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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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At least 523 records · Page 29Linked to original sources

Biophysical boundaries for dormancy release in sunflower achenes.

Reactions leading to dormancy release (DR) in "dry", orthodox seeds are still poorly understood, as well as their dependence on moisture content and temperature. Sunflower achenes were used to explore the effects of MC combined with a wide range of storage temperatures (ST{degrees}) on DR dynamics, tested at 10 and 25{degrees}C. Embryo sensitivity to abscisic acid, oxygen uptake and ageing indicators were followed and complemented with predicted viability loss dynamics. Water status was addressed with sorption isotherms and the glass transition temperature (Tg). Small variations in MC and ST{degrees} had profound effects on achene dormancy and embryo responsiveness to ABA. Using a temperature by humidity coordinate system we mapped three regions allowing DR within the glassy, rubbery and fluid states. Full DR (allowing germination at 10 and 25{degrees}C) occurred only in the rubbery state (above Tg), and optimal ST{degrees} for full DR dropped from +30 to-18{degrees}C with MC increasing from 0.045 to 0.1 gH2O.gDW-1. Differently, under both glassy and fluid states, DR was incomplete (allowing germination at 25{degrees}C but not at 10{degrees}C) and required warm ST{degrees} which also promoted ageing. HighlightSunflower achenes display diverse mechanisms for dormancy release in the "dry" state, depending on the physical state of water produced by different humidity and temperature combinations.

plant biology↗

Defining the Polycystin Pharmacophore Through HTS & Computational Biophysics.

Background and PurposePolycystins (PKD2, PKD2L1) are voltage-gated and Ca2+-modulated members of the transient receptor potential (TRP) family of ion channels. Loss of PKD2L1 expression results in seizure-susceptibility and autism-like features in mice, whereas variants in PKD2 cause autosomal dominant polycystic kidney disease. Despite decades of evidence clearly linking their dysfunction to human disease and demonstrating their physiological importance in the brain and kidneys, the polycystin pharmacophore remains undefined. Contributing to this knowledge gap is their resistance to drug screening campaigns, which are hindered by these channels unique subcellular trafficking to organelles such as the primary cilium. PKD2L1 is the only member of the polycystin family to form constitutively active ion channels on the plasma membrane when overexpressed. Experimental ApproachHEK293 cells stably expressing PKD2L1 F514A were pharmacologically screened via high-throughput electrophysiology to identify potent polycystin channel modulators. In-silico docking analysis and mutagenesis were used to define the receptor sites of screen hits. Inhibition by membrane-impermeable QX-314 was used to evaluate PKD2L1s binding site accessibility. Key ResultsScreen results identify potent PKD2L1 antagonists with divergent chemical core structures and highlight striking similarities between the molecular pharmacology of PKD2L1 and voltage-gated sodium channels. Docking analysis, channel mutagenesis, and physiological recordings identify an open-state accessible lateral fenestration receptor within the pore, and a mechanism of inhibition that stabilizes the PKD2L1 inactivated state. Conclusion and ImplicationOutcomes establish the suitability of our approach to expand our chemical knowledge of polycystins and delineates novel receptor moieties for the development of channel-specific antagonists in TRP channel research.

pharmacology and toxicology↗

Hybrid Generative Model: Bridging Machine Learning and Biophysics to Expand RNA Functional Diversity

Functional RNAs perform diverse catalytic roles, yet natural sequences represent only a narrow subset of what is possible. Rediscovering such activities requires exploring functional sequence diversity beyond natural RNAs. We introduce a hybrid generative model that combines a coevolutionary likelihood with an RNA secondary structure prior. This approach disentangles folding constraints from functional signals, enabling targeted diversification. On synthetic benchmarks, the model generates functional sequences beyond the training distribution. On large-scale ribozyme data, it improves the detection of active sequences and enhances sensitivity to local tertiary contacts. Finally, we introduce structural imprinting, a sampling strategy that uses alternative secondary structures to steer generation across under-sampled regions of sequence space. These results show that folding-informed generative modeling improves RNA design by supporting both extrapolation and control.

bioinformatics↗

Biophysical basis for brain folding and misfolding patterns in ferrets and humans

A mechanistic understanding of neurodevelopment requires us to follow the multiscale processes that connect molecular genetic processes to macroscopic cerebral cortical formations and thence to neurological function. Using magnetic resonance imaging of the brain of the ferret, a model organism for studying cortical morphogenesis, we create in vitro physical gel models and in silico numerical simulations of normal brain gyrification. Using observations of genetically manipulated animal models, we identify cerebral cortical thickness and cortical expansion rate as the primary drivers of dysmorphogenesis and demonstrate that in silico models allow us to examine the causes of aberrations in morphology and developmental processes at various stages of cortical ontogenesis. Finally, we explain analogous cortical malformations in human brains, with comparisons with human phenotypes induced by the same genetic defects, providing a unified perspective on brain morphogenesis that is driven proximally by genetic causes and affected mechanically via variations in the geometry of the brain and differential growth of the cortex. Impact statementPhysical gel models and numerical simulations are created to study ferret brain gyrification and cortical malformations, and comparisons with human phenotypes are presented to link genetics and brain morphogenesis.

neuroscience↗

Few-Shot Viral Variant Detection via Bayesian Active Learning and Biophysics

The early detection of high-fitness viral variants is critical for pandemic response, yet limited experimental resources at the onset of variant emergence hinder effective identification. To address this, we introduce an active learning framework that integrates protein language model ESM3, Gaussian process with uncertainty estimation, and a bio-physical model to predict the fitness of novel variants in a few-shot learning setting. By benchmarking on past SARS-CoV-2 data, we demonstrate that our methods accelerates the identification of high-fitness variants by up to fivefold compared to random sampling while requiring experimental characterization of fewer than 1% of possible variants. We also demonstrate that our framework benchmarked on deep mutational scans effectively identifies sites that are frequently mutated during natural viral evolution with a predictive advantage of up to two years compared to baseline strategies, particularly those enabling antibody escape while preserving ACE2 binding. Through systematic analysis of different acquisition strategies, we show that incorporating uncertainty in variant selection enables broader exploration of the sequence landscape, leading to the discovery of evolutionarily distant but potentially dangerous variants. Our results suggest that this framework could serve as an effective early warning system for identifying concerning SARS-CoV-2 variants and potentially emerging viruses with pandemic potential before they achieve widespread circulation.

bioinformatics↗

Fitness Landscape Ruggedness Arises from Biophysical Complexity

The ruggedness of a proteins fitness landscape constrains how it can evolve and how effectively it can be engineered. What determines whether a fitness landscape is rugged, and where in sequence space that ruggedness concentrates, remains poorly understood. Existing measures of ruggedness rely on complete combinatorial datasets and cannot be applied to landscapes in which only a fraction of possible sequences have been characterized. Here, we introduce a graph-based measure of ruggedness based on how quickly fitness differences diffuse across a landscape, which can be applied to sparse datasets. Applying this measure to 64 protein domains and several experimental systems with characterized folding and binding constraints, we show that ruggedness is not uniformly distributed but concentrates at regions where the dominant constraint on fitness changes. These results establish that the composition of individually simple constraints can generate localized ruggedness, providing a mechanistic basis for the complexity of protein fitness landscapes.

evolutionary biology↗

CHI3L1-Targeted Small Molecules as Glioblastoma Therapies: Virtual Screening-Based Discovery, Biophysical Validation, Pharmacokinetic Profiling, and Evaluation in Glioblastoma Spheroids

Glioblastoma (GBM) remains the most aggressive primary brain malignancy with a 10% three- year survival rate. Chitinase-3-like protein 1 (CHI3L1) has emerged as a critical factor in the progression of GBM progression, invasion, and treatment resistance. However, small molecule inhibitors targeting CHI3L1 are largely unexplored. Microscale thermophoresis (MST) investigation of the direct binding potential of reported CHI3L1 modulators (K284, G721-0282, CHI3L1-IN-1) revealed modest to undetectable direct CHI3L1 binding affinity. Herein, pharmacophore-based virtual screening of in-house library resulted in the discovery of G28 as the most potent small molecule CHI3L1 binder reported to date. The CHI3L1 binding affinity of G28 was validated using MST and surface plasmon resonance (SPR). To evaluate the GBM-modulatory potential of G28, we conducted comprehensive pharmacokinetic and 3D spheroid studies alongside established CHI3L1 modulators. G28 demonstrated outstanding bioavailability and low toxicity, addressing key limitations faced by previous CHI3L1-targeted strategies. Notably, in 3D GBM spheroid models, G28 significantly outperformed reported CHI3L1 small molecule modulators, showing the most pronounced dose-dependent reductions in spheroid weight, migration, and viability. These findings position G28 as the most promising CHI3L1-targeted small molecule to date and a compelling candidate for GBM therapeutic development.

pharmacology and toxicology↗

Biophysical modeling for accurate T cell specificity prediction of viral and tumor antigens

Accurate predictions of T cell receptor (TCR) specificity remain an important open problem in immunology, with broad implications for vaccine design, optimal immunotherapy, and improved management of autoimmune diseases. However, diversity in peptide antigens and TCR sequences at the level of individual patient repertoires remains a formidable computational challenge. Here, we develop a joint experimental and computational approach for predicting the antigen specificity of clinically-derived TCR sequences. Our model is trained on a combination of experimentally pre-identified and in silico-predicted TCR-pMHC structures using AlphaFold3. We apply our structural model in the clinical setting of hematopoietic stem cell transplant (HSCT) and demonstrate that our model is able to effectively discern the specificity of previously unseen donor and patient-derived TCR sequences against tumor associated and viral antigens. Model performance was further enhanced through the integration of sequence-based clustering and structurally diverse training templates. Our results highlight the predictive capabilities of structurally guided machine learning frameworks, trained on a minority test dataset, for antigen specificity prediction on unseen TCR sequences and their potential impact on a wide range of immunological applications.

immunology↗

Expression and Purification of Full-Length hnRNPA2B1 for Biophysical Characterization of Liquid-Liquid Phase Separation

Heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNPA2B1) is a multifunctional RNA-binding protein involved in RNA maturation and mRNA transport. It has recently been shown to undergo liquid-liquid phase separation (LLPS), contributing to the assembly of membraneless organelles. Moreover, dysregulation of LLPS is associated with the formation of pathogenic protein aggregates, in which hnRNPA2B1 is frequently found. Despite its biological and pathological relevance, studies on the full-length protein remain limited due to its intrinsically disordered, low-complexity domain, which renders hnRNPA2B1 highly aggregation-prone and difficult to purify. In this study, we report the successful expression and purification of full-length hnRNPA2B1 with high purity and minimal nucleic acid contamination. By optimizing buffer conditions, specifically ionic strength and pH, we maintain the protein in solution following cleavage of its solubility tag. Preliminary in vitro characterization under near-physiological conditions reveals that purified hnRNPA2B1 undergoes LLPS, forming dynamic liquid-like droplets that grow and mature into amorphous aggregates. Our approach provides a robust method for purifying hnRNPA2B1 suitable for LLPS and aggregation studies. This strategy may also be useful to purify other aggregation-prone, intrinsically disordered proteins.

biochemistry↗

CD33 and Clusterin Interact Biophysically and Genetically to Modulate Alzheimer Risk

We report the results of structural, functional and genetic studies on the CD33 sialic acid- binding receptor that reveal how non-coding variants in CD33 alter risk for Alzheimers disease (AD). The full-length CD33M isoform, whose expression is upregulated by non-coding AD-risk alleles, preferentially forms dimers at the cell surface, where they interact with AD-related proteins (clusterin and A{beta}). This interaction induces CD33M inhibitory signalling and downregulates protective microglial functions including phagocytic removal of amyloid plaques. Human brain expression quantitative trait loci (eQTL) and causal mediation analyses confirm that quantitative interactions between CLU and CD33 genotypes modulate AD phenotypes and suggest that genotypes at these loci might be used to personalise future therapeutic approaches. Our work also highlights several other unexpected aspects of CD33 biology, including a soluble shed extracellular fragment of CD33M and a similar soluble secreted product arising from a truncating mutation in the CD33 extracellular domain (CD33M{Delta}4bp).

neuroscience↗

On the role of L-type Ca2+ and BK channels in a biophysical model of cartwheel interneurons

Cartwheel interneurons (CWCs) in the auditory system exhibit a range of activity patterns relevant to auditory function and pathologies. Although experiments have shown how these patterns can vary across individual neurons and can change under pharmacological manipulations, the field has lacked a computational framework in which to explore the contributions of particular currents to these observations and to generate new predictions about the effects of manipulations on CWCs. In this work, we address this deficiency by presenting a conductance-based CWC computational model. This model captures the diversity of CWC activity patterns observed experimentally and suggests parameter changes that may underlie differences across cells. Bifurcation analysis of this model provides an explanation of how distinct dynamic mechanisms contribute to these differences, while direct simulations suggest how cells with different baseline dynamics will respond to variations in certain experimentally-accessible potassium and calcium channel conductances. In addition to the full model that we introduce, we present a reduced model that preserves CWC dynamic regimes. We classify the reduced model variables in terms of distinct dynamic timescales and show that the key transitions in dynamic patterns can be explained based on equilibria of the averaged dynamics of the slowest model variables, in a regime where the faster model variables exhibit oscillations. Overall, this study predicts how changes in parameters will influence CWC behavior, suggests how bifurcations contribute to changes in CWC dynamics, and provides a theoretical foundation that supports our simulation findings. Author summaryCartwheel interneurons (CWCs) are the most common class of inhibitory interneurons in an auditory brainstem region involved in sound localization and are believed to be important for auditory processing and pathologies. Distinct patterns of CWC activity have been observed experimentally in a variety of conditions. In this work, we present two novel computational models that simulate the factors contributing to CWC dynamics. By harnessing this framework, we are able to reveal the contributions of key ion currents to modulating CWC activity. Indeed, we find that the factors present in CWC neurons can produce a complicated dynamic landscape, with a wide range of output patterns possible as the relative strengths of these factors are varied. Overall, our models represent useful tools for understanding experimental results and generating new predictions about CWC behavior. In particular, in the more reduced of the two models, we can perform mathematical analysis to make more detailed predictions about the effects of current modulation on whether CWC neurons will exhibit regular spiking or more complex forms of outputs.

neuroscience↗

BIOPHYSICAL MECHANISMS UNDERLYING THE GENERATION AND MAINTENANCE OF RULE-LEARNING ENGRAM

Training rodents in a particularly difficult olfactory-discrimination task results with acquisition of high skill to perform the task superbly, termed rule-learning. We show that rule-learning occurs abruptly, in a "light bulb moment". Using whole-cell patch-clamp recordings in the piriform cortex of Fos2A-iCreER/TRAP2 mice, we targeted activated neurons, expressing immediate early genes (IEG). From the onset of training, IEG-positive neurons from trained animals display enhanced intrinsic excitability. Subsequently, synaptic excitation and inhibition is enhanced in these neurons, in a coordinated, cell-wide process. In parallel, the density of IEG-expressing neurons sharply declines. Double labeling with TRAP and c-Fos reveals that nearly two-thirds of the rule-memory engram neurons were engaged from the beginning of training. Silencing TRAP-expressing neurons using DREADDs leads to a complete loss of rule memory. We propose that rule learning occurs at a discrete moment, and is developed through a gradual process that stabilizes the memory of the rule.

neuroscience↗

Blood volume sensitive laminar fMRI with VASO in human hippocampus: Capabilities and biophysical challenges at clinical 7T scanners

Sub-millimeter resolution functional magnetic resonance imaging (fMRI) at ultra-high field ([≥] 7T) has offered an unprecedented opportunity to probe mesoscopic computations at a columnar or laminar level. However, its application has been primarily restricted to the neocortex. Inferior brain regions, particularly the hippocampus (HC), are challenging targets for laminar fMRI. Recent developments in acquisition methods have shown the feasibility of laminar recordings in the HC using gradient-echo blood oxygenation level-dependent (BOLD) contrast. Nonetheless, the spatial specificity of the BOLD signal is compromised by the draining veins bias. Cerebral blood volume (CBV)-sensitive sequences including vascular space occupancy (VASO) have emerged as a promising augmentation tool to capture the vein-free laminar activity. Yet, its feasibility in the HC is unclear and challenged by methodological constraints. Here, we optimized VASO to mitigate the macrovasculature contribution in HC. By evaluating a series of advanced acquisition strategies tailored to HC, we obtained improved VASO signal quality with minimal artifacts. The optimized protocol was further validated with an autobiographical memory task. Our findings show that combining the high detection power of gradient-echo BOLD with the vein-free VASO contrast allows for differentiation between neural activity-related BOLD signals and those biased by draining veins. These results demonstrate the feasibility of submillimeter VASO acquired with conventional 7T scanners in the HC to map the circuit-level mechanisms of memory retrieval across HC subfields, laying a foundation to investigate the microcircuitry of HC-driven complex cognitive functions and their alterations in neurodegeneration.

neuroscience↗

Biophysical mechanisms of morphogenesis in lizard lungs

The lungs of squamate reptiles (lizards and snakes) are highly diverse, exhibiting single chambers, multiple chambers, transitional forms with two to three chambers, along with a suite of other anatomical features, including finger-like epithelial projections into the body cavity known as diverticulae. During embryonic development of the simple, sac-like lungs of anoles, the epithelium is pushed through the openings of a pulmonary smooth muscle mesh by the forces of luminal fluid pressure. This process of stress ball morphogenesis generates the faveolar epithelium typical of squamate lungs. Here, we compared embryonic lung development in brown anoles, leopard geckos, and veiled chameleons to determine if stress ball morphogenesis is conserved across squamates and to understand the physical processes that generate transitional-chambered lungs with diverticulae. We found that epithelial protrusion through the holes in a pulmonary smooth muscle mesh is conserved across squamates. Surprisingly, however, we found that luminal inflation is not conserved. Instead, leopard geckos and veiled chameleons appear to generate their faveolae via epithelial folding downstream of epithelial proliferation. We also found experimental and computational evidence suggesting that the transitional chambers and diverticulae of veiled chameleon lungs develop via apical constriction, a process known to be crucial for airway branching in the bird lung. Thus, distinct morphogenetic mechanisms generate epithelial diversity in squamate lungs, which may underpin their species-specific physiological and ecological adaptations.

developmental biology↗

Adipose-mimetic granular hydrogels uncover biophysical cues driving breast cancer invasion

1.Breast cancer cells invade mammary adipose tissue during initial stages of metastasis but how the physical properties of adipose tissue regulate this process remains unclear. Here, we combined single cell mechanical characterization of primary adipocytes with microfluidic hydrogel fabrication, quantitative multiparametric imaging, Discrete Element Method (DEM) simulations, and in vivo experiments to elucidate these connections. First, we quantified the heterogeneous size and stiffness of primary adipocytes, and replicated these properties by fabricating adipocyte-sized polyacrylamide (PAAm) beads with tunable elasticity. Subsequently, we embedded these beads into type I collagen, the primary fibrillar extracellular matrix (ECM) component of breast adipose tissue, to form 3D granular hydrogels mimicking aspects of native adipose tissue architecture. Granular hydrogels embedded with beads demonstrated increased breast cancer cell invasion relative to bead-free controls, an effect that was more pronounced with soft versus stiff beads and correlated with increased collagen fiber alignment and hierarchical organization. In addition, live cell imaging and DEM simulations revealed that soft beads promoted invasion relative to stiff beads by deforming in response to confined cancer cell migration. Fiber alignment and adipocyte deformation trends were validated in vivo via intravital imaging of cancer cell migration in mammary fat pads of mice, and suggest that adipocyte mechanics regulate breast cancer invasion by coordinating both ECM architecture and cellular confinement. Ultimately, this work highlights the utility of tunable PAAm bead-collagen composites as micromechanical models to study the effect of adipose tissue structure on cancer cell invasion. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=106 SRC="FIGDIR/small/684224v1_ufig1.gif" ALT="Figure 1"> View larger version (56K): org.highwire.dtl.DTLVardef@d3ce0org.highwire.dtl.DTLVardef@117abb1org.highwire.dtl.DTLVardef@10917faorg.highwire.dtl.DTLVardef@55f3e_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical Abstract:C_FLOATNO Polyacrylamide bead granular hydrogels recapitulate adipose tissue structure and alter cancer cell invasion in a stiffness-dependent manner. The invasion of primary breast cancer cells through mammary adipose is influenced by the heterogeneous mechanical properties of native stromal components, namely adipocytes and extracellular matrix (left). Customizable granular hydrogels provide insight into the physical regulation of breast cancer progression by mimicking adipose tissue extracellular matrix structure (collagen I, i.) and stromal cell mechanics (polyacrylamide bead, ii.), whereby the inclusion of soft beads and/or dilute matrix promotes cancer cell migration compared to stiffer hydrogels (right). C_FIG

bioengineering↗

Biophysics of object segmentation in a collision-detecting neuron

Collision avoidance is critical for survival, including in humans, and many species possess visual neurons exquisitely sensitive to objects approaching on a collision course. The most studied such collision-detecting neuron within the optic lobe of grasshoppers has long served as a model for understanding collision avoidance behaviors and their underlying neural computations. Here, we demonstrate that this neuron detects the spatial coherence of a simulated impending object, thereby carrying out a computation akin to object segmentation critical for proper escape behavior. At the cellular level, object segmentation relies on a precise selection of the spatiotemporal pattern of synaptic inputs by dendritic membrane potential-activated channels. One channel type linked to dendritic computations in many neural systems, the hyperpolarization-activated cation channel, HCN, plays a central role in this computation as its pharmacological block abolishes the neuron's spatial selectivity and impairs the generation of visually guided escape behaviors, making it directly relevant to survival. Our results elucidate how active dendritic channels produce neuronal and behavioral object specificity by discriminating between complex spatiotemporal synaptic activation patterns.

neuroscience↗

The expa1-1 mutant reveals a new biophysical lateral root organogenesis checkpoint

In plants, post-embryonic formation of new organs helps shape the adult organism. This requires the tight regulation of when and where a new organ is formed, and a coordination of the underlying cell divisions. To build a root system, new lateral roots are continuously developing, and this process requires asymmetric cell division in adjacent pericycle cells. Characterization of an expansin a1 (expa1) mutant has revealed a novel checkpoint during lateral root formation. Specifically, a minimal pericycle width was found to be necessary and sufficient to trigger asymmetric pericycle cell divisions during auxin-driven lateral root formation. We conclude that a localized radial expansion of adjacent pericycle cells is required to position the asymmetric cell divisions and generate a core of small daughter cells, which is a prerequisite for lateral root organogenesis.\n\nSIGNFICANCE STATEMENTOrgan formation is an essential process in plants and animals, driven by cell division and cell identity establishment. Root branching, where lateral roots form along the primary root axis, increases the root system and aids capture of water and nutrients. We have discovered that tight control of cell width is necessary to co-ordinate asymmetric cell divisions in cells that give rise to a new lateral root organ. While biomechanical processes have been shown to play a role in plant organogenesis, including lateral root formation, our data give new mechanistic insights into the cell size checkpoint during lateral root initiation.

plant biology↗

Determination of biophysical parameters for enhancement of human osteoblast proliferation by mechanical vibration in the infrasonic frequency range

Experimental methods for studying an enhancement of osteoblast proliferation in vitro provide tools for the research of biochemical processes involved in bone turnover in vivo. Some of the current methods used for this purpose are based on the ability of the osteoblasts to enhance proliferation by mechanical stimulation. We describe an experimental approach of biomechanical stimulation of cultured human osteoblast-like cells by vibration. This method is based on the specially designed controlled vibration setup that consists of an electric actuator, with horizontally mounted well plate containing cell cultures. Previously this method found to be effective to enhance cell proliferation, but the exact mechanical parameters of effective vibration were elusive. The current low friction system for mechanical stimulation of osteoblast-like cells in vitro provides recording of narrow range mechanical parameters in the infrasonic spectrum.\n\nWe exposed human osteoblast-like cells in explant monolayer culture to mechanical vibration in the 10-70Hz range of frequencies and found that 50-70 Hz of vibration frequency is optimal for inducing osteoblast proliferation that was deduced from interrelation between unchanged cell number in culture samples with significant decrease in cell death rate (decreased LDH activity in culture media, p<.05) and with parallel decrease of their maturation level (p<.01).\n\nIn this report we determined the optimal mechanical parameters and excitation protocol for induction of osteoblast proliferation in vitro by using a tunable and versatile mechanical platform, which can be used in the research of cell mechanotransduction.

cell biology↗