Biology subjects

Bu, Y.

Publications and source records attributed to Bu, Y..

At least 19 recordsLinked to original sources

bioRxiv2026

Reconstructing True 3D Spatial Omics at Single-Cell Resolution

Capturing the three-dimensional (3D) organization of cells is essential for deciphering complex biological processes, yet comprehensive 3D spatial omics is severely hindered by the destructive nature of physical sectioning and the depth limitations of intact tissue imaging. Current computational methods rely on 2.5D stacking of discrete slices, which inherently disrupts tissue topology and fails to resolve continuous depth-dependent molecular gradients. To bridge this gap, we introduce DO_SCPLOWEEPC_SCPLOWSO_SCPLOWPATIALC_SCPLOW, an Optimal Transport flow matching framework that models tissue evolution as a continuous dynamic vector field. By solving the underlying probability flow ODEs, DO_SCPLOWEEPC_SCPLOWSO_SCPLOWPATIALC_SCPLOW enables the direct extraction of uninterrupted, infinitely resolvable tissue states at arbitrary spatial depths. Using Deep STAR/RIBOmap 3D technologies, we demonstrate that DO_SCPLOWEEPC_SCPLOWSO_SCPLOWPATIALC_SCPLOW achieves improved 3D reconstruction fidelity relative to 2.5D approaches, yielding structures that more closely recapitulate native tissue microenvironments in real-world datasets. Across diverse spatial omics modalities, including spatial proteomics using imaging mass cytometry in human breast cancer and spatial transcriptomics using openST in head and neck squamous cell carcinoma metastatic lymph nodes, DO_SCPLOWEEPC_SCPLOWSO_SCPLOWPATIALC_SCPLOW produces biologically interpretable and high-fidelity reconstructions across datasets. We evaluated the scalability and robustness of DO_SCPLOWEEPC_SCPLOWSO_SCPLOWPATIALC_SCPLOW on a large-scale mouse brain dataset, reconstructing a continuous 3D cellular atlas comprising 39 million cells within 41.6 hours. Systematic downstream characterization validated its ability to recapitulate consistent spatial architectures, cell-type distributions, transcriptomic patterns, and microenvironmental structures across brain regions. Collectively, these results demonstrate DO_SCPLOWEEPC_SCPLOWSO_SCPLOWPATIALC_SCPLOW as a generalizable and efficient solution for true 3D spatial reconstruction across scales and modalities. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=117 SRC="FIGDIR/small/721395v2_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@1a19624org.highwire.dtl.DTLVardef@188361forg.highwire.dtl.DTLVardef@199321corg.highwire.dtl.DTLVardef@a8f411_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2026

Precision DNA Impurity Reduction Approaches for Ultra-Pure rAAV Manufacturing

Recombinant adeno-associated virus (rAAV) vectors are a leading platform for gene delivery in basic and clinical research, yet large-scale manufacturing remains constrained by residual nucleic-acid impurities that compromise safety. In this study, we profiled the DNA species packaged within rAAV capsids and identified plasmid backbone sequences and host cell genomic DNA (hcDNA) as predominant contaminants. To mitigate this critical quality attribute, we implemented upstream strategies designed to fragment or excise backbone DNA, including TelN/TelROL excision, I-SceI meganuclease digestion, CRISPR/Cas9 cleavage, and Cre/LoxP recombination. Quantitatively, TelN/TelROL and I-SceI reduced encapsidated plasmid backbone DNA to approximately 20-30% and 20-40% of baseline levels, respectively, while CRISPR/Cas9 lowered it to about 10-20%. Notably, the Cre/LoxP system eliminated detectable plasmid backbone DNA without compromising vector-genome titers, indicating preserved genomic integrity. Additionlly, supplementating cell culture with a caspase inhibitor significantly reduced hcDNA contamination in rAAV particles to 1-5% of the baseline level. Collectively, these interventions provide practical bioprocess frameworks that markedly enhance rAAV purity via targeted DNA minimization and prevention of hcDNA fragmentation, thereby strengthening the safety profile of rAAV therapeutics in alignment with current Good Manufacturing Practice (cGMP) expectations.

molecular biology↗

bioRxiv2025

Decoupling Efficacy from Toxicity: Engineering Spatial Control in AAV-Mediated Gene Therapy

Hepatotoxicity poses a critical safety challenge for AAV-mediated gene therapy. To mitigate this, we evaluated strategies to minimize off-target hepatic transduction using an antibody expression model. We compared (i) muscle-restricted wild-type AAV9 expression and (ii) a novel myotropic capsid variant, AAV.eM. In humanized B-NDG mice bearing Raji-Luc lymphomas, intravenous administration of AAV9-MHCK7 encoding an CD19CD3 bispecific T-cell engager failed to reduce tumor burden. Conversely AAV.eM-MHCK7-CD19CD3 substantially alleviated tumor burden and achieved lymphoma clearance. By leveraging tissue-specific microRNAs, precise restriction of AAV.eM-mediated transgene expression to skeletal or cardiac muscle was achieved. Incorporating a heart-specific miR-208a binding site into the transgenes 3UTR did not compromise therapeutic efficacy when delivered via AAV.eM-MHCK7. Intramuscular delivery of AAV9-MHCK7-CD19CD3 or AAV.eM-MHCK7-CD19CD3 both cleared Raji-Luc tumors at a dose of 5 x 1012 vg/kg, underscoring the advantage of localized and targeted rAAV delivery over systemic administration. Notably, only AAV.eM-MHCK7-CD19CD3 achieved tumor eradication at a tenfold lower intramuscular dose (5 x 1011 vg/kg), reducing manufacturing costs and risks of dose-dependent immunogenicity and toxicity. Our findings demonstrate that combining tissue-specific targeting--via engineered capsids or tissue-selective promoters--with local delivery robustly reduces off-target hepatic expression, providing a strategic framework for enhancing the safety of AAV-based gene therapies.

cancer biology↗

bioRxiv2025

Phylogenomics of Rhinogobius gobies reveal northern-southern divergence and trait evolution in China

The goby genus Rhinogobius represents a prominent radiation of freshwater fishes in East Asia, with China serving as its evolutionary hotspot harboring over 39 endemic species. Despite their ecological dominance in riverine ecosystems, phylogenetic relationships among mainland Chinese Rhinogobius species remain poorly resolved due to limited sampling and morphological conservatism, particularly in meristic traits traditionally used for classification. To address these knowledge gaps, we employed a nuclear gene enrichment approach to target 4,434 single-copy nuclear loci across 26 species spanning all major river basins in eastern and southern China. Our phylogenomic analyses based on sequence of 2,137 cleaned-up loci (477,822 bp) recovered a robust topology dividing Rhinogobius into three primary lineages: an early-diverging of R. similis sister to two major clades, the northern vs. southern clades exhibiting strong geographic structuring. Fossil-calibrated molecular dating placed the crown age of Rhinogobius at [~]9.33 million years ago (Ma) during the late Miocene, with a pivotal north-south divergence at [~]5.96 Ma coinciding with major paleo-drainage reorganizations in East Asia. Ancestral state reconstruction revealed that the southern clade uniquely evolved unstable first dorsal fin formula and near-universal presence of predorsal scales. By integrating the most extensive geographic and taxonomic sampling to date with phylogenomic data, this study establishes: (1) a revised evolutionary timeline for Rhinogobius diversification linked to East Asian geomorphological history, (2) diagnostic morphological characters for major clades, and (3) a robust phylogenetic framework to guide future taxonomic revisions and conservation efforts for this ecologically significant group. HighlightsO_LIPhylogenomics resolves north/south split in Chinese Rhinogobius gobies. C_LIO_LIFossil-dated 9.3 MYA origin of Rhinogobius and rapid divergence at 5.96 MYA. C_LIO_LISouthern lineages exhibit more predorsal scales and unstable first dorsal fin formula. C_LIO_LIFirst genomic backbone for species revision and adaptation study. C_LI

zoology↗

bioRxiv2025

Engineering novel AAV capsids by global de-targeting and subsequent muscle-specific tropism in mice and NHPs

Recombinant adeno-associated viral (rAAV) vectors are a potent tool, but their clinical application is restricted by insufficient target tissue transduction and liver toxicity. We employed a novel two-step engineering strategy to create novel rAAV capsids with global tissue de-targeting, then produced strong tissue-specific expression by adding a peptide sequence. We created a novel capsid, AAV.Zero1, with globally de-targeted transduction by loop swapping domains from AAV9 into AAV2. Making an R585A substitution (AAV.Zero2) re-targeted tissues but deleting residues 585-587 (AAV.Zero3) abrogated transduction. Inserting a myogenic peptide into AAV.Zero3 produced a novel capsid (AAV.eM) with strong muscle-specific transgene expression while maintaining minimal off-target expression, including in liver, which was conserved in two mouse strains and non-human primates. AAV.eM showed similar expression as the leading myotropic vector MyoAAV.4A but had a more favorable safety profile. Importantly, AAV.eM was able to functionally rescue a mouse model of Duchenne Muscular Dystrophy following systemic delivery of a micro-dystrophin gene. Thus, AAV.eM is an improved myotropic rAAV capsid that de-targets other tissues, especially the liver, and proof-of-concept for a platform to create capsids with specific properties that translate across species by addition of peptides onto low transduction backbones.

bioengineering↗

bioRxiv2024

Deep Learning Unravels Potential Antibiotic-Resistance Drugs against Klebsiella Pneumoniae

BackgroundKlebsiella pneumoniae (KP) is a critical pathogen recognized by the World Health Organization due to its high mortality rates and resistance to antibiotics. Developing effective treatments for KP is essential to mitigate the growing threat of multidrug-resistant infections. This study leverages deep learning techniques to identify and validate potential drug candidates against KP, accelerating the drug discovery process and reducing associated risks. ResultsA comprehensive dataset was constructed, comprising 3,475 drugs selected from the DrugBank database, filtered for their potential efficacy and safety. Using convolutional neural networks (CNNs) for drug-drug interaction analysis, a prediction accuracy of 72% was achieved. Evolutionary Scale Modeling (ESM) was employed to calculate molecular similarities between drugs and KP strains, identifying five promising candidates with structural similarities exceeding 85% to known KP targets. A drug repurposing knowledge graph (DRKG) further validated these findings, with the PairRE model outperforming other approaches in ranking potential drug candidates. ConclusionsThis study presents a scalable and efficient framework for identifying and validating drug candidates to combat antibiotic-resistant KP. By integrating deep learning models, molecular similarity analysis, and knowledge graph techniques, this approach shortens the drug discovery timeline from years to months. The identified candidates hold significant promise for preclinical and clinical testing, offering a pathway to effective therapies against KP and other multidrug-resistant pathogens.

bioinformatics↗

bioRxiv2024

Deep Learning with Evolutionary Scale Modeling to discover Antibiotic-Resistance Drug against Klebsiella Pneumoniae

The development of effective antibiotics is crucial in combating antibiotic-resistant pathogens such as Klebsiella pneumoniae (KP). This study utilizes deep learning, specifically Evolutionary Scale Modeling (ESM), to identify potential drug candidates. By analyzing molecular similarities between known drugs and KP strains, we aim to accelerate the discovery of new treatments and reduce the risks associated with clinical trials. This approach may shed new lights in the field of drug repositioning.

bioinformatics↗

bioRxiv2024

The m6A Reader ECT1 Mediates Seed Germination via the DAG2-ECT1-PHYB Regulatory Cascade in Arabidopsis

The N6-methyladenosine (m6A) RNA modification, a significant epitranscriptomic mark, is integral to plant growth and development. m6A reader proteins are capable of recognizing m6A modifications to impact RNA metabolic and biological processes. Here, we discovered ECT1 as an m6A reader protein that directly binds to the m6A site and plays a role in positively regulating seed germination in the context of gibberellins (GAs). ECT1 undergoes m6A modification to ensure its own stability and engages in self-interaction to enhance its protein stability. Moreover, ECT1 establishes a regulatory circuit with DAG2, which has been reported to play a positive regulatory role in GA-mediated seed germination. We found that DAG2 directly binds to the ECT1 promoter to control its transcription. ECT1 modulates the mRNA stability of DAG2. Furthermore, we identified PHYB, which encodes a key photoreceptor controlling GA-regulated seed germination, as a common downstream target of DAG2 and ECT1. DAG2 and ECT1 regulate the expression of PHYB at the transcriptional and post-transcriptional levels, respectively. ECT1 binds directly to PHYB, thereby influencing its stability. While DAG2 binds to the PHYB promoter to regulate its transcription. Together, these results unveil that ECT1 participates in the m6A signaling pathway through complex and multifaceted molecular mechanisms, broadening the spectrum of m6A reader functions and deepening our insight into the m6A signaling network in Arabidopsis.

plant biology↗

bioRxiv2024

Unraveling the Role of Urea Hydrolysis in Salt Stress Response during Seed Germination and Seedling Growth in Arabidopsis thaliana

Urea is intensively utilized as a nitrogen fertilizer in agriculture, originating either from root uptake or from catabolism of arginine by arginase. Despite its extensive use, the underlying physiological mechanisms of urea, particularly its adverse effects on seed germination and seedling growth under salt stress remains unclear. In this study, we demonstrate that salt stress induces excessive hydrolysis of arginine-derived urea, leading to an increase in cytoplasmic pH within seed radical cells, which, in turn, triggers salt-induced inhibition of seed germination (SISG) and hampers seedling growth. Our findings challenge the long-held belief that ammonium accumulation and toxicity are the primary causes of SISG, offering a novel perspective on the mechanism underlying these processes. This study provides significant insights into the physiological impact of urea hydrolysis under salt stress, contributing to a better understanding of SISG.

plant biology↗

bioRxiv2024

Dual Targeting of STING and PI3Kγ Eliminates Regulatory B Cells to Overcome STING Resistance for Pancreatic Cancer Immunotherapy

The immune suppression in tumors and lymph nodes of pancreatic ductal adenocarcinoma (PDAC), regulated by suppressive myeloid cells and regulatory B (Breg) cells, hinders the effectiveness of immunotherapy. Although STING agonists activate myeloid cells to overcome immune suppression, it expands Breg cells, conferring STING resistance in PDAC. We discovered that blocking PI3K{gamma} during STING activation abolished IRF3 phosphorylation to eliminate Breg cells, while PI3K{gamma} inhibition sustained STING-induced IRF3 phosphorylation to preserve STING function in myeloid cells. Therefore, we developed a dual functional compound SH-273 and its albumin nanoformulation Nano-273, which stimulates STING to activate myeloid cells and inhibits PI3K{gamma} to eliminates Breg cells overcoming STING resistance. Nano-273 achieved systemic antitumor immunity through intravenous administration, which decreases Breg cells and remodels microenvironment in tumors and lymph nodes. Nano-273, combined with anti-PD-1, extended median survival to 200 days in transgenic KPC PDAC mice (KrasG12D-P53R172H-Cre), offering potential for PDAC treatment.

pharmacology and toxicology↗

bioRxiv2024

A new story of four Hexapoda classes: Protura as the sister to all other hexapods

Insects represent the most diverse animal group, yet previous phylogenetic analyses based on the morphological and molecular data have failed to agree on the evolutionary relationships of early insects and their six-legged relatives (together constituting the clade Hexapoda). In particular, the phylogenetic positions of the three early-diverging hexapod groups, the coneheads (Protura), springtails (Collembola), and two-pronged bristletails (Diplura), have been debated for over a century, with alternative topologies implying drastically different scenarios of the evolution of the insect body plan and hexapod terrestrialisation. We addressed this issue by sampling of all hexapod orders, and experimented with a broad range of across-site compositional heterogeneous models designed to tackle ancient divergences. Our analyses support Protura as the earliest-diverging hexapod lineage (Protura-sister) and Collembola as a sister group to the Diplura, a clade we refer to as Antennomusculata characterised by the shared possession of internal muscles in the antennal flagellum. The universally recognized Ellipura hypothesis is recovered under the site-homogenous LG model. Our cross-validation analysis shows that the CAT-GTR model that recovers Protura-sister fits significantly better than homogenous model. Furthermore, as a very unusual group, Protura as the first diverging lineage of hexapods is also supported by other lines of evidence, such as mitogenomics, comparative embryology, and sperm morphology. The backbone phylogeny of hexapods recovered in this study will facilitate the exploration of the underpinnings of hexapod terrestrialisation and mega-diversity.

evolutionary biology↗

bioRxiv2023

Status and Genetic Diversity of Hipposideros pratti in the Northernmost Part of Its Distribution

Hipposideros pratti is species of bat distributed in caves from Southeast Asia to the Qinling-Funiu Mountain area in China. To understand the dynamic changes in its distribution and evaluate the current health status of the species, we reviewed 48 caves in the northernmost area of the H. pratti distribution and conducted the first analysis of genetic diversity for the species in the region. H. pratti was only detected in three caves, the four previously distributed caves and the remaining 41 previously undistributed caves were not observed. An analysis based on microsatellite markers revealed a few key points. 1) The average number of observed alleles (Na) of H. pratti in the region was 3.94, and the average observed heterozygosity (Ho) was 0.5293. All three populations were in Hardy-Weinberg equilibrium. 2) Intra-population was the predominant genetic variation of the H. pratti population. 3) H. pratti in the region experienced a bottleneck effect. We found that the three caves where H. pratti are currently distributed face varying degrees of human interference and the populations are threatened. Management strategies, including appropriate countermeasures to reduce human interference in caves, are urgently needed.

zoology↗

bioRxiv2023

Human co-ripples facilitate neuronal interactions between cortical locations.

Synchronous bursts of high frequency oscillations ( ripples) are hypothesized to contribute to binding by facilitating integration of neuronal firing across cortical locations. We tested this hypothesis using local field-potentials and single-unit firing from four 96-channel microelectrode arrays in supragranular cortex of 3 patients. Neurons in co-rippling locations showed increased short-latency co-firing, prediction of each-others firing, and co-participation in neural assemblies. Effects were similar for putative pyramidal and interneurons, during NREM sleep and waking, in temporal and Rolandic cortices, and at distances up to 16mm. Increased co-prediction during co-ripples was maintained when firing-rate changes were equated, and were strongly modulated by ripple phase. Co-ripple enhanced prediction is reciprocal, synergistic with local upstates, and further enhanced when multiple sites co-ripple. Together, these results support the hypothesis that trans-cortical co-ripples increase the integration of neuronal firing of neurons in different cortical locations, and do so in part through phase-modulation rather than unstructured activation.

neuroscience↗

bioRxiv2022

Measurement of Cervical Neuronal Activity during Stress Challenge Using Novel Flexible Adhesive Surface Electrodes

This study introduces a flexible, adhesive-integrated electrode array that was developed to enable non-invasive monitoring of cervical nerve activity. The device uses silver-silver chloride as the electrode material of choice and combines it with a novel electrode array consisting of a customized biopotential data acquisition unit and integrated graphical user interface (GUI) for visualization of real-time monitoring. Preliminary testing demonstrated this novel electrode design can achieve a high signal to noise ratio during cervical neural recordings. To demonstrate the capability of the surface electrodes to detect changes in cervical neuronal activity, the cold-pressor test (CPT) and a timed respiratory challenge were employed as stressors to the autonomic nervous system. This sensor system recording, a new novel technique, was termed Cervical Electroneurography (CEN). By applying a custom spike sorting algorithm to the novel electrode measurements, neural activity was classified in two ways: 1) pre-to-post CPT, and 2) during a timed respiratory challenge. Unique to this work: 1) rostral to caudal channel position-specific (cephalad to caudal) firing patterns and 2) cross challenge biotype-specific change in average CEN firing, were observed with both CPT and the timed respiratory challenge. Future work is planned to develop an ambulatory CEN recording device that could provide immediate notification of autonomic nervous system activity changes that might indicate autonomic dysregulation in healthy subjects and clinical disease states.

physiology↗

bioRxiv2022

Extracellular matrix deposition precedes muscle-tendon integration during murine forelimb morphogenesis

The development of a functional vertebrate musculoskeletal system requires the combination of contractile muscle and extracellular matrix (ECM)-rich tendons that transmit muscle-generated force to bone. Despite the different embryologic origins, muscle and tendon integrate at the myotendinous junction (MTJ) to seamlessly connect cells and ECM across this interface. While the cell-cell signaling factors that direct development have received considerable attention, how and when the ECM linking these tissues is deposited remains unknown. To address this gap, we analyzed the 3D distribution of different ECM and the influence of skeletal muscle in forelimbs from wildype (WT) and muscle-less Pax3Cre/Cre mice. At E11.5, prior to MTJ integration, an aligned ECM was present at the presumptive insertion of the long triceps into the WT ulna. Mechanically robust tendon-like and muscle compartmentalization structures, positive for type I collagen, type V collagen, and fibrillin-2, still formed when muscle was knocked out. However, MTJ-specific ECM was not observed when muscle was absent. Our results show that an ECM-based template forms independent of muscle, but muscle is needed for the proper assembly of ECM at the MTJ. Summary statementAn aligned ECM template connects tendon and muscle during limb development, independent of muscle progenitor migration into the limb; however, the assembly of MTJ-specific ECM requires the presence of muscle.

developmental biology↗

bioRxiv2021

Quantitative imaging in whole-mount zebrafish embryos traces morphogen gradient maintenance and noise propagation in BMP signaling

Dorsoventral (DV) embryonic patterning relies on precisely controlled interpretation of morphogen signaling. In all vertebrates, DV axis specification is informed by gradients of Bone Morphogenetic Proteins (BMPs). We developed a 3D single-molecule mRNA quantification method in whole-mount zebrafish to quantify the inputs and outputs in this pathway. In combination with 3D computational modeling of zebrafish embryo development, data from this method revealed that Sizzled (Szl), shaped by BMP and Nodal signaling, kept a consistent inhibition level with Chordin (Chd) to maintain the BMP morphogen gradient. Intriguingly, BMP morphogen intrinsic expression is highly noisy at the ventral marginal layer in early zebrafish gastrula, where the gradient for DV patterning is established, which implies an unexpected role for noise in gradient shaping.

systems biology↗

bioRxiv2021

Detection of Small Fiber Neuronal Activity with Optically Pumped Magnetometers

Electrodiagnosis is routinely integrated into clinical neurophysiology practice for peripheral nerve disease diagnoses such as neuropathy, demyelinating disorders, nerve entrapment/impingement, plexopathy or radiculopathy. Measured with conventional surface electrodes, the propagation of peripheral nerve action potentials along a nerve is the result of ionic current flow which, according to Amperes Law, generates a small magnetic field that is also detected as an "action current" by magnetometers such as superconducting quantum interference device (SQUID) Magnetoencephalography (MEG) systems. Optically pumped magnetometers (OPMs) are an emerging class of quantum magnetic sensors with a demonstrated sensitivity at the 1 fT/{surd}Hz level, capable of cortical action current detection. But OPMs were ostensibly constrained to low bandwidth therefore precluding their use in peripheral nerve electrodiagnosis. With careful OPM bandwidth characterization, we hypothesized OPMs may also detect compound action current signatures consistent with both Sensory Nerve Action Potential (SNAP) and the Hoffmann Reflex (H-Reflex). In as much, our work confirms OPMs enabled with expanded bandwidth can detect the magnetic signature of both the SNAP and H-Reflex. Taken together, OPMs now show potential as an emerging electrodiagnostic tool.

neuroscience↗

bioRxiv2020

MACMIC Reveals Dual Role of CTCF in Epigenetic Regulation of Cell Identity Genes

Numerous studies of relationship between epigenomic features have focused on their strong correlation across the genome, likely because such relationship can be easily identified by many established methods for correlation analysis. However, two features with little correlation may still colocalize at many genomic sites to implement important functions. There is no bioinformatic tool for researchers to specifically identify such feature pair. Here, we develop a method to identify feature pair in which two features have maximal colocalization but minimal correlation (MACMIC) across the genome. By MACMIC analysis of 3,385 feature pairs in 15 cell types, we reveal a dual role of CTCF in epigenetic regulation of cell identity genes. Although super-enhancers are associated with activation of target genes, only a subset of super-enhancers colocalized with CTCF regulate cell identity genes. At super-enhancers colocalized with CTCF, the CTCF is required for the active marker H3K27ac in cell type requiring the activation, and also required for the repressive marker H3K27me3 in other cell types requiring the repression. Our work demonstrates the biological utility of the MACMIC analysis and reveals a key role for CTCF in epigenetic regulation of cell identity.

bioinformatics↗