Biology subjects

Agarwal, A.

Publications and source records attributed to Agarwal, A..

At least 19 recordsLinked to original sources

bioRxiv2026

A stromal metabolic program suppresses NK-cell immunity to drive tumor progression in HER2-low breast cancer

Cancer-associated fibroblasts (CAFs) are major regulators of the tumor microenvironment, yet how distinct CAF states suppress innate immunity in HER2-low breast cancer remains poorly understood. Here, we identify an S100A4-enriched CAF population that expands during HER2-low breast tumor progression and establishes a metabolically immunosuppressive niche. Spatial transcriptomics and multiplex imaging of human HER2-low tumors reveal progressive CAF accumulation and an inverse spatial association between S100A4-enriched CAFs and immune infiltration, including natural killer (NK) cells. Using an immunocompetent HER2-low mammary tumor model, we show that S100A4-enriched CAFs promote tumor initiation and progression while suppressing NK-cell cytotoxicity, IFN-{gamma} production, perforin, and granzyme B. Fractionation of CAF-conditioned media and metabolic profiling identify a low-molecular-weight immunosuppressive program characterized by enhanced branched-chain amino acid catabolism and accumulation of branched-chain -keto acids (BCKAs). Mechanistically, BCKAs directly suppress NK-cell IFN-{gamma} production, whereas inhibition of the branched-chain aminotransferase BCAT1 reduces CAF-mediated NK-cell suppression and restores antitumor cytotoxicity. BCAT1 inhibition also suppresses HER2-low tumor growth in vivo, an effect attenuated by NK-cell depletion, establishing NK-cell restoration as a functional component of its antitumor activity. Together, these findings uncover a CAF-driven metabolic immune checkpoint in which S100A4-enriched CAFs exploit BCAT1-dependent BCKA production to suppress NK-cell surveillance and promote HER2-low breast tumor progression. Targeting stromal BCAT1 therefore represents a potential strategy to dismantle CAF-mediated immune suppression and restore innate antitumor immunity.

cancer biology↗

bioRxiv2026

Targeting Macrophage Connexin 43 Hemichannels Suppresses Synovial Inflammation and Osteoarthritis

Osteoarthritis (OA) is the most common chronic joint disease and a leading cause of disability worldwide, yet no disease-modifying therapies are available. Although OA has traditionally been viewed as a cartilage-centered degenerative disease, growing evidence suggests that synovial inflammation is a critical driver of disease progression. However, the upstream mechanisms and actionable targets that govern this inflammatory response remain poorly defined. Connexin 43 (Cx43), a key regulator of musculoskeletal homeostasis that forms both gap junctions and hemichannels (HCs), is upregulated in osteoarthritic cartilage and synovium, but the specific contribution of Cx43 channels remains unclear. Here, we show that selective inhibition of Cx43 HCs attenuates OA progression in a murine post-traumatic OA model. Using adeno-associated virus (AAV)-mediated gene deletion, we found that Cx43 knockdown in joint tissues reduced cartilage erosion, synovial hyperplasia, subchondral bone sclerosis, and OA-associated pain. To dissect HC-specific functions, we developed a monoclonal antibody, Cx43(M1), that blocks Cx43 HCs without affecting junctional communication. Cx43(M1) preserved cartilage integrity, suppressed synovial inflammation, and alleviated pain behaviors, even when treatment was initiated after established cartilage damage. Mechanistically, the antibody preferentially localized to synovial macrophages, where it inhibited HC activity, reduced ATP and PGE2 release, and suppressed pro-inflammatory gene expression. Macrophage-specific Cx43 deletion recapitulated these protective effects, and Cx43 HC blockade in primary human OA synovial macrophages similarly inhibited inflammatory and catabolic responses. These findings identify synovial macrophage Cx43 HCs as upstream drivers of OA and support a paradigm shift from a predominantly chondrocyte-centered view toward synovial macrophage directed intervention.

immunology↗

bioRxiv2026

Derivation of primary fetal epithelial organoids from cryopreserved human amniotic fluid cells.

Primary fetal human epithelial organoids are canonically derived from tissue samples obtained after termination of pregnancy. Recently, we have demonstrated that these organoids can be consistently generated from amniotic fluid cells isolated prenatally during diagnostic and interventional procedures. Amniotic Fluid-derived Organoids (AFOs) are promising fetal epithelial lung, kidney and small intestine tissue models that bypass some of the ethical and legal constraints associated with obtaining primary fetal tissue. Despite this, the widespread adoption of this technology is limited by the lack of standardised and accessible pipelines for the biobanking, culturing and distribution of organoid-forming amniotic fluid cells. To enable broader AFO use for research and clinical purposes, this work investigates two cryopreservation strategies to optimise AF cell recovery for organoid derivation. In Strategy 1, viable AF cells were sorted before freezing, while in Strategy 2 unsorted AF was frozen, and viable AF cells were sorted for viability after thawing. We present here a comprehensive evaluation of 5 commercially available GMP-compliant freezing media (FM1-5) alongside a standard lab-grade control (FM CT). AFOs were assessed for formation efficiency, morphological characteristics, proliferation capacity, epithelial identity, and tissue type. Our results demonstrate that organoid-forming AF cells can be successfully cryopreserved both pre- and post-sorting. Strategy 1 yielded higher organoid formation efficiency, with AFOs derived from cryopreserved and fresh cells exhibiting comparable expansion potential. Finally, FM4 provided minimal to no decline in survival rate, making it the most effective GMP-grade freezing medium tested. In conclusion, we present two viable cryopreservation strategies adaptable to different laboratory settings and identify optimal GMP-compliant freezing media, to support safe distribution and centralised processing, thereby facilitating scalability and collaborative work on the AFO technology.

cell biology↗

bioRxiv2026

Multiple Fault Analysis and Drug Therapy on Signaling Pathways Using Dynamic Bayesian Network-based Model

Cancer-associated signaling pathways often exhibit abnormal activation under simultaneous dysregulation of multiple molecular components. This study presents a probabilistic temporal Dynamic Bayesian Network (DBN)-based framework for analyzing multi-fault behaviour and intervention response in Growth Factor (GF) and Mitogen-Activated Protein Kinase (MAPK) signaling pathways. Unlike deterministic Boolean propagation, the proposed model represents each pathway component through an activation probability and propagates these probabilities over discrete time steps using soft-logic update rules. One-, two-, three-, and four-fault scenarios were systematically evaluated under a common lowest-burden input vector. The resulting output probabilities were summarized using an encoded pathway-burden score, and known-drug combinations were ranked using efficiency scores relative to no-intervention baselines. Pareto analysis was further used to balance intervention efficiency against drug-vector burden, while a custom dual-target search was performed to identify computational intervention hypotheses beyond predefined drug targets. Results showed that encoded burden increased with fault order in both pathways, with MAPK producing a higher baseline burden than GF. Among known-drug vectors, U0126+LY294002+Temsirolimus consistently emerged as the strongest low-burden candidate, achieving efficiency close to the maximum six-drug vector. Custom dual-target analysis identified ERK1/2+RPS6KB1 in GF and Raf+MEK1 in MAPK as high-impact computational target pairs. Runtime benchmarking showed that batched vectorized NumPy execution substantially improved scalability for higher-order fault simulations. Overall, the framework provides an interpretable and scalable approach for probabilistic pathway-level fault analysis and intervention prioritization.

bioinformatics↗

bioRxiv2026

A systematic imputation framework for sparse, multimodal space biology datasets: application to retinal imaging and omics from the RR9 mission

Missing data is a fundamental challenge in space biology, where high experimental costs, limited sample availability, and tissue allocation constraints produce datasets that are sparse, multimodal, and heterogeneous. We present a systematic four-stage framework for diagnosing, implementing, and validating data imputation strategies tailored to these characteristics, and demonstrate its application to retinal imaging and omics data from the NASA Rodent Research 9 (RR9) mission. Using logistic regression-based missingness diagnosis, we identify a Missing At Random (MAR) mechanism driven by experimental design constraints across nine assay modalities. We implement and optimize three imputation strategies: K-Nearest Neighbors (KNN), Multiple Imputation by Chained Equations with weak ElasticNet regularization (MICE-Elastic), and a per-column hybrid strategy, evaluated against a random sample imputer baseline. Validation across seven complementary metrics including supervised classification, unsupervised clustering, correlation structure preservation, masked value recovery, cross-dataset generalization, and permutation testing reveals that MICE-Elastic and the Hybrid strategy preserve genuine biological signal in both RNA-seq and TUNEL modalities, while KNN and the random sample imputer do not despite achieving comparable cross-validation accuracy. A critical finding is that imputation substantially improves supervised classification performance while consistently degrading unsupervised clustering structure, a trade-off researchers must understand before applying these methods. This framework provides practical, actionable guidance for space biologists and data scientists managing sparse multimodal datasets, and represents a foundational step toward digital twin development for space medicine.

bioinformatics↗

bioRxiv2026

Glucose selectively drives a rapid oxidative burst and immunometabolic reprogramming in human neutrophils during Mycobacterium tuberculosis infection

Neutrophil functions have been linked to tuberculosis (TB)-associated tissue damage; however, the mechanisms driving immunopathology in the human TB lung remain poorly understood, due partly to the scarcity of human tissue for study. Here, we examine the metabolic and bioenergetic reprogramming of human neutrophils in response to Mycobacterium tuberculosis (Mtb) infection. In human necrotic TB granulomas, levels of NETosis-associated proteins are increased and co-localize with GLUT3, linking nutrient uptake to tissue damage. In vitro, Mtb elicits an immediate, contact-dependent oxidative burst in human neutrophils, and the magnitude of this response is carbon source-dependent. Glucose enables the most robust responses, indicating that glucose metabolism is a key driver of neutrophil-mediated inflammatory damage during TB. Mtb-induced responses are distinct from those induced by PMA, non-tuberculous mycobacteria, or other pathogenic intracellular bacteria, and are mediated through multiple neutrophil surface receptors. Notably, our data show that while the oxidative burst is carbon source-dependent, cytokine production is not. Further, Mtb infection reprograms neutrophil metabolism from glycolysis to the pentose phosphate pathway (PPP), generating NADPH required for the oxidative burst. Inhibiting G6PD, NADPH oxidase, or PAD4 significantly reduces this response, highlighting the PPP as a promising host target for mitigating TB immunopathology.

microbiology↗

bioRxiv2026

C-Terminal Domain of Mycobacterium tuberculosis Glutamate Decarboxylase determines the bacterial stress-adaptive metabolic state that steers macrophage polarisation supporting intracellular persistence

Mycobacterium tuberculosis (Mtb) adapts metabolically to survive hostile conditions within alveolar macrophages, but whether specific enzyme-driven metabolic states of Mtb link stress adaptation to host immunomodulation remains unclear. Using LC-MRM/MS metabolite profiling and metabolic modeling, we previously identified elevated {gamma}-aminobutyric acid (GABA) production via the GABA shunt during adaptation to oxidative and acidic stress. Here, we show that stress-responsive enzymatic efficiency of glutamate decarboxylase (GadB) in producing GABA, not only mitigates acidic and oxidative stress but also drives macrophage polarisation towards the M2 phenotype, extending its role beyond bioenergetics into host-pathogen cross-talk. We identify the C-terminal domain (CTD) of mycobacterial GadB as a key regulator. Enzyme kinetics and fluorescence spectroscopy revealed that CTD deletion abolishes pH-dependent regulation of catalytic efficiency (kcat/Km) and substrate affinity (Ka). Under oxidative stress, the CTD enhances catalytic efficiency without altering substrate affinity, suggesting condition-specific roles. Accordingly, in vitro, Mtb::MtbGadB with higher GABA and [~]40% lower ROS outgrew Mtb::MtbGadB{Delta}Ct under acidic and oxidative conditions. These biochemical differences influenced host immunity, wherein infection with high-GABA-producing Mtb::MtbGadB strain induced M2 polarisation, with decreased CD86, reduced RNS, limited endosomal maturation, as indicated by Rab5/Rab7 ratio and Lysotracker-based confocal microscopy, and decreased TNF-, IL-12p70, and IFN-{gamma} release compared to low-GABA-producing Mtb::MtbGadB{Delta}Ct. Consequently, Mtb::MtbGadB{Delta}Ct showed higher clearance. We conclude that by enabling pH-dependent activity of GadB, the CTD orchestrates proton consumption, ROS reduction, and GABA-mediated immunomodulation. Distinct structural features of CTDs in human versus Mtb GADs highlight the CTD as a selective drug target for pathogen-specific, host-compatible therapies. ImportanceThis work establishes a direct mechanistic link between metabolic state of Mycobacterium tuberculosis (Mtb) actively shaping host immune responses, deciding infection outcome. The study identifies the C-terminal domain (CTD) of MtbGadB in steering enzymatic efficiency, regulating intrabacterial GABA production and ROS quenching, determining the stress-responsive metabolic state of the bacteria. Infection with metabolically distinct strains had differential impacts on host immunity, wherein infection with high-GABA-producing Mtb::MtbGadB induced M2-polarisation, reduced RNS, limited endosomal maturation, and reduced proinflammatory cytokine release, compared to low-GABA-producing Mtb::MtbGadB{Delta}Ct. Consequently, Mtb::MtbGadB{Delta}Ct could be cleared, but Mtb::MtbGadB persisted, deciding the fate of the infection. Structural differences between CTDs of human and Mtb GADs, make it a therapeutically-exploitable determinant for developing pathogen-specific drugs that remain compatible with the host.

microbiology↗

bioRxiv2026

Robot-based 3D-multispectral monitoring of soybean in a spatially heterogenous agrivoltaic environment

Agrophotovoltaic (APV) systems provide a unique opportunity for improving agricultural land-use efficiency by combining solar energy capture via photovoltaic panels with crop production. However, in-depth information on plant growth patterns within the spatially heterogenous microclimate created by the intermittent shading of APVs is largely missing. In the present study, we implement a customized robot-mounted 3D-multispectral imaging system to closely monitor the growth and spectral reflectance patterns of a conventional soybean cultivar "Eiko" (EK) and a chlorophyll-deficient mutant variety MinnGold (MG) under an APV system. Weekly trends in canopy morphometric features revealed significant variations in plant height, 3D leaf area, light penetration, and canopy volume across the APV field depending on the proximity with the overhead solar panels for both EK and MG, with plants receiving adequate rainfall and intermittent shade performing the best. Furthermore, although spectral indices exhibited variations between EK and MG due to intrinsic differences in pigmentation, symptoms of stress could be detected for both genotypes within rain-shaded areas of the APV plot. Hence, the present investigation depicts the potential for complementary usage of robotics and machine vision for high-precision high-throughput crop monitoring under APVs, which would enable better crop management within such non-homogenous cultivation systems.

plant biology↗

bioRxiv2026

METTL16 promotes taxane resistance in Triple-Negative Breast Cancer through m6A-dependent translational upregulation of ABCB1

Triple-negative breast cancer (TNBC) commonly develops resistance to taxane-based chemotherapy, resulting in recurrence and poor clinical outcomes. Defining the molecular mechanisms that sustain chemoresistance is essential for improving therapeutic efficacy. Using unbiased insertional mutagenesis, we identified the RNA methyltransferase METTL16 as a previously unrecognized epi-transcriptomic driver of taxane resistance. METTL16 overexpression conferred resistance to docetaxel and paclitaxel across multiple TNBC models, and METTL16 expression was elevated in paclitaxel-resistant cells. Genetic depletion of METTL16 in paclitaxel-resistant cells restored taxane sensitivity. Because enhanced drug efflux is a well-established mechanism of taxane resistance, we investigated whether METTL16 regulates the multidrug transporter ABCB1 (P-glycoprotein). Paclitaxel-resistant TNBC cells exhibited elevated METTL16 and ABCB1 expression compared to parental cells. METTL16 binds to ABCB1 mRNA and catalyzes its N6-methyladenosine (m6A) modification, promoting increased ribosome loading and translational upregulation without altering transcript abundance. Inactivation of METTL16 impaired ABCB1 polysome association and restored paclitaxel sensitivity, demonstrating that the methyltransferase activity is essential for resistance. Consistent with this mechanism, METTL16 overexpression increased ABCB1 protein levels, whereas METTL16 down-regulation increased intracellular paclitaxel accumulation. Analysis of TNBC patient datasets revealed a positive correlation between METTL16 and ABCB1 expression, supporting the clinical relevance of this mechanism. Antisense-mediated inhibition of METTL16 using a translation-blocking Vivo-Morpholino reduced the survival of resistant TNBC cells and suppressed tumor growth in vivo. Surprisingly, genetic ablation of METTL16 caused profound loss of TNBC cell viability, while having only modest effects on nonmalignant mammary epithelial cells, indicating a cancer-selective dependency. Collectively, these findings define a METTL16-ABCB1 interaction that drives taxane resistance and establish METTL16 as a therapeutic target in TNBC.

cancer biology↗

bioRxiv2026

Multisensory coding of audiovisual movies in the human hippocampus

The hippocampus receives convergent input from multiple sensory systems, yet in humans it has been studied almost exclusively through vision. Here we examine how the hippocampus contributes to sensory processing beyond the visual modality and to the multisensory integration of visual information with these other modalities. Participants were exposed to multiple repetitions of short naturalistic movie clips, each presented in four formats: auditory-only, visual-only, congruent audiovisual, and incongruent audiovisual (audio and video from different movies). Using high-resolution fMRI, we measured univariate activation and multivariate representations across the longitudinal axis and subfields of the human hippocampus. Whereas univariate analyses detected only visual responses across hippocampal subregions, with no activation for auditory stimuli and no benefit of congruent stimuli, multivariate analyses revealed robust representations of both auditory and visual scenes. The posterior hippocampus showed enhanced pattern similarity for congruent stimuli relative to unisensory stimuli, especially in the left hemisphere, demonstrating multisensory facilitation. The anterior hippocampus showed crossmodal decoding between auditory and visual versions of the same clip, driven by the right hemisphere, suggesting a more abstract representation. Finally, whole-brain searchlight analyses revealed parallel effects in cortical regions known to support multisensory integration. These findings advance understanding of auditory and multisensory coding in the human hippocampus, revealing specialization and lateralization along its longitudinal axis.

neuroscience↗

bioRxiv2025

β-catenin-independent regulation by TCF7L2 underlies isoform redundancy during embryonic thalamic development

Alternative promoter usage generates multiple transcription factor isoforms during brain development, yet their functional significance remains poorly defined. One such example is TCF7L2, a transcription factor critical for the development of the thalamus and recurrently affected by de novo mutations in autism spectrum disorder. TCF7L2 exists in two isoforms driven from different promoters: the long isoform (L-TCF7L2) with the {beta}-catenin-binding domain, and the shorter isoform (S-TCF7L2), lacking this domain and classically considered a dominant-negative regulator of the L isoform. We investigated the role of TCF7L2 isoforms in thalamic development using total and isoform-specific knockout strategies. Integrated phenotypic and transcriptomic analyses revealed functional redundancy of TCF7L2 isoforms during embryogenesis. {beta}-catenin subcellular localization and chromatin occupancy uncovered a developmental switch in TCF7L2 activity, from a {beta}-catenin-independent and isoform-redundant mode in the embryonic thalamus to a {beta}-catenin-dependent program postnatally. More broadly, these findings point to distinct embryonic and postnatal regulatory strategies, with alternative promoter usage potentially supporting robust availability of regulatory proteins during brain development.

neuroscience↗

bioRxiv2025

The MTL200: a surface-based, probabilistic atlas of the medial temporal lobe

The human medial temporal lobe (MTL) includes a number of cortical regions surrounding the hippocampus implicated in long-term memory and high-level perception, including temporopolar cortex (TPC), entorhinal cortex (ERC), perirhinal cortex (PRC), and parahippocampal cortex (PHC). A critical prerequisite to studying the function of these areas is to define their anatomical boundaries. While several atlases or segmentations methods for automated boundary definition exist, they have limitations: 1) they are derived from small sample sizes; 2) most do not include TPC; and 3) they are volumetric rather than surface-based and are therefore not straightforward to use for surface-based analysis of MRI data. Here we present the MTL200: a surface-based, probabilistic atlas of MTL regions TPC, ERC, PRC, and PHC. The atlas is derived from hand drawings of regions based on gross anatomical features corresponding to cytoarchitectonic boundaries, using anatomical MRI data from 200 human participants. This work provides a valuable tool for researchers studying the anatomy and function of MTL regions, particularly the poorly characterized area TPC.

neuroscience↗

bioRxiv2025

Polysialic Acid-Functionalized MAP Scaffolds Promote Regulatory Immune Responses After Ischemic Stroke

Glycosylation regulates immune and neural functions within the central nervous system (CNS), yet biomaterials rarely leverage glycans due to their structural complexity. Polysialic acid (PSA), comprising 2,8-linked sialic acid residues, is a promising candidate owing to its potent immunomodulatory interactions with inhibitory Siglec receptors. Systematic screening of multiple sialic acid derivatives identifies PSA as uniquely effective in inducing anti-inflammatory polarization of bone marrow-derived macrophages (BMDMs). Based on these findings, an injectable microporous annealed particle (MAP) scaffold presenting PSA covalently via its reducing end (MAP-PSA) is engineered, recapitulating physiological glycan orientation. MAP-PSA exhibits robust mechanical properties, stable glycan immobilization, and resistance to enzymatic degradation. Using ischemic stroke as a CNS injury model, MAP-PSA significantly reduces neutrophil infiltration and inflammatory activation while enhancing reparative macrophage and microglial phenotypes. These immunomodulatory effects persist into subacute stages, characterized by sustained reductions in inflammation and enhanced microglial homeostasis. Overall, MAP-PSA scaffolds demonstrate a novel therapeutic paradigm for CNS injuries such as stroke, with translational potential for broader neuroinflammatory and regenerative applications.

bioengineering↗

bioRxiv2025

Cell-type-selective synaptogenesis during the development of excitatory connectivity in the mammalian neocortex

The function of mammalian neocortex relies on the timing of axon extension and establishment of cell-type-biased patterns of excitatory synaptic connections. A subtype of excitatory neurons, layer 6 corticothalamic neurons (L6CThNs), ultimately exhibit a marked preference for synapsing onto parvalbumin-positive (PV) inhibitory interneurons over more common excitatory cells in layers 6 and 4 (L6, L4). We show that the intracortical axons of L6CThNs develop in phases, elongating within L6, then pausing before extending translaminar branches into L4. Decreasing L6CThN excitability selectively enhanced axon growth in L6 but not later elaboration in L4. For both layers, we tested whether preferential synaptogenesis onto rarer PV interneurons, or promiscuous synapse formation followed by selective pruning, generated adult connectivity. We found that L6CThNs formed functional AMPA-receptor-containing synapses preferentially onto PV interneurons. Silent L6CThN synapses were not detected. Our findings show that cell-type-biased synaptogenesis underlies the formation of functional cell-type-specific excitatory connections in the neocortex.

neuroscience↗

bioRxiv2025

Closing the loop: Teaching single-cell foundation models to learn from perturbations

The application of transfer learning models to large scale single-cell datasets has enabled the development of single-cell foundation models (scFMs) that can predict cellular responses to perturbations in silico. Although these predictions can be experimentally tested, current scFMs are unable to "close the loop" and learn from these experiments to create better predictions. Here, we introduce a "closed-loop" framework that extends the scFM by incorporating perturbation data during model fine-tuning. Our closed-loop model improves prediction accuracy, increasing positive predictive value in the setting of T-cell activation three-fold. We applied this model to RUNX1-familial platelet disorder, a rare pediatric blood disorder and identified two therapeutic targets (mTOR and CD74-MIF signaling axis) and two novel pathways (protein kinase C and phosphoinositide 3-kinase). This work establishes that iterative incorporation of experimental data to foundation models enhances biological predictions, representing a crucial step toward realizing the promise of "virtual cell" models for biomedical discovery.

bioinformatics↗

bioRxiv2025

Dual effect of alpha-synuclein disease variants on condensate formation

-synuclein is a pre-synaptic protein1,2 implicated in synucleinopathies like Parkinsons disease and Dementia with Lewy Bodies, where it accumulates in intracellular aggregates termed Lewy bodies and Lewy neurites3. Recent studies have reported that -synuclein undergoes phase separation to form biomolecular condensates both in vitro and in mammalian cells4-7. -synuclein condensates are thought to contribute to disease through progressive aggregation5,8,9. Here we show that specific PD-associated -synuclein variants fail to form biomolecular condensates. We demonstrate that only two -synuclein variants associated with familial disease, E46K and E83Q, enhance condensate formation in vitro and in cells. However, variants including A30G, G51D, and A53E fail to form or have reduced levels of condensate formation in cells. The same phenotypes are reflected in the budding yeast model showing differential inclusion formation. In iPSC-derived neurons, the propensity of -synuclein variants to undergo VAMP2-mediated phase separation reflects the level of synaptic enrichment. We show that the intrinsic propensity of -synuclein to form condensates and the ability to bind lipid membranes are important to mediate condensate formation in cells. Our results emphasize that -synuclein pathology follows divergent pathways, with both increased and decreased condensate formation contributing to disease. This study establishes biomolecular condensates as a key intermediate in -synuclein dysfunction, providing a novel foundation for translational research.

neuroscience↗

bioRxiv2025

Synergistic 3D, multispectral, and thermal image analysis via supervised machine learning for improved detection of root rot symptoms in hydroponically-grown flat-leaf parsley

Root rot in hydroponically-grown leafy vegetables is difficult to detect via conventional manual and machine vision-based approaches as symptoms of infection are not clearly visible on the canopy at earlier stages of infection. Hence, the present study investigates the potential of using machine learning for assessing canopy information obtained from multiple imaging platforms synergistically to improve root rot detection. Herein, flat-leaf parsley seedlings were grown in an experimental hydroponic vertical farm and inoculated with Pythium irregulare and Phytophthora nicotianae. Subsequently, the seedlings were imaged via 3D, multispectral, and thermal sensors at various stages of growth to obtain twenty-six image-based plant features. Following a preliminary screening of redundant features via regression analysis, data for seventeen image features associated with morphometric, spectral, and thermal attributes was co-analyzed using supervised machine learning by Support Vector Machines (SVM). Exhaustive feature selection using different SVM kernels and maximum feature thresholds was performed to identify optimal feature subsets. It was observed that combining parameters obtained from all three imaging platforms enabled better identification of infected samples (>99%) than using a higher number of attributes from individual imaging systems. In addition, model performance was improved considerably by including temporal information during model training. Hence, it may be inferred that fusion of data from multiple imaging systems and using it with temporal information can enable better real-time high-throughput monitoring of root rot.

plant biology↗

bioRxiv2025

A cholinergic spinal pathway for the adaptive control of breathing

The ability to amplify motor neuron (MN) output is essential for generating high intensity motor actions. This is critical for breathing that must be rapidly adjusted to accommodate changing metabolic demands. While brainstem circuits generate the breathing rhythm, the pathways that directly augment respiratory MN output are not well understood. Here, we mapped first-order inputs to phrenic motor neurons (PMNs), a key respiratory MN population that initiates diaphragm contraction to drive breathing. We identified a predominant spinal input from a distinct subset of genetically-defined V0C cholinergic interneurons. We found that these interneurons receive phasic excitation from brainstem respiratory centers, augment phrenic output through M2 muscarinic receptors, and are highly activated under a hypercapnia challenge. Specifically silencing cholinergic interneuron neurotransmission impairs the breathing response to hypercapnia. Collectively, our findings identify a novel spinal pathway that amplifies breathing, presenting a potential target for promoting recovery of breathing following spinal cord injury.

neuroscience↗