Biology subjects

Cao, J.

Publications and source records attributed to Cao, J..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

Structural basis for constitutive activation of oncogenic gp130 mutants in human inflammatory hepatocellular adenomas

Inflammatory hepatocellular adenomas (IHCAs) are benign liver tumors primarily driven by somatic mutations in the gp130 receptor, resulting in its constitutive activation. To uncover the structural mechanisms underlying this activation, we employed cryo-electron microscopy and mutagenesis to study five representative IHCAs-associated gp130 mutants and two engineered mutants with ligand-independent activity. Most mutants formed "X"-shaped dimers through a 3D domain-swapping mechanism, characterized by large juxtamembrane separations and reliant on D1 domain-mediated "tip-to-tip" interactions for dimer clustering and activation. A rare mutant displayed a distinct "Y"-shaped dimer conformation, with closely aligned juxtamembrane domains enabling direct activation independent of dimer clustering. These findings highlight the diversity of gp130 mutant activation mechanisms and provide a foundation for developing therapeutic strategies targeting aberrant gp130 signaling in IHCAs.

biochemistry↗

bioRxiv2025

Zebrafish otic vesicle and mouse epididymis as model systems for studying columnar epithelial cell division

Epithelial cell division maintains tissue architecture through coordinated nuclear migration, cell shape changes, and spindle orientation. In columnar epithelia, interkinetic nuclear migration (INM) involves apical nuclear translocation in G2 phase and basal return post-mitosis, yet its regulation remains incompletely understood in vertebrates, in part due to limited in vivo live-imaging models. In this methodological study, we adapted the zebrafish embryonic otic vesicle as an in vivo model to investigate cell division dynamics in simple columnar epithelium using high-resolution live imaging and genetic tools. We demonstrated that apical INM initiates in mid-to-late G2 and is driven by dynein, not myosin II. Mitotic rounding is achieved via actomyosin-mediated basolateral constriction while maintaining basal attachment. Inhibiting myosin II impairs rounding and planar division, causing apical retention of daughter cells, suggesting planar division ensures proper integration. We additionally analyzed the mouse epididymal epithelium, a simple columnar epithelial tissue, to allow cross-species comparison of nuclear migration dynamics. Together, these optimized in vivo vertebrate models uncover conserved and tissue-specific mechanisms underlying epithelial organization and function, and importantly, provide tools for deeper mechanistic dissection in the future.

cell biology↗

bioRxiv2025

Embryonic Atrial Injury Programs an Anti-Fibrotic Atrial Fibrillation Substrate

Atrial myopathy is associated with atrial fibrillation in adults, yet the role of embryonic atrial injury in predisposing individuals to atrial fibrillation remains unclear. Using an inducible, cardiomyocyte-specific diphtheria toxin A-mediated injury mouse model, we found that injured embryonic atria exhibited upregulation of apoptosis and TNF signaling pathways, alongside downregulation of calcium handling and atrial fibrillation-associated genes. Single-cell RNA sequencing revealed two distinct cardiomyocyte populations with differential injury responses. Importantly, although myofibroblasts increased in the fibroblast population, these cells displayed an extracellular matrix gene expression profile distinct from the pro-fibrotic phenotype typically observed in fibrillating adult atria and showed no evidence of fibrosis. Concurrently, monocytes/macrophages were activated and showed upregulated expression of the anti-fibrotic factor GPNMB. These findings suggest that adaptive tissue remodeling is counterintuitively prevented from progressing to fibrosis, potentially through GPNMB-mediated mechanisms. This study elucidates molecular pathways linking developmental atrial injury to myopathy and proposes novel strategies to attenuate atrial fibrillation progression.

pathology↗

bioRxiv2025

Decoder-FFPE-seq enables sensitive, genome-wide spatial transcriptomics of archival tissues at single-cell resolution

Formalin-fixed paraffin-embedded (FFPE) tissue-compatible spatial transcriptomics (ST) is transforming biology and histopathology, but achieving high sensitivity, genome-wide coverage, single-cell resolution and researcher-accessibility remains challenging. Here, we present a dendrimeric DNA coordinate-barcoding design for FFPE-adapted spatial RNA sequencing (Decoder-FFPE-seq), by integrating RNA-to-DNA probe conversion with blotting-assisted barcoding. Decoder-FFPE-seq provides single-cell resolution with a 2.54-4.25-fold sensitivity increase in mouse brain transcriptomic profiling over similar assays with an available protocol, successfully mapping rare genes and sparse cell types. It robustly resolves tumor microenvironments (TMEs) in clinical archives preserved 6-20 years. With expanded probe libraries, Decoder-FFPE-seq captures long non-coding and viral RNAs, revealing spatial co-expression networks with mRNAs in oral squamous cell carcinomas (OSCCs). In an OSCC immunotherapy cohort, it delineates TME heterogeneity across pathology response groups and tracks spatial cellular dynamics pre- and post-immuno-chemotherapy. Decoder-FFPE-seq establishes a robust, high-resolution framework for sensitive ST profiling in fundamental and translational studies of archival tissues.

genomics↗

bioRxiv2025

Multi-omics analysis reveals genetic regulatory mechanismsof muscle traits in wild and domesticated Bactrian camels

Bactrian camels, large domesticated animals adapted to extreme environments, have evolved under natural and artificial selection to suit their living conditions and meet human needs. In arid and semi-arid regions, they provide valuable resources through economically important traits such as meat, milk, and wool production, contributing significantly to both economy and ecology. However, research on gene function, adaptive evolution, and molecular mechanisms underlying complex traits in Bactrian camels remains limited, constraining the full utilization of their breeding potential and conservation efforts.To address this gap, we performed whole-genome resequencing (WGS) on 35 domesticated Bactrian camels from the Chinese Sunite local population and integrated this data with publicly available WGS data from 112 Asian Bactrian camels. Our comprehensive analysis revealed significant genetic differences and genomic features between wild and domesticated Bactrian camels. We further integrated 27 epigenomic (ATAC-seq) and transcriptomic (RNA-seq) datasets from eight biologically important tissues of four Bactrian camels, along with skeletal muscle scRNA-seq and scATAC data. This allowed us to identify tissue-specific genes, regulatory elements, and genes under positive selection.Our integrated analysis identified target genes and their upstream regulatory elements, including potential regulatory transcription factors, with a focus on positively selected regions in Bactrian camel muscle tissue. We constructed regulatory pathways from key SNPs to regulatory elements to genes, and for the first time, identified regulatory elements in major tissues of the Bactrian camel genome. Our results also summarized the single-cell transcriptional landscape of Bactrian camel skeletal muscle, revealing cellular heterogeneity and genetic expression patterns across two developmental stages.This comprehensive regulatory element map provides a valuable resource for genetic and genomic studies in Bactrian camels. It reveals key regulatory networks, important genes, and regulatory elements influencing muscle development, as well as potentially significant functional variants affecting meat production traits. These findings will provide crucial data support for future genetic improvement and breeding efforts in Bactrian camels.

genetics↗

bioRxiv2025

The Pdgfd-Pdgfrb axis orchestrates tumor-nerve crosstalk in pancreatic cancer

Nerves are an integral component of the tumor microenvironment, contributing to cancer progression, metastasis, morbidity, and mortality. In pancreatic ductal adenocarcinoma (PDAC), worse clinical outcomes are associated with perineural invasion (PNI), a process by which cancer cells surround and invade nerves. Here, we employed whole-transcriptome and single-cell spatial transcriptomics to identify candidate tumor-nerve interactions that promote PNI. We discovered that Pdgfd signaling promotes key features of nerve invasion. Mechanistically, Pdgfd stimulated cancer cell invasiveness, neurite outgrowth, and direct physical engagement with glia. Pharmacological blockade of this axis reduced each of these processes in vitro as well as PNI in vivo. Thus, Pdgfd-Pdgfrb signaling mediates PNI by coordinating multifaceted cancer-neuron-glia interactions and represents a promising therapeutic strategy aimed at disrupting harmful cancer-nerve crosstalk.

cancer biology↗

bioRxiv2025

Droplet-based microfluidics platform for investigation of protoplast development of three exemplary plant species

Microfluidic technologies offer powerful tools for miniaturized and highly controlled biological experiments, yet their application in plant research remains underexploited. In this study, we present a droplet-based microfluidic platform tailored for the encapsulation and cultivation of plant protoplasts, enabling long-term observation of cell development at nearly single-cell resolution. Protoplasts isolated from leaves of Nicotiana tabacum, Brassica juncea, and Kalanchoe daigremontiana were used to evaluate the platforms suitability across diverse plant species. Our results demonstrate species-dependent responses to microfluidic cultivation, with tobacco protoplasts showing the highest viability. The system permits dynamic tracking of cell fate within individual droplets and supports the quantification of stochastic and concentration-dependent responses to chemical stimuli. Using tobacco protoplasts, we further investigated the effect of low concentrations of cytokinins (BAP) and auxins (NAA) for the early protoplast culture, up to the first division. Low concentrations (20-80 {micro}g{middle dot}L-1) significantly enhanced cell survival and cell growth, while higher doses did not yield additional benefits. This work underscores the potential of droplet-based microfluidics as a high-resolution, low-volume platform for protoplast-based assays and dose-response screening, with applications across diverse plant biotechnology studies.

bioengineering↗

bioRxiv2025

Serotonin shapes the temporal window for associative fear learning

Fear learning is a critical adaptive mechanism that enables the association of an environmental cue (the conditioned stimulus, CS) with a potential threat (the unconditioned stimulus, US), even when these events are temporally discontiguous. When associations are instead formed between unrelated cues and threats, the maladaptive learning process can give rise to neurological disorders. Therefore, maintaining accurate temporal boundaries for forming such associations is crucial for distinguishing real threats from irrelevant stimuli. Despite its importance, the mechanisms defining this precise time window remain poorly understood. Here, we report that serotonin plays a crucial role in modulating the associable interval in trace fear conditioning. Specifically, we show that serotonergic projections from the dorsal raphe nuclei (DRN) to the CA1 region of the ventral hippocampus (vCA1) dynamically regulate the association of discontiguous CS and US. Mechanistically, we demonstrate that sustained serotonin release in the vCA1 maintains the ensemble activity of 5-HT2C receptor-expressing pyramidal neurons, facilitating the association between temporally separated events. In contrast, when the trace interval exceeds the associable window, transient serotonin release fails to drive vCA1 neuronal ensemble activity, thus preventing maladaptive learning. These findings highlight a key role for serotonin in regulating the temporal precision of fear learning and reveal the DRN-vCA1 serotonergic pathway as a potential therapeutic target for related disorders.

neuroscience↗

bioRxiv2025

Model of leaf biomass partitioning coefficient in different leaf ranks of rapeseed (Brassica napus L.) main-stem

Leaf growth is a dynamic process, and understanding the number of main-stem leaves and the distribution of leaf biomass at different development stages can provide a theoretical basis for the development of functional-structural rapeseed model. To address the gap in research on the leaf biomass partitioning coefficient model in the crop growth model, we developed the leaf biomass partitioning coefficient model for the main-stem at different leaf ranks in rapeseed. The outdoor experiment with cultivars, Ningyou 18 (V1, conventional), Ningyou 16 (V2, conventional), and Ningza 19 (V3, hybrid), and designed treatment of cultivars x fertilizer x transplanting density in the 2012-2015 growth season. The model was fitted by calculating the ratio of leaf biomass at different leaf ranks on the main-stem to the total leaf biomass on the main-stem, and normalizing the leaf ranks into (0-1] interval. We parameterized the model variables to elucidate how cultivar and environmental conditions influence the leaf biomass partitioning coefficients for different leaf ranks. The descriptive models were validated with independent experimental data, the correlation (r) of simulation and observation values all greater than 0.9 with significant levels at p < 0.001. The absolute values of the average absolute difference (da) are all less than 0.011 g g-1. The ratio of da to the average observation (dap) for the normalized leaf rank, and leaf biomass partitioning coefficient at seedling stage, and budding and bolting stage, and blooming stage, and mature stage are 13.599%, 11.236%, 11.312%, 3.077%, and 8.661%, respectively. The RMSE values are all less than 0.032 g g-1. The models developed in this paper have good performance and reliability for predicting leaf biomass partitioning coefficient of main-stem at different leaf ranks in rapeseed. This study integrates the leaf morphology model of rapeseed with the rapeseed growth model through biomass partitioning, contributing to the development of the functional-structural rapeseed model.

bioinformatics↗

bioRxiv2025

Germline de novo mutation rate of the highly heterozygous amphioxus genome

Germline de novo mutations (DNMs) are the ultimate source of heritable variation, yet their patterns in highly heterozygous genomes remain poorly understood. Amphioxus, an early-branching chordate with exceptionally high genomic heterozygosity (3.2[~]4.2% in sequenced species), offers a unique model to explore mutational dynamics in such contexts. It is unclear whether high heterozygosity in amphioxus is due to a large effective population size, an increased mutation rate, or both. Here, we perform deep short-read whole genome sequencing of a two-generation pedigree of the amphioxus Branchiostoma floridae comprising two parents and 104 offspring, and develop a framework based on allele-aware parental assemblies as the reference to accurately identify DNMs. We detect 205 high-confidence DNMs, yielding a genome-wide mutation rate of 5.10 x 10-9 per base per generation, which is comparable to that of vertebrates. Combining this estimate with observed nucleotide diversity, we obtain an effective population size of [~]1.9 million, indicating that the elevated heterozygosity mainly results from a large effective population size. We observe a maternal-origin bias when considering all DNMs but a paternal-origin bias for early-occurring ones. Amphioxus harbors a much smaller fraction of CpG>TpG DNMs relative to vertebrates, attributable to its low methylation levels. We also investigate putative post-zygotic mutations in the offspring, revealing an unexpected paternal-origin bias. These suggest some distinct mutational mechanisms in amphioxus. Our study not only provides the first DNM measurement for amphioxus but also offers a generalizable strategy for studying DNMs in highly heterozygous genomes, facilitating mutation rate studies across chordates and other lineages.

genomics↗

bioRxiv2025

Thioredoxin-1 inhibits granulosa cell ferroptosis to rescue ovarian aging through mitophagy-dependent activation of BNIP3L

Ovarian aging is closely associated with a decline in fertility and an increase in reproductive dysfunction. Ovarian granulosa cells (GCs) support oocyte homeostasis and development, yet insight into GC dysfunction during aging is limited. Here, we show that aged GCs of humans and mice have indications of elevated ferroptosis, including increased ferroptosis-related metabolites, lipid peroxidation, and iron accumulation. The ferroptosis inhibitor Ferrostatin-1 reversed ovarian impairment and fertility of aged mice in vivo. We show that the age-related reduction in the expression of TXN (thioredoxin) leads to ferroptosis in human and mouse GCs by blocking BNIP3L-dependent mitophagy. Exogenous activation of TXN could promote mitophagy, thereby clearing excessive ROS and inhibiting ferroptosis. These results suggest that anti-ferroptosis-related treatments may assist in treating aging-related reproductive disorders. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=107 SRC="FIGDIR/small/664497v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@1b97e8forg.highwire.dtl.DTLVardef@123f8d0org.highwire.dtl.DTLVardef@a44c61org.highwire.dtl.DTLVardef@92b32e_HPS_FORMAT_FIGEXP M_FIG C_FIG Key pointsO_LIFerroptosis signatures are upregulated in aged GCs from human and mouse ovaries C_LIO_LITXN is deregulated in aged GCs, leading to mitochondrial and ROS metabolic dysfunction C_LIO_LITXN binds to DNA to regulate autophagy and mitophagy genes, including BNIP3L C_LIO_LIInhibition of ferroptosis can ameliorate GC dysfunction C_LI

cell biology↗

bioRxiv2025

A Set of Novel Far-Red Fluorescent Proteins for Temporal Domain Multiplexing and Super-Resolution Imaging

The diverse spectral and photochemical properties of fluorescent proteins enable a variety of imaging applications in cell biology, ranging from cellular and organelle labeling to super-resolution microscopy and multiplexed live cell imaging. Here, we report a set of novel far-red fluorescent proteins, named mfRFP, mfRFP-A, and mCardinal-A, which are characterized by similar fluorescence spectra with excitation/emission at [~]600/660 nm while exhibiting distinct photobleaching rates. Differences in photostability allowed us to perform per-pixel unmixing of the three far-red FPs imaged simultaneously by employing a recently introduced temporal domain multiplexing approach. We demonstrated the application of the temporal domain multiplexing approach with different combinations of far-red fluorescent proteins possessing nearly identical emission spectra by acquiring BrainBow-like images of cellular populations and distinguishing subcellular structures in mammalian cells using a single imaging channel without applying any hardware modifications to the conventional microscope. Unlike previous temporal domain multiplying approaches employing photophysical properties of fluorescent proteins, the current approach is a wide range of microscopy modalities, including 3D imaging with a spinning disk and point scanning confocal microscopy. The most photostable fluorescent protein in the set, mfRFP, was further benchmarked against spectrally similar FPs and applied for super-resolution imaging of structural proteins in mammalian cells and for neuroimaging of model organisms, including mice, zebrafish, and C. elegans.

biochemistry↗

bioRxiv2025

Organism-wide cellular dynamics and epigenomic remodeling in mammalian aging

To investigate organism-wide cellular alterations and epigenomic dynamics during aging, we constructed a single-cell chromatin accessibility atlas spanning 21 mouse tissues across three age groups and both sexes. We found that around one quarter of 536 organ-specific cell types and 1,828 finer-grained subtypes exhibited significant age-related population shifts. Cellular states from broadly distributed lineages displayed synchronized dynamics with age, indicating systemic signals that coordinate these changes. Molecular analyses identified both intrinsic regulators (chromatin peaks, transcription factor activity) and extrinsic factors (cytokine programs) underlying these shifts. Moreover, [~]40% of aging-associated population dynamics were sex-dependent, with tens of thousands of peaks altered exclusively in one sex. Together, these findings present a comprehensive framework for how aging reshapes the chromatin landscape and cellular composition across diverse tissues.

genomics↗

bioRxiv2025

Lactate metabolism antagonizes antiviral innate immunity bypromoting VDAC1-lactylation-mediated mtDNA release

Foot-and-mouth disease (FMD) is an acute, highly contagious disease caused by the foot-and-mouth disease virus (FMDV). Due to its zoonotic nature, this disease not only significantly impacts livestock productivity but also poses a serious threat to human health. FMDV exhibits distinct host specificity, mainly infecting cloven-hoofed animals such as cattle, sheep, pigs, and deer, while perissodactyl animals like horses and donkeys show natural resistance. Elucidating the interspecies differences in response to FMDV infection is essential for understanding the pathogenesis of the virus. In this study, we report that FMDV infection in horse epithelial cells induces the production of lactate, which modulates the function of VDAC1 protein through lactylation, facilitating the release of mtDNA from mitochondria into the cytoplasm. This process activates the cGAS-STING signaling pathway, which stimulates interferon-{beta} expression and suppresses FMDV replication. Our findings provide new insights into the mechanism behind horse resistance to FMDV and lay a theoretical foundation for further research concerning the differential pathogenesis of FMDV infection across species.

cell biology↗

bioRxiv2025

EnrichSci: Transcript-guided Targeted Cell Enrichment for Scalable Single-Cell RNA Sequencing

Large-scale single-cell atlas efforts have revealed many aging-or disease-associated cell types, yet these populations are often underrepresented in heterogeneous tissues, limiting detailed molecular and dynamic analyses. To address this, we developed EnrichSci--a highly scalable, microfluidics-free platform that combines Hybridization Chain Reaction RNA FISH with combinatorial indexing to profile single -nucleus transcriptomes of targeted cell types with full gene-body coverage. When applied to profile oligodendrocytes in the aging mouse brain, EnrichSci uncovered aging-associated molecular dynamics across distinct oligodendrocyte subtypes, revealing both shared and subtype-specific gene expression changes. Additionally, we identified aging-associated exon-level signatures that are missed by conventional gene-level analyses, highlighting post-transcriptional regulation as a critical dimension of cell-state dynamics in aging. By coupling transcript-guided enrichment with a scalable sequencing workflow, EnrichSci provides a versatile approach to decode dynamic regulatory landscapes in diverse cell types from complex tissues.

genomics↗

bioRxiv2025

Spatiotemporal profiling reveals the impact of caloric restriction on mammalian brain aging

Aging induces functional declines in the mammalian brain, increasing its vulnerability to cognitive impairments and neurodegenerative disorders. Among various interventions to slow the aging process, caloric restriction (CR) has consistently demonstrated the ability to extend lifespan and enhance brain function across different species. Yet the precise molecular and cellular mechanisms by which CR benefits the aging brain remain elusive, especially at region-specific and cell type-specific resolution. In this study, we performed spatiotemporal profiling of mouse brains to elucidate the detailed mechanisms driving the anti-aging effects of CR. Utilizing highly scalable single-nucleus genomics and spatial transcriptomics platforms, EasySci and IRISeq, we profiled over 500,000 cells from 36 mouse brains across three age groups and conducted spatial transcriptomic analysis on twelve brain sections from aged mice under CR and control conditions. This comprehensive approach allowed us to explore the impact of CR on over 300 cellular states and assess region-specific molecular alterations. Our findings reveal that CR effectively modulates key aging-associated changes, notably by delaying the expansion of inflammatory cell populations and preserving cells critical to the neurovascular system and myelination pathways. Moreover, CR significantly reduced the expression of aging-associated genes involved in oxidative stress, unfolded protein stress, and DNA damage stress across various cell types and regions. A notable reduction in senescence-associated genes and restoration of circadian rhythm genes were observed, particularly in ventricles and white matter. Furthermore, CR exhibited region-specific restoration in genes linked to cognitive function and myelin maintenance, underscoring its targeted effects on brain aging. In summary, the integration of single-nucleus and spatial genomics provides a novel framework for understanding the complex effects of anti-aging interventions at the cellular and molecular levels, offering potential therapeutic targets for aging and neurodegenerative diseases.

genomics↗

bioRxiv2025

Uncovering Convergent Cell State Dynamics Across Divergent Genetic Perturbations Through Single-Cell High-Content CRISPR Screening

High-throughput genomic studies have uncovered associations between diverse genetic alterations and disease phenotypes; however, elucidating how perturbations in functionally disparate genes give rise to convergent cellular states remains challenging. Here, we present PerturbFate, a high-throughput, cost-effective, combinatorial-indexing single-cell platform that enables systematic interrogation of massively parallel CRISPR perturbations across the full spectrum of gene regulation, from chromatin remodeling and nascent transcription to steady-state transcriptomic phenotypes. Using PerturbFate, we profiled over 300,000 cultured melanoma cells to characterize multi-modal phenotypic and gene regulatory responses to perturbations in more than 140 Vemurafenib resistance-associated genes. We uncovered a shared dedifferentiated cell state marked by convergent transcription factor (TF) activity signatures across diverse genetic perturbations. Combined inhibition of cooperative TF hubs effectively reversed cellular adaptation to Vemurafenib treatment. We further dissected phenotypic responses to perturbations in Mediator Complex components, linking module-specific biochemical properties to convergent gene activations. Together, we reveal common regulatory nodes that drive similar phenotypic outcomes across distinct genetic perturbations. We also delineate how perturbations in functionally unrelated genes reshape cell state. PerturbFate thus establishes a versatile platform for identifying key molecular regulators by anchoring multi-modal regulatory dynamics to disease-relevant phenotypes.

genomics↗

bioRxiv2025

Holistic Motor Control of Zebra Finch Song Syllable Sequences

How brain circuits are organized to skillfully produce learned sequences of behaviors is still poorly understood. Here, we functionally examine how the cortical song premotor region HVC, which is necessary for zebra finch song, controls the sequential production of learned song syllables. We find that HVC can generate the complete sequence of learned song syllables independently of its main synaptic input pathways. Thalamic input to HVC is permissive for song initiation but it is not required for transitions between syllables or completing song. We show that excitation of HVC neurons during song reliably causes vocalizations to skip back to the beginning of song, reminiscent of a skipping record. This restarting of syllable sequences can be induced at any moment in song and depends on local circuits within HVC. We identify and computationally model a synaptic network including intratelencephalic premotor and corticostriatal neurons within HVC that are essential for completing song syllable sequences. Together, our results show that the learned zebra finch song is controlled by a cortical sequence-generating network in HVC that, once started, can sustain production of all song syllables independent of major extrinsic input pathways. Thus, sequential neuronal activity can be organized to fuse well-learned vocal-motor sequences, ultimately achieving holistic control of this naturally learned behavior.

neuroscience↗