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Brenner, J.

Publications and source records attributed to Brenner, J..

13 recordsLinked to original sources

Somnus, a tunable, open-source platform for automated EEG sleep scoring in physiological and disease mouse models

Many studies using mouse models of human brain disorders rely on extended recordings of brain electrical activity through electroencephalogram, or EEG. The features that researchers measure in these recordings, such as seizures and epileptiform activity, are deeply influenced by whether the animal is awake, in non-rapid eye movement (NREM) sleep, or in rapid eye movement (REM) sleep. Because total sleep duration varies naturally across animals, rigorous experimental comparisons require classifying the sleep state of every few seconds of a recording. Manual annotation is highly labor-intensive, often requiring hours per subject. While automated scoring tools accelerate this process, they are typically trained exclusively on healthy cohorts and frequently fail when applied to disease models that exhibit atypical or fragmented sleep architecture. Somnus is an open-source Python package designed to automate rodent sleep staging using EEG, muscle activity (EMG), and optional video-tracking data. It includes a robust base model trained on diverse recordings from six independent laboratories, enabling out-of-the-box classification across various equipment setups without requiring prior manual labeling. A central feature of Somnus is its capacity for rapid adaptation to pathological sleep structure. The software provides a graphical interface that directs researchers to the specific epochs where the base model is least certain. Users can manually correct these targeted epochs in a built-in viewer by comparing the model's classification to the raw EEG signal and video, and then use their corrections to fine-tune the model. This workflow empowers researchers to generate accurate custom scorers for complex disease models or novel treatments with just a few minutes of manual correction, bypassing the need to hand-score entire experimental cohorts or develop their own models from scratch.

neuroscience↗

Nanoparticle Albumin-Bound Paclitaxel Targets Pulmonary Neutrophils

Nanoparticle albumin-bound paclitaxel (Abraxane) was among the first clinically approved nanomedicines and is a first-line chemotherapeutic. It enhances therapeutic efficacy and reduces side effects of paclitaxel. Despite its extensive clinical use, mechanisms of Abraxane's therapeutic effects are the subject of ongoing study. This work posits a new mechanism for Abraxane's effects in lung cancers and metastasis. Tracing the albumin component of Abraxane with radioisotopes or fluorophores, we find that Abraxane homes to the lungs. Abraxane's lung tropism is elevated by inflammation and is eliminated in complement protein C3 knockout mice, showing that complement pathway drives the uptake. Flow cytometry and histology show that neutrophils are the dominant cell type taking up Abraxane in the lungs. We demonstrate Abraxane's tropism to pulmonary neutrophils in mice and in human donor lungs. We note previously undocumented side effects associated with Abraxane's lung tropism, including pro-thrombotic and pro-inflammatory responses in acute lung inflammation. We show that Abraxane's efficacy against lung metastases is affected when lung uptake is eliminated by C3 knockout. Our results provide insight into why Abraxane is effective in lung cancer and metastases. These findings may be used to improve outcomes in patients receiving Abraxane and help design next-generation cancer nanomedicines.

bioengineering↗

Percolation-inspired criticality in complement activation: universal scaling and transport-limited complement surface amplification

The collective spatial phenomenon of complement protein opsonization on nanoparticle surfaces is a key component of the immune response to viruses, engineered nanoparticles, and diseased cells. Recent work showed this opsonization follows a sharp, percolation-like transition versus the spacing d between surface-bound attachment sites, leaving two open questions: 1) whether the transition exhibits hallmarks of true criticality, such as diverging susceptibility, and 2) whether it can be distinguished from an alternative first-order cooperative (Hill-type) process producing an equally sharp threshold without true criticality. Here, we resolve both questions using a hierarchical statistical-mechanics treatment spanning stochastic, mean-field, and spatial reaction-diffusion models. The variance of two order parameters, peak complement activity and activation lifetime, diverges near threshold and sharpens systematically with system size, the defining signature of a critical point rather than a smooth cooperative response. Extending the analysis across site spacing and intrinsic kinetic rate constants traces a two-dimensional locus of critical points with consistent critical exponents throughout, establishing a single, robust universality class. The mean-field dynamic exponent for activation lifetime agrees quantitatively with the exact value predicted for the general epidemic process. Finally, a reaction-diffusion model of the nanoparticle surface shows the critical locus is set by a diffusion-limited length scale, establishing complement percolation as a fundamentally transport-limited surface reaction. These results place complement activation within the percolation universality class and identify the physical parameters, diffusion, catalysis, and decay, that govern its critical threshold, with direct implications for rational design of complement-evading nanomaterials, immunology, and evolutionary biology.

biophysics↗

Cathepsin-dependent amyloid formation drives mechanical rupture of lysosomal membranes

Lysosomal membrane integrity is essential for cellular homeostasis, and its failure drives lysosomal storage disorders (LSD) and neurodegeneration. The dipeptide L-leucyl-L-leucine methyl ester (LLOMe) is widely used to model lysosomal damage, yet its mechanism remains poorly understood. The prevailing view holds that LLOMe polymerizes into membrane-permeabilizing peptide chains within the lysosomal lumen. Using cryo-electron tomography in cultured cells and primary neurons, we visualized the structural basis of LLOMe-induced lysosomal damage. We reveal that LLOMe forms amyloid structures within lysosomes that directly interact with and rupture the limiting membrane through mechanical stress. In vitro reconstitution confirms this amyloid-mediated mechanism. These findings establish a structural paradigm for lysosomal membrane disruption and provide insights into how disease-relevant protein aggregates, implicated in neurodegeneration and LSD, may compromise lysosomal integrity.

cell biology↗

Biomolecular condensate architecture of an autophagic cargo at molecular resolution in situ

Biomolecular condensates organise cellular biochemistry, yet their molecular architecture in situ remains poorly understood. During selective autophagy, macromolecules frequently accumulate into biomolecular condensates, forming discrete entities for autophagic engulfment and degradation - ideal systems for structural analysis. We employed in situ cryo-electron tomography to determine the near-atomic resolution structure of Aminopeptidase 1 condensates within cells. These condensates form densely packed, spherical assemblies with amorphous organisation and liquid-like properties, elucidating the requirements of a selective autophagic cargo for exclusive targeting. Structural analysis and multiscale simulations reveal that the short, transient -helical structures in the disordered N-terminus of Aminopeptidase 1 enable site-specific, coiled-coil-like interactions required for condensate formation and properties. A single point mutation that increases -helical propensity directly modulates condensate viscosity and dynamics from a liquid-like to a glass-like state, while preserving local molecular packing. Our results demonstrate that disordered regions encode both specificity and material properties through transiently structured motifs, linking sequence specificity to phase behaviour in cells and expanding the molecular logic of phase separation in cells.

cell biology↗

Exploring shaped focused ion beams for lamella preparation

Focused ion beams (FIB) are widely used instruments in transmission electron microscopy (TEM) sample preparation across scientific disciplines. Generally, site-specific ablation of material is achieved by scanning a highly focused probe across a selected area, leading to the removal of material. However, the geometries of TEM lamellae milled with the FIB are usually highly non-isometric, with their thickness generally being orders of magnitude smaller than their width and length. Here, we explore a changed probe shape for milling. Instead of using a Gaussian-like spot probe ion beam, we characterize the use of the stigmator as quasi-cylindrical lens to create a highly astigmatic beam that we term ion knife. Using the ion knife allows for material ablation by spreading the current over a larger area and changes the dimension of the probe with an anisotropic change in apparent beam resolution, as observed in spot burn cross-sections. We demonstrate a method to approximate beam shapes by imaging that allows for convenient alignment of parameters for beam shaping. Finally, exploring shaped probes in cryogenic lamella preparation, we demonstrate the feasibility of cellular lamella milling and sectioning of cryo-lift-out volumes with the ion knife.

biophysics↗

Making plant tissue accessible for cryo-electron tomography

Cryo-Electron Tomography (cryo-ET) allows to visualize the molecular architecture of pristinely preserved cells and tissues. The workflow of sample preparation for cryo-ET is rather complex; it involves vitrification by rapid freezing followed by cryo-Focused Ion Beam (FIB) milling rendering the volumes of interest thin enough for cryo-ET data acquisition. The established protocols for single cells grown on or deposited on EM-grids are not suitable for multicellular plant tissues. Plunge-freezing does not yield vitrified samples in most cases and must be replaced by high-pressure freezing. This, in turn, necessitates extensive modifications of the subsequent FIB milling procedures. In this communication we describe procedures for sample screening, targeted FIB milling guided by cryo-fluorescence microscopy and a novel lamella trimming step that allows to obtain homogenously thin lamellae suitable for cryo-ET. We have tested all the steps along the workflow with a variety of plant tissues including the moss Physcomitrium patens and tissues of Arabidopsis thaliana and Limonium bicolor. We could demonstrate that the workflow optimized for plant tissues allows to attain subnanometer resolution in cases where subtomogram averaging is applicable.

plant biology↗

TSC2 loss in neural progenitor cells suppresses translation of ASD/NDD-associated transcripts in an mTORC1- and MNK1/2-reversible fashion

Tuberous sclerosis complex (TSC) is an inherited neurodevelopmental disorder (NDD) with frequent manifestations of epilepsy and autism spectrum disorder (ASD). TSC is caused by inactivating mutations in TSC1 or TSC2 tumor suppressor genes, with encoded proteins hamartin (TSC1) and tuberin (TSC2) forming a functional complex inhibiting mechanistic target of rapamycin complex 1 (mTORC1) signaling. This has led to treatment with allosteric mTORC1 inhibitor rapamycin analogs ("rapalogs") for TSC tumors; however, rapalogs are ineffective for treating neurodevelopmental manifestations. mTORC1 signaling controls protein synthesis by regulating formation of the eIF4F complex, with further modulation by MNK1/2 kinases via phosphorylation of the eIF4F subunit eIF4E. While both these pathways modulate translation, comparing their impact on transcriptome-wide mRNA translation, as well as effects of inhibiting these pathways in TSC has not been explored. Here, employing CRISPR-modified, isogenic TSC2 patient-derived neural progenitor cells (NPCs), we have examined transcriptome-wide changes in mRNA translation upon TSC2 loss. Our results reveal dysregulated translation in TSC2-Null NPCs, which significantly overlaps with the translatome from TSC1-Null NPCs. Interestingly, numerous non-monogenic ASD-, NDD-and epilepsy-associated genes identified in patients harboring putative loss-of-function mutations, were translationally suppressed in TSC2-Null NPCs. Importantly, translation of these ASD- and NDD-associated genes was reversed upon inhibition of either mTORC1 or MNK1/2 signaling using RMC-6272 or eFT-508, respectively. This study establishes the importance of mTORC1-eIF4F- and MNK-eIF4E-sensitive mRNA translation in TSC, ASD and other neurodevelopmental disorders laying the groundwork for evaluating drugs in clinical development that target these pathways as a treatment strategy for these disorders.

molecular biology↗

Lysosomal storage disease proteo/lipidomic profiling using nMOST links ferritinophagy with mitochondrial iron deficiencies in cells lacking NPC2

Lysosomal storage diseases (LSDs) comprise [~]50 monogenic disorders marked by the buildup of cellular material in lysosomes, yet systematic global molecular phenotyping of proteins and lipids is lacking. We present a nanoflow-based multi-omic single-shot technology (nMOST) workflow that quantifies HeLa cell proteomes and lipidomes from over two dozen LSD mutants. Global cross-correlation analysis between lipids and proteins identified autophagy defects, notably the accumulation of ferritinophagy substrates and receptors, especially in NPC1-/- and NPC2-/- mutants, where lysosomes accumulate cholesterol. Autophagic and endocytic cargo delivery failures correlated with elevated lyso-phosphatidylcholine species and multi-lamellar structures visualized by cryo-electron tomography. Loss of mitochondrial cristae, MICOS- complex components, and OXPHOS components rich in iron-sulfur cluster proteins in NPC2-/- cells was largely alleviated when iron was provided through the transferrin system. This study reveals how lysosomal dysfunction affects mitochondrial homeostasis and underscores nMOST as a valuable discovery tool for identifying molecular phenotypes across LSDs.

cell biology↗

Thirty years of deforestation within the entire ranges of nine endangered lemur species (3 CR, 4 EN, 2 VU) in northwestern Madagascar

Forest cover change is of particular concern in tropical regions. In this study, we investigate the degree of deforestation in the entire ranges of nine highly threatened lemur species in northwestern Madagascar. Landsat satellite images were acquired from four different time stages (1990, 2000, 2011, 2020), classified into forest/non-forest, and changes quantified. Forest cover declined from 17.5% to 9.3% within the last 30 years. This decline varied across four protected areas (PAs) investigated: the forest cover of Ankarafantsika National Park (ANP) declined only moderately over time (from 76.3% to 67.4%), while it declined drastically in other PAs (e.g., from 54.9% to 18.9%, Bongolava Forest Corridor). Two lemur taxa are most affected (Lepilemur otto, Microcebus bongolavensis) by having only very few isolated forest patches left within their ranges (approximately 542.7 km{superscript 2}). For two other species (L. ahmansoni, L. aeeclis), most of the remaining forest is concentrated in two coastal PAs (in total 627.2 and 477.9 km{superscript 2}, respectively), while those species occurring inside ANP (5 taxa) experienced rather stable forest coverage until 2020. A reversal of these deforestation trends and active reforestation measures are desperately needed to reduce habitat loss for these nine lemur species. A practical experience-based guideline is therefore provided.

ecology↗

Period Doubling Bifurcations in a Forced Cell-Free Genetic Oscillator

Complex non-linear dynamics such as period doubling and chaos have been previously found in computational models of the oscillatory gene networks of biological circadian clocks, but their experimental study is difficult. Here, we present experimental evidence of period doubling in a forced synthetic genetic oscillator operated in a cell-free gene expression system. To this end, an oscillatory negative feedback gene circuit is established in a microfluidic reactor, which allows continuous operation of the system over extended periods of time. We first thoroughly characterize the unperturbed oscillator and find good agreement with a four-species ODE model of the system. Guided by simulations, microfluidics is then used to periodically perturb the system by modulating the concentration of one of the oscillator components with a given amplitude and frequency. When the ratio of the external zeitgeber period and the intrinisic period is close to 1, we experimentally find period doubling and quadrupling in the oscillator dynamics, whereas for longer zeitgeber periods, we find stable entrainment. Our theoretical model suggests favorable conditions for which the oscillator can be utilized as an externally synchronized clock, but also demonstrates that related systems could, in principle, display chaotic dynamics.

synthetic biology↗

Different Munc18 Proteins Mediate Baseline and Stimulated Airway Mucin Secretion

Airway mucin secretion is necessary for ciliary clearance of inhaled particles and pathogens, but can be detrimental in pathologies such as asthma and cystic fibrosis. Exocytosis in mammals requires a Munc18 scaffolding protein, and airway secretory cells express all three Munc18 isoforms. Using conditional airway epithelial deletant mice, we found that Munc18a has the major role in baseline mucin secretion, Munc18b has the major role in stimulated mucin secretion, and Munc18c does not function in mucin secretion. In an allergic asthma model, Munc18b deletion reduced airway mucus occlusion and airflow resistance. In a cystic fibrosis model, Munc18b deletion reduced airway mucus occlusion and emphysema. Munc18b deficiency in the airway epithelium did not result in any abnormalities of lung structure, particle clearance, inflammation, or bacterial infection. Our results show that regulated secretion in a polarized epithelial cell may involve more than one exocytic machine at the apical plasma membrane, and that the protective roles of mucin secretion can be preserved while therapeutically targeting its pathologic roles.

cell biology↗

Exploring the Diversity of Bacillus whole genome sequencing projects using Peasant, the Prokaryotic Assembly and Annotation Tool

BackgroundThe persistent decrease in cost and difficulty of whole genome sequencing of microbial organisms has led to a dramatic increase in the number of species and strains characterized from a wide variety of environments. Microbial genome sequencing can now be conducted by small laboratories and as part of undergraduate curriculum. While sequencing is routine in microbiology, assembly, annotation and downstream analyses still require computational resources and expertise, often necessitating familiarity with programming languages. To address this problem, we have created a light-weight, user-friendly tool for the assembly and annotation of microbial sequencing projects.\n\nResultsThe Prokaryotic Assembly and Annotation Tool, Peasant, automates the processes of read quality control, genome assembly, and annotation for microbial sequencing projects. High-quality assemblies and annotations can be generated by Peasant without the need of programming expertise or high-performance computing resources. Furthermore, statistics are calculated so that users can evaluate their sequencing project. To illustrate the computational speed and accuracy of Peasant, the SRA records of 322 Illumina platform whole genome sequencing assays for Bacillus species were retrieved from NCBI, assembled and annotated on a single desktop computer. From the assemblies and annotations produced, a comprehensive analysis of the diversity of over 200 high-quality samples was conducted, looking at both the 16S rRNA phylogenetic marker as well as the Bacillus core genome.\n\nConclusionsPeasant provides an intuitive solution for high-quality whole genome sequence assembly and annotation for users with limited programing experience and/or computational resources. The analysis of the Bacillus whole genome sequencing projects exemplifies the utility of this tool. Furthermore, the study conducted here provides insight into the diversity of the species, the largest such comparison conducted to date.

bioinformatics↗