Biology subjects

Pan, R.

Publications and source records attributed to Pan, R..

At least 19 recordsLinked to original sources

bioRxiv2026

A single-domain antibody targets aggregation-prone region of α-synuclein to reduce synucleinopathy, rescue neurodegeneration and improve function

Synucleinopathies are a group of neurodegenerative disorders characterized by the accumulation of aggregated -synuclein (-syn), including Parkinsons disease, Dementia with Lewy Bodies, and Multiple System Atrophy. These diseases are marked by locomotor and non-motor impairments, as well as mitochondrial dysfunction and the loss of dopaminergic (DA) neurons. We have developed several anti--syn single-domain antibodies (sdAbs) and demonstrated the diagnostic imaging potential of two of them and the acute therapeutic benefit of one in clearing -syn in a mouse model. However, whether these sdAbs can suppress -syn-mediated neuronal loss and locomotor impairment in vivo remains unclear. We evaluated the therapeutic potential of five anti--syn sdAbs to clear pathological -syn in mouse neuronal culture and then demonstrated their in vivo efficacy in a Drosophila model of synucleinopathy. The sdAbs differed in their efficacy to lower levels of phospho-serine 129 -syn, prevent loss of DA neurons, alleviate mitochondrial dysfunction, improve motor function, and prolong survival in synucleinopathy flies. The most effective sdAb, 2H1, has not been reported before. It binds strongly to the aggregation prone region of -syn and robustly improves all these disease parameters. Additionally, that sdAb is associated with -syn in the fly neurons, as shown through proximity dependent turboID biotinylation assays. The sdAb-turboID also biotinylated -syn-associated proteins involved in synapse/vesicle trafficking pathways, pinpointing the location of their intracellular interaction. Our findings provide an insight into the therapeutic mechanism of action of these sdAbs and strongly support their clinical development.

neuroscience↗

bioRxiv2026

Tau pSer396 and pSer404 Define Distinct Epitope Regions Linked to Different Antibody Functions

Hyperphosphorylated tau is a central pathological feature of Alzheimers disease and related tauopathies, and antibodies targeting the pSer396/pSer404 epitope region represent a promising therapeutic strategy. However, direct comparisons of pSer396- and pSer404-selective antibodies and the impact of humanization on their functional properties remain limited. We generated two new monoclonal antibodies (mAbs), 9E (pSer404-specific) and G10 (pSer396-specific), and evaluated them alongside 4E6 (pSer404) and PHF-1 (pSer396) in murine and partially humanized chimeric formats. Antibodies were assessed in mixed cortical cultures using extracellular (PHF + Ab) and intracellular (PHF [->] Ab) paradigms. Efficacy in preventing tau-induced toxicity and seeding differed substantially among antibodies and was variably altered by chimerization, despite identical variable regions. Antibodies targeting pSer404 were more effective than those targeting pSer396, and antibodies that preferentially bound soluble pathological tau species in competition ELISA were consistently more efficacious, whereas neuronal uptake was comparable across variants. To define structural determinants of phospho-epitope recognition, we determined the crystal structures of the Fab regions of 9E, G10, and PHF-1, and additionally solved the co-crystal structure of Fab PHF-1 in complex with a pSer396 tau peptide at 2.55 [A] resolution. The PHF-1 complex reveals a heavy-chain-dominant binding mode in which pSer396 is anchored within an electropositive pocket and Tyr394 adopts a flipped conformation that stabilizes a {beta}-strand-like motif, consistent with a phosphorylation-dependent conformational switch. These findings demonstrate that epitope selectivity, aggregate preference, structural binding mode, and Fc context collectively govern antibody efficacy, and that humanization can substantially alter therapeutic properties.

neuroscience↗

bioRxiv2026

A basophil-specific GPCR mediates the immune response to helminth infection

Basophils are rare innate immune cells that contribute to type 2 immunity in allergy and parasitic helminth infection, yet the receptors governing their activation remain poorly defined. Here, we identify the G protein-coupled receptor (GPCR) Mrgpra6 as a basophil-specific gene that mediates the host response to helminth infection. Mrgpra6 expression is restricted to basophils in blood and lung and is uniformly expressed across the population. Knockout of Mrgpra6 impairs early innate immunity to invading helminths, resulting in increased larval burden, elevated mortality, and disrupted parasite progression within the host. Transcriptional profiling of FACS-sorted basophils during helminth infection reveals how Mrgpra6 shapes the basophil transcriptional landscape needed for effective host defense. Together, these findings identify Mrgpra6 as a functional regulator of basophil-mediated anti-helminth immunity.

immunology↗

bioRxiv2025

Single-Domain Antibody-Based Autophagosome-Targeting Chimera for Tau Clearance and Motor Function Restoration in Tauopathies

Tauopathies are neurodegenerative diseases characterized by pathological tau accumulation, leading to motor and neuropsychiatric symptoms. Effective tau-targeting therapies remain a major challenge. Here, we present 1D9-LIR{Delta}TP53INP2, a single-domain antibody (sdAb)-based protein degrader that facilitates tau clearance via the autophagy-lysosomal pathway. This engineered molecule combines the anti-tau sdAb 1D9 with an LC3-interacting region (LIR{Delta}TP53INP2) to promote autophagosomal recruitment, mimicking autophagy receptors by simultaneously binding tau and LC3. In frontotemporal dementia (FTD) patient-derived neurons and JNPL3 tauopathy mice, both harboring the P301L tau mutation, 1D9-LIR{Delta}TP53INP2 significantly reduced tau levels and improved motor function in mice. These findings underscore the therapeutic potential of sdAb-based protein degraders for tauopathies. Given the challenges of brain delivery for conventional antibodies, sdAbs with enhanced brain penetration and efficacy offer a promising strategy for treatment of neurodegenerative diseases.

neuroscience↗

bioRxiv2025

A parallelly distributed microscope and software system for scalable high-throughput multispectral 3D imaging

Recent advances in high throughput optical microscopy have achieved whole-organism scale imaging at diffraction-limited resolutions. Current microscopes, however, require making compromises between achieving the optimal resolution, imaging depth, multispectral capability, and data throughput due to limitations in optical design and data stream handling. We have created a parallel-line scanning confocal microscope (plSCM) which provides 1.1 Gigavoxels/second high speed imaging while achieving an optical resolution of [~]180 x 220 x 650 nm with 2 millimeters of imaging depth in 3 simultaneous spectral channels. To handle such a massive imaging data stream, we have engineered a scalable network-distributed image acquisition/processing framework (SNDiF), which allows continuous capture, real-time processing, and cluster storage of petabyte-scale single image datasets in days. Together, our parallelly distributed microscope and software system presents a general solution for large-scale high-throughput and high-resolution multicolor imaging which can operate for days at a time.

bioengineering↗

bioRxiv2025

Inferring non-coding RNA regulatory network from transcriptomic data and curated databases

Non-coding RNAs (ncRNAs) have played an indispensable role in regulating gene expression and cellular processes. However, the complex regulatory circuits between ncRNAs and coding genes remain understudied. Major challenges for inferring ncRNA regulatory network (NRN) on a transcriptome-wide scale include high dimensionality of both ncRNA and coding gene expression data, their context-dependent interaction, and lack of validation. We hereby propose a comprehensive analytical framework, namely Construction and Analysis of non-coding RNA regulatory NETwork (CAR-NET), specifically designed to infer NRN from transcriptomic data and curated databases on experimentally validated ncRNA-gene interactions and disease-related ncRNAs. At its core is a novel computationally efficient Bayesian network structure learning method that leverages the semi-bipartite graph structure of NRN for dimension reduction, edge orientation and global network search. We also developed and applied algorithms to perform differential network, subnetwork and pathway analysis of the inferred network. We showed the strength of CAR-NET in simulations and three case studies, encompassing lncRNA regulation in brain development, miRNA differential regulation in renal cell carcinoma and regulation by different classes of ncRNAs in different cell lines using bulk or single cell RNA-seq data. An R-shiny app (https://github.com/kehongjie/CAR-NET) was developed to implement the method(s) with interactive GUI.

bioinformatics↗

bioRxiv2024

Non-destructive in situ monitoring of structural changes of 3D tumor spheroids during the formation, migration, and fusion process

For traditional laboratory microscopy observation, the multi-dimensional, real-time, in situ observation of three-dimensional (3D) tumor spheroids has always been the pain point in cell spheroid observation. In this study, we designed a side-view observation petri dish/device that reflects light, enabling in situ observation of the 3D morphology of cell spheroids using conventional inverted laboratory microscopes. We used a 3D-printed handle and frame to support a first surface mirror, positioning the device within a cell culture petri dish to image cell spheroid samples. The imaging conditions, such as the distance between the mirror and the 3D spheroids, the light source, and the impact of the culture medium, were systematically studied to validate the in-situ side-view observation. The results proved that placing the surface mirror adjacent to the spheroids enables non-destructive in situ real-time tracking of tumor spheroid formation, migration, and fusion dynamics. The correlation between spheroid thickness and dark core appearance under light microscopy and the therapeutic effects of chemotherapy doxorubicin and Natural Killer cells on spheroidss 3D structure was investigated.

cancer biology↗

bioRxiv2024

Mapping individual differences in intermodal coupling in neurodevelopment

Within-individual coupling between measures of brain structure and function evolves in development and may underlie differential risk for neuropsychiatric disorders. Despite increasing interest in the development of structure-function relationships, rigorous methods to quantify and test individual differences in coupling remain nascent. In this article, we explore and address gaps in approaches for testing and spatially localizing individual differences in intermodal coupling, including a new method, called CEIDR. CEIDR controls false positives in individual differences in intermodal correlations that arise from mean and variance heterogeneity and improves statistical power by adopting adaptive cluster enhancement. Through a comparison across different approaches to testing individual differences in intermodal coupling, we delineate subtle differences in the hypotheses they test, which may ultimately lead researchers to arrive at different results. Finally, we illustrate these differences in two applications to brain development using data from the Philadelphia Neurodevelopmental Cohort.

neuroscience↗

bioRxiv2024

Single-Domain Antibody-Based Protein Degrader for Synucleinopathies

Synucleinopathies are a group of neurodegenerative diseases characterized by the accumulation of -synuclein (-syn) in the brain, leading to motor and neuropsychiatric symptoms. Currently, there are no known cures for synucleinopathies, and treatments mainly focus on symptom management. In this study, we developed a single-domain antibody (sdAb)-based protein degrader with features designed to enhance proteasomal degradation of -syn. This sdAb derivative targets both -syn and Cereblon (CRBN), a substrate-receptor for the E3-ubiquitin ligase CRL4CRBN, and thereby induces -syn ubiquitination and proteasomal degradation. Our results indicate that this therapeutic candidate enhances proteasomal degradation of -syn, in addition to the endogenous lysosomal degradation machinery. By promoting proteasomal degradation of -syn, we improved clearance of -syn in primary culture and mouse models of synucleinopathy. These findings indicate that our sdAb-based protein degrader is a promising therapeutic candidate for synucleinopathies. Considering that only a small percentage of antibodies enter the brain, more potent sdAbs with greater brain entry than whole antibodies could enhance clinical benefits of antibody-based therapies.

neuroscience↗

bioRxiv2023

Cell-autonomous targeting of arabinogalactan by host immune factors inhibits mycobacterial growth

Deeper understanding of the crosstalk between host cells and Mycobacterium tuberculosis (Mtb) provides crucial guidelines for the rational design of novel intervention strategies against tuberculosis (TB). Mycobacteria possess a unique complex cell wall with arabinogalactan (AG) as critical component. AG has been identified as a virulence factor of Mtb which is recognized by host galectin-9. Here we demonstrate that galectin-9 directly inhibited mycobacterial growth through AG-binding property of carbohydrate-recognition domain 2. Furthermore, IgG antibodies with AG specificity were detected in serum of TB patients. Based on the interaction between galectin-9 and AG, we developed monoclonal antibody (mAb) screening assay and identified AG-specific mAbs which profoundly inhibit Mtb growth. Mechanistically, proteomic profiling and morphological characterizations revealed that AG-specific mAbs regulate AG biosynthesis, thereby inducing cell wall swelling. Thus, direct AG-binding by galectin-9 or antibodies contributes to protection against TB. Our findings pave the way for the rational design of novel immunotherapeutic strategies for TB control.

immunology↗

bioRxiv2023

BRD4 directs mitotic cell division by inhibiting DNA damage

BRD4 binds to acetylated histones to regulate transcription and drive cancer cell proliferation. However, the role of BRD4 in normal cell growth remains to be elucidated. Here we investigated the question by using mouse embryonic fibroblasts with conditional Brd4 knockout (KO). We found that Brd4KO cells grow more slowly than wild type cells: they do not complete replication, fail to achieve mitosis, and exhibit extensive DNA damage throughout all cell cycle stages. BRD4 was required for expression of more than 450 cell cycle genes including genes encoding core histones and centromere/kinetochore proteins that are critical for genome replication and chromosomal segregation. Moreover, we show that many genes controlling R-loop formation and DNA damage response (DDR) require BRD4 for expression. Finally, BRD4 constitutively occupied genes controlling R-loop, DDR and cell cycle progression. We suggest that BRD4 epigenetically marks those genes and serves as a master regulator of normal cell growth.

cell biology↗

bioRxiv2023

IRF8 configures enhancer landscape in postnatal microglia and directs microglia specific transcriptional programs

Microglia are innate immune cells in the brain. Transcription factor IRF8 is highly expressed in microglia. However, its role in postnatal microglia development is unknown. We demonstrate that IRF8 binds stepwise to enhancer regions of postnatal microglia along with Sall1 and PU.1, reaching a maximum after day 14. IRF8 binding correlated with a stepwise increase in chromatin accessibility, which preceded the initiation of microglia-specific transcriptome. Constitutive and postnatal Irf8 deletion led to a loss of microglia identity and gain of disease-associated microglia-like genes. Combined analysis of scRNA-seq and scATAC-seq revealed a correlation between chromatin accessibility and transcriptome at a single-cell level. IRF8 was also required for microglia-specific DNA methylation patterns. Lastly, in the 5xFAD model, constitutive and postnatal Irf8 deletion reduced the interaction of microglia with A{beta} plaques and the size of plaques, lessening neuronal loss. Together, IRF8 sets the epigenetic landscape, which is required for postnatal microglia gene expression.

immunology↗

bioRxiv2023

Evolution of the gene regulatory network of body axis by enhancer hijacking in amphioxus

A central goal of evolutionary developmental biology is to decipher the evolutionary pattern of gene regulatory networks (GRNs) that control morphogenesis during embryonic development, and the underlying molecular mechanism of the GRNs evolution. The Nodal signaling that governs the body axes of deuterostomes exhibits a conserved GRN orchestrated principally by Nodal, Gdf1/3 and Lefty. Despite that the GRN retains a conserved function in deuterostomes, the expression patterns of Gdf1/3 and Nodal are derived in cephalochordate amphioxus, implying a specific rewiring of the GRN in this lineage. Here we examined the regulatory mechanism and evolution of this GRN in amphioxus by functional genetic manipulations. We found that while the amphioxus Gdf1/3 orthologue shows nearly no expression during embryogenesis, its duplicate Gdf1/3-like linking to Lefty is zygotically expressed in a similar pattern as Lefty. Mutant and transgenic analyses revealed that Gdf1/3 is no longer crucial for amphioxus axial development. Instead, Gdf1/3-like assumes this responsibility, likely through hijacking Lefty enhancers. We also showed that amphioxus Nodal has become an indispensable maternal factor to compensate for the loss of maternal Gdf1/3 expression. We therefore demonstrated a case that the evolution of an ancestral GRN could be triggered by enhancer hijacking events. This pivotal event has allowed the emergence of a new GRN in the extant amphioxus, presumably evolving through a stepwise process. The co-expression of Gdf1/3-like and Lefty achieved by shared regulatory region may have provided developmental robustness during body axis formation in extant amphioxus, which provides a selection-based hypothesis for the phenomena called developmental system drift.

evolutionary biology↗

bioRxiv2023

Spatial-extent inference for testing variance components in reliability and heritability studies

Clusterwise inference is a popular approach in neuroimaging to increase sensitivity, but most existing methods are currently restricted to the General Linear Model (GLM) for testing mean parameters. Statistical methods for testing variance components, which are critical in neuroimaging studies that involve estimation of narrow-sense heritability or test-retest reliability, are underdeveloped due to methodological and computational challenges, which would potentially lead to low power. We propose a fast and powerful test for variance components called CLEAN-V (CLEAN for testing Variance components). CLEAN-V models the global spatial dependence structure of imaging data and computes a locally powerful variance component test statistic by data-adaptively pooling neighborhood information. Correction for multiple comparisons is achieved by permutations to control family-wise error rate (FWER). Through analysis of task-fMRI data from the Human Connectome Project across five tasks and comprehensive data-driven simulations, we show that CLEAN-V outperforms existing methods in detecting test-retest reliability and narrow-sense heritability with significantly improved power, with the detected areas aligning with activation maps. The computational efficiency of CLEAN-V also speaks of its practical utility, and it is available as an R package.

neuroscience↗

bioRxiv2022

An e-cigarette aerosol generation, animal exposure and toxicants quantification system to characterize in vivo nicotine kinetics in arterial and venous blood

The increasing e-cigarette use worldwide presents an urgent need to characterize their nicotine delivery property, brain stimulation and potential long-term health effects. We constructed an end-to-end system enabling combustible-cigarette (c-cigarette) and e-cigarette aerosol generation, animal exposure, and effect assessment. The system consists of (1) a 10-channel aerosol generator resembling human smoking/vaping scenarios, (2) nose-only and whole-body exposure chambers suitable for long- or short-duration studies, (3) a lab protocol for animal exposure and collecting arterial and venous blood <1 minute after the exposure, and (4) chromatograph and mass spectrometry to quantify nicotine concentrations in aerosol and biospecimens. We applied the system in a proof-of-principle study characterizing in vivo nicotine delivery after e-cigarette aerosol inhalation. Groups of Sprague-Dawley rats were exposed to e-cigarette aerosols for 1, 2 and 4 minutes, respectively. Arterial and venous blood samples were collected immediately after the exposure. We also directly compared nose-only and whole-body exposure approaches. After nose-only e-cigarette aerosol exposure, the nicotine concentration in arterial blood was substantially higher (11.32 ng/mL in average) than in veins. Similar arterio-venous concentration difference was observed in whole-body exposure experiments. In summary, we described a complete system ideal for e- and c-cigarette in vivo nicotine kinetics and long-term health research. Our findings highlight arterial blood as the suitable bio-specimen for e-cigarette nicotine delivery studies. HighlightO_LIWe constructed a combustible- and e-cigarette aerosol generation - exposure - effect assessment system resembling real world human smoking/vaping scenarios. C_LIO_LIProof-of-principle study characterized in vivo nicotine delivery from e-cigarette aerosol to arterial and venous blood at high temporal resolution. C_LIO_LIAfter exposure, the nicotine concentration was substantially higher (11.32 ng/mL) in arterial blood than in veins. C_LIO_LIOur results suggest arterial blood as the suitable bio-specimen to study nicotine delivery and brain stimulation. C_LI

pharmacology and toxicology↗

bioRxiv2022

DNAscope: High accuracy small variant calling using machine learning

We present DNAscope, an accurate and efficient germline small-variant caller. DNAscope combines the robust and well-established preprocessing and assembly mathematics of the GATKs HaplotypeCaller with a machine-learned genotyping model. Benchmarks of DNAscope and DNAseq (Sentieons GATK-matching germline variant calling pipeline) demonstrate that DNAscope achieves superior SNP and insertion/deletion accuracy with reduced computational cost.

bioinformatics↗

bioRxiv2022

PanGu Drug Model: Learn a Molecule Like a Human

Recent achievements in large-scale pre-trained models like GPT-3 and PanGu- showed amazing performances in many downstream tasks, which makes AI friendlier toward industrial users. Deep learning has been recognized as the most promising technology for pharmaceuticals, a powerful molecule pre-trained model could save researchers tons of time. In chemistry classes, the students learn two molecule representations, the molecular formula and the structure formula, and learn to translate them from one way to the other. Inspired by this, we developed a novel deep learning architecture using a graph-to-sequence asymmetric conditional variational autoencoders, called PanGu Drug Model, which can appropriately characterize the molecule from both representations and improve the performance of downstream drug discovery tasks. After pretrained with 1.7 billion small molecules, our proposed model achieved the state-of-the-art results in 20 drug discovery tasks, such as molecule property prediction (predict ADMET properties, compound-target interactions, drug-drug interactions and chemical reaction productivity), molecule generation and molecule optimization. A new drug screening library of 100 million drug-like small molecules with 99.68% novelty was generated by PanGu molecule generator, which could efficiently produce novel compounds with similar physiochemical properties to given distribution, this library could be used to supplement existing compound databases. In addition, PanGu molecule optimizer could optimize the chemical structures of starting molecule with improved molecular property of interest. An automatic multi-objective optimization web application implemented by PanGu Drug Model is available at http://www.pangu-drug.com/.

biochemistry↗

bioRxiv2022

Engineering bacterial blight-resistant plants through CRISPR/Cas9-targeted editing of the MeSWEET10a promoter in cassava

Cassava starch is a widely used raw materials for industrial production. However, cassava bacterial blight (CBB) caused by Xanthomonas axonopodis pv. manihotis (Xam) results in severe yield losses and is the most destructive bacterial disease in all worldwide cassava planting regions. This study showed that editing of the promoter in the disease-susceptibility gene MeSWEET10a of SC8 cassava confers resistance to CBB. All mutated cassava lines had normal morphological and yield-related traits as the wild type. The results lay a research foundation for breeding cassava resistant to bacterial blight. HighlightsMeSWEET10a gene in SC8 cultivar is hijacked by TALE20 from Xam11 strain. Editing of the MeSWEET10a promoter in SC8 cultivar confers resistance to CBB. All the mutated cassava lines had similar yield-related traits compared to wild-type.

plant biology↗