to the environment as well as to humans, renewable and sustainable bio-hybrid fuels are explored and investigated in the cluster of excellence "The Fuel Science Center" at RWTH Aachen University. However, data on the effects of bio-hybrid fuels on human health is scarce, leaving a data gap regarding their hazard potential. To help close this data gap, this study investigates potential toxic effects of a Ketone-Ester-Alcohol-Alkane (KEAA) fuel blend on A549 human lung cells. Experiments were performed using a commercially available air-liquid interface exposure system which was optimized beforehand. Then, cells were exposed at the air-liquid interface to 50-2000 ppm C3.7 of gaseous KEAA for 1 h. After a 24 h recovery period in the incubator metabolic activity and cytotoxicity of cells were assessed. Our data support the international occupational exposure limits of the single KEAA constituents and moreover indicate no adverse effect to A549 cells when exposed to a fuel mixture. This finding applies only to the exposure scenario tested in this study and is difficult to extrapolate to the complex in vivo situation.

pharmacology and toxicology↗

Single cell resolution of SARS-CoV-2 tropism, antiviral responses, and susceptibility to therapies in primary human airway epithelium

The human airway epithelium is the initial site of SARS-CoV-2 infection. We used flow cytometry and single cell RNA-sequencing to understand how the heterogeneity of this diverse cell population contributes to elements of viral tropism and pathogenesis, antiviral immunity, and treatment response to remdesivir. We found that, while a variety of epithelial cell types are susceptible to infection, ciliated cells are the predominant cell target of SARS-CoV-2. The host protease TMPRSS2 was required for infection of these cells. Importantly, remdesivir treatment effectively inhibited viral replication across cell types, and blunted hyperinflammatory responses. Induction of interferon responses within infected cells was rare and there was significant heterogeneity in the antiviral gene signatures, varying with the burden of infection in each cell. We also found that heavily infected secretory cells expressed abundant IL-6, a potential mediator of COVID-19 pathogenesis.

microbiology↗

Functional dispersion of wetland birds, invertebrates and plants more strongly influenced by hydroperiod than each other

The relative role of biological and abiotic filters on the assembly of co-occurring taxa is widely debated. While some authors point to biological interactions (e.g., competition) as the stronger driver of ecological selection, others assert that abiotic conditions are more important because they filter species at the regional level. Because communities influenced by a dominant abiotic filter, (e.g., Prairie Pothole Region (PPR) wetlands, each varying in ponded water permanence), often have strong cross-taxon relationships, we can study these communities to better understand the relative influence of abiotic vs biotic filters on community structure. Using functional dispersion as our measure of communities, we test six alternate hypotheses about the relative importance of various pathways representing influence of biological and permanence filters on birds, aquatic macroinvertebrates and wetland plants in the northwest PPR using structural equation modeling. We aimed to understand whether: 1) ponded water permanence alone explained functional dispersion; 2) the influence of permanence on functional dispersion was direct or mediated; and 3) abiotic filtering by permanence was stronger than biotic filtering by co-occurring taxa. The best model suggests that there is a direct influence of permanence on the functional dispersion of each taxonomic group and that both bird and macroinvertebrate functional dispersion are causally related to plant functional dispersion, though for invertebrates the influence of plants is much less than that of permanence. Thus, the relative importance of wetland permanence and the functional dispersion of co-occurring taxa depends on which taxon is considered in PPR wetlands.

ecology↗

A Rapid, Cost-Effective Tailed Amplicon Method for Sequencing SARS-CoV-2

The global COVID-19 pandemic has led to an urgent need for scalable methods for clinical diagnostics and viral tracking. Next generation sequencing technologies have enabled large-scale genomic surveillance of SARS-CoV-2 as thousands of isolates are being sequenced around the world and deposited in public data repositories. A number of methods using both short- and long-read technologies are currently being applied for SARS-CoV-2 sequencing, including amplicon approaches, metagenomic methods, and sequence capture or enrichment methods. Given the small genome size, the ability to sequence SARS-CoV-2 at scale is limited by the cost and labor associated with making sequencing libraries. Here we describe a low-cost, streamlined, all amplicon-based method for sequencing SARS-CoV-2, which bypasses costly and time-consuming library preparation steps. We benchmark this tailed amplicon method against both the ARTIC amplicon protocol and sequence capture approaches and show that an optimized tailed amplicon approach achieves comparable amplicon balance, coverage metrics, and variant calls to the ARTIC v3 approach and represents a cost-effective and highly scalable method for SARS-CoV-2 sequencing.

genomics↗

Analytical Validation of a COVID-19 qRT-PCR Detection Assay Using a 384-well Format and Three Extraction Methods

The COVID-19 global pandemic is an unprecedented health emergency. Insufficient access to testing has hampered effective public health interventions and patient care management in a number of countries. Furthermore, the availability of regulatory-cleared reagents has challenged widespread implementation of testing. We rapidly developed a qRT-PCR SARS-CoV-2 detection assay using a 384-well format and tested its analytic performance across multiple nucleic acid extraction kits. Our data shows robust analytic accuracy on residual clinical biospecimens. Limit of detection sensitivity and specificity was confirmed with currently available commercial reagents. Our methods and results provide valuable information for other high-complexity laboratories seeking to develop effective, local, laboratory-developed procedures with high-throughput capability to detect SARS-CoV-2.

pathology↗

Emergence of stealth polymorphs that escape α-synuclein amyloid monitoring, take over and acutely spread in neurons

The conformational strain diversity characterizing -synuclein (-syn) amyloid fibrils is possibly at the origin of the different clinical presentations of synucleinopathies. Experimentally, various -syn fibril polymorphs have been obtained from distinct fibrillization conditions by altering the medium constituents and were selected by amyloid monitoring using the probe Thioflavin T (ThT). We report here that besides classical ThT positive products, fibrillization in saline simultaneously gives rise to competing fibril polymorphs that are invisible to ThT (stealth polymorphs), and that can take over. Due to competition, spontaneous generation of such stealth polymorphs bears on the apparent fibrillization kinetics and on the final plateau values. Their emergence has thus been ignored so far or mistaken for fibrillization inhibitions/failures. Compared to their ThT-positive counterparts, and as judged from their chemical shift resonances fingerprint, these new stealth polymorphs present a yet undescribed atomic organization and show an exacerbated propensity (approx. 20-fold) towards self-replication in cortical neurons. They also trigger a long distance synucleinopathic spread along nigro-striatal projections in vivo. In order to rapidly screen fibrillization products for the presence of such stealth polymorphs, we designed a simple multiplexed assay that can be easily and rapidly operated. This assay allows us to demonstrate the sustainability of the conformational replication of these novel and particularly invasive strains. It should also be of help to avoid erroneous upstream interpretations of fibrillization rates based on sole ThT, and to expedite further structural and functional characterization of stealth amyloid assemblies. One Sentence Summarystealth -synuclein fibrils take over

neuroscience↗

A MULTI-PURPOSE WORM TRACKER BASED ON FIM

The analysis of behavioural traits of Caenorhabditis elegans is an important method for understanding neuromuscular functions and diseases. Since C. elegans is a small and translucent animal which conducts a variety of complex movement patterns many different imaging and tracking protocols are used for different behavioural traits. Thus a unified multi-purpose imaging and tracking system for multiple behavioural assays would be favourable to improve statistical strength and comparability. Here we present a novel worm tracking toolbox based on the FIM (Frustrated total internal reflection (FTIR) based Imaging Method) system incorporating a variety of different behavioural assays into a single imaging and tracking setup.\n\nFirst, we apply the FTIR-based imaging method to C. elegans, thus we are able to improve the overall image quality compared to state of the art recording techniques. This method is easy to use and can be utilised to image animals during crawling on agar and trashing in water. Second, we extended the existing FIMTrack software to extract skeleton-based posture and motion features of multiple worms with very high accuracy in a comparatively large field-of-view. Third, we integrated a variety of different assays into this system. We carried out chemotaxis assays both with attractant and repellent chemicals. A novel electrotaxis dome compatible with FIM allows locomotion analyses that are not corrupted by random aberrations in unrestricted movement. Additionally, the FIM based worm tracker is able to analyse thrashing behaviour of multiple worms automatically with a high accuracy. Finally we demonstrate the capacity of the FIM based worm tracker to observe GFP signals in C. elegans worms. We tested our new C. elegans tracking suite with mutant strains of the ubiquitin-fold modifier 1 (Ufm1) cascade. We identified intermediate chemosensory phenotypes in Ufm1 cascade mutants which were previously undetected.

animal behavior and cognition↗