Biology subjects

Hong, B.

Publications and source records attributed to Hong, B..

At least 19 recordsLinked to original sources

bioRxiv2026

BCIJelly: An integrated ecosystem for brain-computer interface research

Brain-computer interface (BCI) research relies on multistage computational pipelines, but progress has been slowed by fragmented data formats, heterogeneous decoder implementations and hardware-specific deployment toolchains. Here, we introduce BCIJelly, a unified ecosystem that standardizes 18 BCI datasets into AI-ready inputs and integrates 15 benchmark decoders, 80 reusable modules, automated architecture search (AAS) and hardware-aware neuromorphic deployment. Our AAS constructs task-specific decoders without manual design and extends into a large language model (LLM)-driven closed-loop mode supporting single-task, multitask and cross-species decoder design. A single-command pipeline compiles trained decoders for neuromorphic hardware, reducing power consumption by 30 to 50 times while preserving decoding performance. An interactive visualization software enables code-free exploration of neural recordings and decoding outputs. BCIJelly is validated across five BCI paradigms (motor, visual, speech, emotion and auditory) in humans, macaques and mice, providing an extensible ecosystem connecting data standardization, decoder development, systematic evaluation and hardware-aware deployment for BCI research.

neuroscience↗

bioRxiv2026

Tyrosine hydroxylase–mediated neuroimmune crosstalk regulates antitumor immunity in glioblastoma during oncolytic herpes virotherapy

Neuroimmune crosstalk is increasingly recognized as a key regulator of tumor progression and therapeutic response, yet its role in central nervous system (CNS) tumors remains poorly understood. Here, we investigate tyrosine hydroxylase (TH)-mediated neuronal signaling in glioblastoma (GBM) and its impact on antitumor immunity and response to oncolytic virotherapy (OV). We show that TH cells are widely distributed within the GBM microenvironment, including neurons, astrocytes, and immune cells, and are enriched at the tumor margin. In addition, TH cells are present in the tumor-draining lymph nodes (TDLNs) of GBM, where they localize near lymphatic vessels and are associated with lymphangiogenesis. Notably, a subset of CD3TH T cells is detected within lymphatic structures of TDLNs, suggesting immune-intrinsic catecholamine signaling. Single-cell RNA sequencing reveals that noradrenergic signaling, particularly via {beta}2-adrenergic receptors (ADRB2), predominates in tumor-infiltrating myeloid cells and is dynamically regulated by therapy. Intratumoral administration of oncolytic herpes simplex virus (oHSV) upregulates ADRB2 expression in macrophages, whereas systemic chemo-immunotherapy induces distinct receptor modulation patterns in tumors and TDLNs. Functionally, pharmacologic {beta}-adrenergic blockade significantly enhances the efficacy of oHSV therapy in orthotopic GBM and subcutaneous melanoma models, resulting in reduced tumor growth, increased tumor cell death, and enhanced CD8 T cell infiltration. Similarly, direct inhibition of TH enzymatic activity suppresses tumor progression and further potentiates OV. Mechanistically, TH inhibition not only promotes tumor-infiltrated cytotoxic immune cells CD8, NK and {gamma}{delta} T cells, but also suppresses the activity of immunosuppressive myeloid cells, including transcriptional (Fos), and metabolism (Arg) modification in M2 macrophages and other immune cells. Collectively, these findings identify TH-mediated neuroimmune signaling as a critical regulator of tumor immunity in GBM and demonstrate that targeting catecholaminergic pathways or downstream neuroimmune crosstalk pathways can enhance the efficacy of OV. This study provides a rationale for integrating neural modulation into immunotherapeutic strategies for CNS malignancies.

Cancer Biology↗

bioRxiv2026

BTK inhibition enhances immunovirotherapy in glioblastoma via tertiary lymphoid structure modulation

Glioblastoma (GBM) is a highly aggressive type of glioma that is resistant to immunotherapy and is associated with poor prognosis, largely due to its immunosuppressive tumor microenvironment. Brutons tyrosine kinase (BTK) is a non-receptor kinase that not only plays an important role in oncogenic signaling, particularly in tumor growth, but also regulates the activity of tumor-infiltrating myeloid cells, including dendritic cells, macrophages, and microglia in brain tumors. High BTK expression is associated with poor survival in patients with glioma. Oncolytic herpes simplex virus type 1 (oHSV)-derived virotherapy, a novel treatment strategy, has demonstrated effectiveness against GBM; however, its efficacy is limited by the tumor microenvironment. In this study, we found that BTK is predominantly expressed in GBM-infiltrating myeloid cells. Intratumoral injection of oHSV not only promotes infiltration of myeloid cells and T cells but also activates BTK in these myeloid cells, thereby limiting oHSV infection and replication in tumor cells. Combination treatment with BTK inhibitor ibrutinib improves anti-tumor efficacy of oHSV in both human GBM12 xenograft and syngeneic murine GSC005 models. Mechanistically, BTK inhibition increases oHSV-mediated tumor cell death (cleaved caspase-3) and cytotoxic CD8 T cell infiltration, while decreasing tumor cell proliferation (Ki-67). BTK inhibition not only suppresses oHSV clearance by tumor-infiltrating microglia and macrophages but also reduces their pro-invasive effects on tumor cells. Addition of IDO inhibitor, an immune modulator, further prolongs survival in tumor-bearing mice in a syngeneic GBM model. Single-cell mRNA sequencing (scRNA-seq) analysis indicates that combination treatment modifies key signaling pathways in both tumor-infiltrating myeloid cells (macrophages and microglia) and CD8 T cells. Further analysis shows that BTK inhibition, with or without IDO inhibition, promotes the formation of tumor-infiltrating tertiary lymphoid structures (TLS) during intratumoral oHSV treatment, subsequently remodeling T cell, NKT cell, and monocyte-macrophage populations. These results indicate that BTK inhibition exerts multifaceted effects in enhancing the anti-tumor efficacy of oHSV therapy.

cancer biology↗

bioRxiv2026

A dataset of simultaneous two-photon calcium imaging and auditory discrimination behavior

The emerging field of artificial intelligence for neuroscience relies heavily on high-quality, standardized datasets. Data linking high-dimensional neural population dynamics with complex behavior is crucial for advancing the field of brain-computer interfaces. However, few existing datasets provide these paired modalities in a standardized, AI-ready format, limiting their immediate utility for computational modeling. Here, we present a large-scale dataset comprising simultaneous two-photon calcium imaging of the primary auditory cortex (A1) and detailed behavioral records in mice. The animals performed an auditory two-alternative forced-choice (2AFC) categorization task, classifying tones as low or high frequency by licking left or right water ports. The raw neural and behavioral data were preprocessed, synchronized, and formatted into trial-aligned multi-dimensional tensors. Comprehensive validation confirms that the recorded populations exhibit consistent, high-fidelity temporal dynamics and distinct frequency tuning across all experimental conditions. To evaluate the datasets capacity for neural decoding, we benchmarked it across diverse machine learning architectures and deep learning networks. By bridging biological recordings and behavior, this open-access dataset serves as a valuable benchmark for developing novel decoding algorithms and testing brain-inspired computational models.

neuroscience↗

bioRxiv2026

Aligned recordings of neural spiking activity and licking behavior in thirsty mice

Electrophysiological data, which serve as a biological signal that bridges neural activity and behavioral tasks, provide an innovative approach to neuroscience research. In this study, we constructed a dataset that contains over 2000 neurons across 117 days recorded in 20 mice containing 28,573 trials. Data for 5 mice were collected from the Secondary Motor Cortex (M2) region 8 mice was derived from the Ventrolateral Striatum (VLS) and 7 mice were from Substantia Nigra pars Reticulata (SNR). We induced licking behavior in head-fixed mice by periodically delivering water through a spout while simultaneously recording spiking activity from three brain regions and behavior related electrical signals. This dataset ensures precise temporal alignment between neural activity and behavioral events, offering a robust foundation for investigating neural encoding mechanisms and simulation of neural activities. This dataset establishes a precise spike-to-event mapping, which enables high decoding accuracy using Multilayer Perceptron (MLP) and Support Vector Machine (SVM). It can serve as a high-quality benchmark for developing encoding and decoding algorithms in neural networks, particularly Spiking Neural Networks (SNNs).

neuroscience↗

bioRxiv2026

The prevalence of protein misfolding as a mechanism for hereditary deafness

Hearing loss is the most common sensory deficit impacting [~]5% of the worlds population. The Deafness Variation Database (DVD) is a public resource of deafness variants, containing over 380,000 missense variants across 224 genes, with 303,577 classified as a variant of uncertain significance (VUS). To address the challenge of evaluating each deafness associated VUS, we evaluate a family of probabilistic frameworks to quantify the strength of computational evidence based on ACMG/AMP recommendations. First, CADD and REVEL are compared using Bayesian models parameterized using either a ClinVar 2019 dataset or labeled DVD variants. The REVEL model built using the DVD dataset demonstrates the best accuracy, sensitivity, and specificity. Incorporation of (in)tolerance to missense variation based on sorting each gene into three bins (tolerant, average, intolerant) shows that intolerant DVD genes are consistent with a higher prior probability of being pathogenic (25.7%) than average (10.7%) or tolerant (8.7%) genes. Finally, the impact of protein folding stability was incorporated using a 2D likelihood, which surpassed the simpler models while also offering a biophysical rationale for the disease mechanism. The protein folding-informed Bayesian model results in 28,866 prioritized VUSs reaching a posterior probability of pathogenicity above 98% with a false positive rate of only 0.14%. Overall, 54,752 missense variants are predicted to cause protein folding destabilization of greater than 1.0 kcal/mol, while 18,706 of the 28,886 prioritized VUS (65%) surpass this threshold. From these VUSs, we identify twelve probands where the patients genetic diagnosis is upgraded to likely pathogenic/pathogenic. We highlight two variants that cause clear structural disruption, demonstrating the impact of biophysical characterization on variant evaluation. Author SummaryWe investigate the impacts of single amino acid changes on protein structure and folding in the context of hearing loss. Hearing loss is the most common impairment of the main senses affecting nearly 5% of the worlds population. About 45% of people with hearing loss receive a diagnosis after targeted genetic testing. Here, we integrate biophysical data that quantifies the effect of a change to protein sequence on protein folding in combination with genetic data to improve our ability to identify protein amino acid changes that are likely to impact hearing. Our work leads to 12 patients receiving an upgraded diagnosis with their variant disrupting protein stability. Although the method is applied to hearing loss, it can be used for interpreting protein sequence changes in other disease contexts.

genetics↗

bioRxiv2025

Conserved early steps of stemmadenine biosynthesis

Stemmadenine acetate is a pivotal intermediate in the production of pharmacologically active monoterpene indole alkaloids. Here, we identify orthologs of stemmadenine acetate pathway genes (SGD, GS, GO, Redox1, Redox2, SAT) from Tabernanthe iboga. We characterize these enzymes in vitro, and additionally, we reconstitute stemmadenine acetate biosynthesis in Nicotiana benthamiana, comparing the formation of intermediates and shunt products that are produced when previously characterized orthologs from the related plant Catharanthus roseus are used. Ortholog pairs are catalytically indistinguishable, except in the case of GS. Surprisingly, the T. iboga ortholog catalyzes formation of an alternate stereoisomer 19Z-geissoschizine, seeding a low-flux Z-series in heterologous reconstitution systems in vitro and in planta. We additionally characterize the major shunt products that arose during reconstitution of stemmadenine acetate biosynthesis. We show that the substrate promiscuity of Redox2 results in formation of the shunt products 16(R/S)-isositsirikines, hampering pathway flux and yields. Additionally, we show that stemmadenine can be oxidized by endogenous N. benthamiana enzymes, leading to the shunt product condylocarpine. Nevertheless, we could produce stemmadenine at a 6 mg yield from 19E-geissoschizine by heterologous expression in N. benthamiana. Overall, we highlight the prospects for milligram production of important biosynthetic intermediates in N. benthamiana.

biochemistry↗

bioRxiv2025

Mapping Human Survivability at Extreme Wet-Bulb Temperatures 32-35°C

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/677706v1_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@70b72dorg.highwire.dtl.DTLVardef@9fbfeorg.highwire.dtl.DTLVardef@17f05a5org.highwire.dtl.DTLVardef@4b977b_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO C_FIG HighlightsWe report the first empirical human survival map across wet-bulb temperatures of 32-35{degrees}C. HydraAon and behavior cooling extend survival beyond the 6-hour theoreAcal limit. Females consistently tolerate extreme humid heat longer than males in controlled trials. Our findings refine heat-health models and inform equitable climate adaptaAon strategies. In briefExtreme humid heat, measured by wet-bulb temperature (Tw), is nearing the threshold of human survivability under climate change. In controlled trials with 36 adults, we provide the first empirical survival map across Tw=32-35{degrees}C, showing that hydration and behavioral adaptations extend endurance beyond the theoretical 6-hour limit, with women tolerating heat longer than men. These findings refine human heat tolerance thresholds and inform equitable climate adaptation and heat-health strategies. SCIENCE FOR SOCIETYAs climate change accelerates, extreme humid heat events--measured by wet-bulb temperature (Tw)--pose a growing threat to human survival. This study provides the first empirical benchmarks of human survivability across Tw=32-35{degrees}C, clarifying how long healthy adults can endure these conditions under full hydration and minimal activity. We find that while the theoretical 6-hour survival limit at Tw=35{degrees}C is partly supported, both men and women can withstand these conditions for 7-8 hours when hydrated, with women demonstrating greater tolerance. At Tw=34{degrees}C, projected heatstroke onset extends beyond 12-16 hours, while conditions at Tw=32-33{degrees}C remain largely compensable for more than 30 hours. These results challenge prevailing heat tolerance models, underscore the role of behavioral and physiological adaptations, and provide critical data for designing equitable heat-health policies and early warning systems that account for sex differences and vulnerable populations. Climate change-driven extreme heat events increasingly threaten human health. Here, we provide the first empirical map of human survivability across four extreme wet-bulb temperatures (Tw=32- 35{degrees}C) in controlled trials. Thirty-six healthy young adults (20 males, 16 females) were exposed to twelve dry-bulb temperature and relative humidity combinations in shaded indoor settings under full hydration. Participants remained seated and performed light office tasks until core temperature (Tcore) reached 39{degrees}C or exposures lasted 8 hours. Clinical heatstroke threshold (Tcore=40.5{degrees}C) were projected by extrapolating from our direct Tcore measurements during controlled exposure trials. At Tw=35{degrees}C, males and females reached heatstroke in approximately 7.1-7.7 and 8.3-8.6 hours, respectively--partly supporting the 6-hour theoretical limit while revealing enhanced short-duration resilience when hydrated. At Tw=34{degrees}C, projected times are 12.5-12.9 hours (males) and 15.5-16.2 hours (females). Conditions at Tw=32{degrees}C and 33{degrees}C remained (quasi-)compensable, with projected heatstroke onset after 33-35 hours. Females consistently tolerated heat stress longer than males. These physiological benchmarks are critical for improving heat-health models and embedding sex-specific vulnerability in equitable adaptation strategies amid accelerating global warming.

physiology↗

bioRxiv2025

DeepHalo: Deep Learning-Powered Exploration of Halogenated Metabolites Uncovering Antibacterial Depsipeptides

In the omics era, confident high-throughput analytical tools are crucial for the efficient identification of metabolites. Here, we present DeepHalo, a deep learning-integrated and hierarchically optimized workflow designed for high-throughput exploration of halogenated metabolites from high-resolution mass spectrometry-based metabolomics. DeepHalo leverages deep learning models combined with a comprehensive scoring to enhance the reliability of halogen predictions. It integrates PyOpenMS for fast isotope pattern detection and incorporates a halogen-based dereplication algorithm with GNPS molecular networking to efficiently exploit and annotate halogenates from complex biological matrices. To validate its performance, DeepHalo was applied to explore halogenated metabolites from 1,296 microbial culture crude, leading to the discovery of six families of structurally diverse halogenated molecules. This included a new class of cyclic depsipeptides, aglomycins A-E, featuring rare 3-chloroanthranilic acid and/or epoxyvaline blocks. Additionally, a plausible biosynthetic pathway of aglomycins was proposed through bioinformatics analyses and targeted gene knockout experiments. Bioassays revealed that aglomycin A exhibits synergistic antibacterial activity with linezolid against vancomycin-resistant Enterococcus faecium (VRE) both in vitro and in vivo. We envision that DeepHalo (freely available at https://github.com/xieyying/DeepHalo) will become a powerful tool for accelerating the discovery of halogenated "dark matter".

biochemistry↗

bioRxiv2025

Reversion of a RND transporter pseudogene uncovers latent stress resistance in Brucella ovis

Small-molecule screens can advance therapeutic discovery while yielding new insights into pathogen biology. Through a luminescence-based screen, we identified clinically approved dihydropyridines that reduced the fitness of the intracellular pathogen Brucella ovis within mammalian phagocytes. Given the established role of dihydropyridines as inhibitors of mammalian L-type calcium channels and our observation that drug treatment perturbed calcium and manganese levels in host phagocytes, we initially hypothesized a host-directed mechanism of action. However, dose-response assays in axenic medium revealed that these drugs can directly inhibit B. ovis growth. To investigate the genetic basis of B. ovis susceptibility to dihydropyridine treatment, we selected for mutants capable of growing in the presence of cilnidipine. Cilnidipine-resistant isolates carried single-base deletions in the bepE pseudogene that restored an open reading frame encoding an RND-family transporter subunit. B. ovis is an ovine venereal pathogen that has experienced significant pseudogenization in its recent evolutionary history. Frameshift mutations that restored bepE function in B. ovis increased its resistance not only to dihydropyridines but also to a broad range of membrane-disrupting agents, including bile acid. Conversely, deleting bepE in Brucella abortus, a related zoonotic species that retains an intact version of the gene, increased its sensitivity to bile acid in vitro and to cilnidipine in the intracellular niche. We conclude that bepE is a key determinant of chemical stress resistance in Brucella spp., and that its pseudogenization in B. ovis contributes to the documented hypersensitivity of this host-restricted lineage to chemical stressors. Author SummaryBrucella species are intracellular bacterial pathogens that infect diverse wildlife and livestock and can also cause disease in humans. We developed a high-throughput assay to identify compounds that reduce the intracellular fitness of Brucella ovis, a globally distributed pathogen of sheep. This screen identified two dihydropyridine-class hypertension drugs as inhibitors of B. ovis fitness within mammalian cells. While dihydropyridines are established inhibitors of calcium channels in mammals, our results revealed that they also directly inhibited B. ovis growth. The B. ovis genome contains over 300 pseudogenes, and we discovered that resistance to dihydropyridines and other chemical stressors can emerge through frameshift mutations that resurrect the function of bepE, a conserved pseudogene encoding a membrane transporter subunit. In other Brucella species, bepE remains intact, and we show that deleting this gene in the zoonotic pathogen Brucella abortus increases susceptibility to both dihydropyridine treatment and membrane-disrupting stress. Our study provides evidence that pseudogenes like bepE can serve as a latent reservoir of adaptive functions in B. ovis, including drug and stress resistance.

microbiology↗

bioRxiv2025

It runs in the family: Discovery of enzymes in the oleuropein pathway in Olive (Olea europaea) by comparative transcriptomics

Olive (Olea europaea L.) is one of the most important crop trees, with olive oil being a key ingredient of the Mediterranean diet. Oleuropein, an oleoside-type secoiridoid, is the major determinant of flavor and quality of olive oil. Iridoid biosynthesis has been elucidated in Catharanthus roseus, which produces secologanin-type secoiridoids, but iridoid biosynthesis in other species remains unresolved. In this work, we sequenced RNA from olive fruit mesocarp of six commercial olive cultivars with varying oleuropein content, during maturation and ripening. Using this data we discovered three polyphenol oxidases with oleuropein synthase (OS) activity, a novel oleoside-11-methyl ester glucosyl transferase (OMEGT) synthesizing a potential intermediate in the route, and a 7-epi-loganic acid O-methyltransferase (7eLAMT). Interestingly, integrating transcriptomics data from 15 plant species from three iridoid-producing plant orders (Lamiales, Gentianales, and Cornales), and tissue expression panels from Jasminum sambac and Fraxinus excelsior, we discovered two 2-oxoglutarate dependent dioxygenases (named 7eLAS) that synthesize 7-epi-loganic acid; in contrast C. roseus 7-deoxy-loganic acid hydroxylase (7DLH), a known bottleneck in MIA production, is a cytochrome p450. This comparative co-expression method, which combines guilt by association and comparative transcriptomics approaches, can successfully leverage big datasets for untargeted discovery of enzymes. Key FindingsO_LIExpression of genes involved in iridoid biosynthesis, from the early MEP pathway to the last step of oleuropein biosynthesis, decreases during olive fruit maturation. C_LIO_LIWe discovered an oxoglutarate dependent dioxygenase, 7-epi-loganic acid synthase (7eLAS), catalyzing the stereoselective oxidation of 7-deoxy-loganic acid to 7-epi-loganic acid, in a reaction analogous to C. roseus 7-deoxy-loganic acid hydroxylase (7DLH), a cytochrome p450. C_LIO_LIWe report a 7-epi-loganic acid O-methyltransferase (7eLAMT) orthologous to Catharanthus roseus loganic acid O-methyltransferase and found a novel oleoside-11-methyl ester glucosyl transferase (OMEGT) synthesizing 7-{beta}-1-D-glucopyranosyl-oleoside-11-methyl ester, a potential intermediate in the oleuropein biosynthesis route. C_LIO_LIWe discovered three olive polyphenol oxidases that have oleuropein synthase (OS) activity, catalyzing the conversion of ligstroside to oleuropein. C_LI

biochemistry↗

bioRxiv2025

Elucidation and de novo Reconstitution of Glyceollins Biosynthesis

Glyceollins are phytoalexins produced by soybeans in response to stressors such as pathogen invasion, injury, and environmental challenges. In addition to their antibacterial and antifungal activities, these compounds have attracted significant attention for their potential anticancer, anti-inflammatory, and antioxidant properties. However, their limited accessibility--due to the challenges and high costs associated with organic synthesis or purification from treated soybean seedlings--has hindered further physiological and biochemical studies. Moreover, the incomplete understanding of glyceollin biosynthesis, particularly the final cyclization steps, remains a major barrier to elucidating the physiological functions of glyceollins and achieving sustainable production through synthetic biology. In this study, we identified previously uncharacterized genes encoding two reductases for 7,2,4-trihydroxyisoflavanol (THI) biosynthesis and five P450 enzymes responsible for the final oxidative cyclization in glyceollin I, II, and III biosynthesis, thereby completing the entire glyceollin biosynthetic pathway. By reconstructing the pathway de novo through synthetic biology, we achieved the successful production of glyceollins from simple carbon sources in bakers yeast. This work advances the understanding of glyceollin biosynthesis in soybean, facilitates sustainable production in microbial hosts and offers new opportunities for their application in agriculture and biology.

biochemistry↗

bioRxiv2024

ATF5-Dependent GDF15 Expression Mediates Anesthesia-Induced Neuroprotection Against Stroke

The incidence of perioperative stroke, a rare but severe complication, is increasing in aging populations. Although anesthetics such as sevoflurane may provide protective preconditioning against ischemic injury, clinical outcomes have been inconsistent. In this study, we demonstrated that sevoflurane-induced neuroprotection is associated with the upregulation of genes involved in the mitochondrial unfolded protein response (UPRmt) and mitochondrial bioenergetic metabolism. Our findings emphasize the critical role of ATF5, a key transcription factor, in mediating these protective effects. Specifically, we observed that sevoflurane preconditioning significantly upregulates ATF5 and its downstream target GDF15 - a regulator of mitochondrial function - in the cerebral cortex. Notably, we found that this mechanistic pathway was not activated in the brain of aged mice, suggesting that age-specific strategies may be necessary for reducing the risk of perioperative stroke. Considering the steadily increasing age of patients, therapeutic approaches that enhance mitochondrial function in the aged brain may provide additional protection against perioperative stroke.

neuroscience↗

bioRxiv2024

Divergent biosynthesis of monoterpene indole alkaloids from geissoschizine

Plants can generate structural diversity by enzymatic rearrangement of a central intermediate. 19E-geisssochizine is one such chemically versatile intermediate that plays a central role in the biosynthesis of monoterpene indole alkaloids such as strychnine, ibogaine and vinblastine. Here we report how 19E-geissoschizine undergoes oxidative transformations to generate four distinct alkaloid scaffolds through the action of three biosynthetic enzymes. Using in vitro enzymatic assays and gene silencing, we demonstrate how these three cytochrome P450 enzymes in the medicinal plant Catharanthus roseus transform 19E-geisssochzine into strychnos, sarpagan, akuammiline-type, and mavacurane-type alkaloids. We use mutational analysis to show how minimal changes to the active site of these similar enzymes modulate product specificity. This work highlights how substrate reactivity and enzyme mutations work synergistically to generate chemical diversity.

biochemistry↗

bioRxiv2024

Connectomic Organization of the Suprachiasmatic Nucleus

The suprachiasmatic nucleus (SCN), the mammalian master clock, is a special structure dedicated to the time computation. However, a connectomic understanding of the nucleus as a whole is lacking. Using serial section electron microscopy, we reconstructed multiscale and multimodal SCN communication networks. Intra-SCN synaptic network consists of 9,566 morphologically similar neurons and 4.3 million synapses, organized into multiscale circuitries that interact with SCN-traversing axon fascicles. Strikingly, SCN neurons engage in unique soma-soma ephaptic interaction, forming 2,038 electrotonic integrative units with the largest overlapping the light-responsive area (LRA). SCNs paracrine modality contains 47,396 m3 dense core vesicles, with notable scarcity in the LRA, suggesting cross-modal coordination and functional integration. These distinct network features provide comprehensive insights into the system architecture of the mammalian circadian clock.

neuroscience↗

bioRxiv2024

Independent evolution of ipecac alkaloid biosynthesis

Ipecac alkaloids are medicinal monoterpenoid-derived tetrahydroisoquinoline alkaloids found in two distantly related plants: Carapichea ipecacuanha (Gentianales) and Alangium salviifolium (Cornales). We have elucidated ipecac alkaloid biosynthesis in both species, conclusively demonstrating that biosynthesis of the structurally complex ipecac alkaloid protoemetine has evolved independently. We show that although protoemetine biosynthesis proceeds via the same chemical logic in both species, each plant uses a distinct monoterpene precursor. Moreover, we provide evidence that both plants initiate ipecac biosynthesis by a non-enzymatic Pictet-Spengler reaction, and we elucidate the biosynthetic fate of both the 1R and 1S stereoisomers that are produced in this non-stereoselective reaction. Phylogenetic analyses clearly show independent pathway evolution through parallel and convergently evolved enzymes. This work provides insight into how nature can capitalize on highly reactive starting substrates, the manner in which multi-step pathways can arise, and also lays the foundation for metabolic engineering of these important medicinal compounds.

plant biology↗

bioRxiv2024

Streamlined screening platforms lead to the discovery of pachysiphine synthase from Tabernanthe iboga

Plant-specialized metabolism is largely driven by the oxidative tailoring of key chemical scaffolds catalyzed by cytochrome P450 (CYP450s) enzymes. The monoterpene indole alkaloids tabersonine and pseudo-tabersonine, found in the medicinal plant Tabernanthe iboga, are extensively modified by oxidative reactions. Here we developed a streamlined screening strategy to screen the activity of T. iboga CYP450s in Nicotiana benthamiana. Using multigene constructs encoding the biosynthesis of tabersonine and pseudo-tabersonine scaffolds, we set out to uncover the CYP450s responsible for oxidative transformations of these scaffolds. Our approach identified two T. iboga cytochrome P450 enzymes: pachysiphine synthase (PS) and 16-hydroxy-tabersonine synthase (T16H). These enzymes catalyze an epoxidation and site-specific hydroxylation of tabersonine to produce pachysiphine and 16-OH-tabersonine, respectively. We further demonstrated that these genes produced the expected products when expressed in Catharanthus roseus flowers. This work provides new insights into the biosynthetic pathways of MIAs and underscores the utility of N. benthamiana and C. roseus as platforms for the functional characterization of plant enzymes.

biochemistry↗

bioRxiv2024

Precise cell recovery by cell nucleus united transcript (CellCUT) for enhanced spatial transcriptomics

Cell segmentation is the first step in parsing spatial transcriptomic data, often a challenging task. Existing cell segmentation methods do not fully leverage spatial cues between nuclear images and transcripts, tending to produce undesirable cell profiles for densely packed cells. Here, we propose CellCUT to perform cell segmentation and transcript assignment without additional manual annotations. CellCUT provides a flexible computational framework that maintains high segmentation accuracy across diverse tissues and spatial transcriptomics protocols, showing superior capabilities compared to state-of-the-art methods. CellCUT is a robust model to deal with undesirable data such as low contrast intensity, localized absence of transcripts, and blurred images. CellCUT supports a human-in-the-loop workflow to enhance its generalizability to customized datasets. CellCUT identifies subcellular structures, enabling insights at both the single-cell and subcellular levels.

bioinformatics↗