Biology subjects

Zhu, Y.

Publications and source records attributed to Zhu, Y..

At least 163 records · Page 9Linked to original sources

bioRxiv2025

Torsins Organize CLCC1 Assembly to Safeguard ER Bilayer and Lipid Homeostasis

The TMEM41B scramblase and its regulatory partner CLCC1 initiate lipid flux by equilibrating newly-synthesized phospholipids across the endoplasmic reticulum (ER) bilayer, a fundamental process required for diverse events ranging from membrane biogenesis to bulk lipid supply. Loss of CLCC1/TMEM41B causes ER bilayer imbalance, inducing giant ER-enclosed lipid droplets (geLDs) and driving rapid progression into severe MASH. Here we identify CLCC1 as the long-missing client of the lumenal Torsin ATPases, which selectively engage oligomerized CLCC1 at sites of ER bilayer imbalance. Hepatic TorsinA inactivation triggers geLD formation amid disrupted lipoprotein biogenesis and severe MASH, closely phenocopying CLCC1/TMEM41B deficiency. Mechanistically, Torsins act as foldases that drive CLCC1 oligomerization for its recruitment to imbalanced bilayers. Remarkably, ectopic CLCC1 expression reverses cellular and systemic lipid disorders arising from hepatic TorsinA deficiency. Hence, Torsin ATPases emerge as fundamental regulators that organizes CLCC1 and the downstream TMEM41B scramblase to govern lipid partitioning and membrane homeostasis.

cell biology↗

bioRxiv2025

Multimodal analysis defines GNG4 as a distinguishing feature of germinal center-positioned CD4 T follicular helper cells in humans

CD4 T follicular helper (Tfh) cells coordinate humoral immune responses within germinal centers (GC) of lymphoid tissue. Despite their critical roles in vaccination and autoimmunity, the gene expression programs that define functionally distinct human Tfh states-- and the molecular pathways engaged by Tfh positioned within the GC niche--remain incompletely understood. This gap has limited translational efforts to monitor or therapeutically target specific Tfh states for clinical benefit. Here, we delineate human CD4 T cell heterogeneity in tonsils and peripheral blood using trimodal single-cell sequencing and spectral flow cytometry to define epigenomic, transcriptional, and proteomic features of distinct Tfh states. Tfh with a GC-like phenotype exhibited markedly increased chromatin accessibility and both mRNA and protein expression of G protein subunit gamma 4 (GNG4). In tonsil, single-cell spatial transcriptomics defined GNG4 expression as a distinguishing feature of activated Tfh states within spatially demarcated GC compartments, with greater specificity than conventionally GC-associated features such as BCL6, TOX2, and S1PR2. In contrast, GNG4- Tfh primarily localized to nonGC regions and exhibited a resting, Th17-polarized phenotype. Together, these data highlight GNG4 as a central feature of activated, GC-positioned Tfh cell identity in humans. One Sentence SummaryGNG4 expression defines activated CD4 T follicular helper cells localized to the germinal center of human lymphoid tissue.

immunology↗

bioRxiv2025

TMEM Doorway Mediated Metastasis in Pancreatic Ductal Adenocarcinoma by Tie2 Signaling

Pancreatic ductal adenocarcinoma (PDAC) is almost invariably fatal due to early hematogenous dissemination that occurs before the primary tumor is clinically detectable, yet the cellular mechanism of tumor cell intravasation has remained unknown. Using multiphoton intravital imaging in autochthonous and orthotopic PDAC models, we demonstrate that intravasation occurs at Tumor Microenvironment of Metastasis (TMEM) doorways--tri-cellular structures comprising a MENA-expressing tumor cell, a Tie2 macrophage, and an endothelial cell in direct contact. These structures are abundant in human PDAC, enriched for Tie2 macrophages, and markedly reduced after neoadjuvant chemotherapy. Selective pharmacologic inhibition of Tie2 with rebastinib decreases TMEM-associated transient vascular openings, suppresses circulating and hepatic disseminated tumor cells, and--when combined with perioperative FOLFIRINOX after curative-intent resection--improves median survival in murine PDAC. These findings establish TMEM doorways as a common, druggable mechanism of intravasation across epithelial cancers and identify Tie2 macrophages as a therapeutic target to prevent metastatic seeding in PDAC, a disease with no anti-metastatic therapies. TMEM doorway-mediated intravasation in PDAC supports its role as a common gateway for hematogenous metastasis in carcinoma.

cancer biology↗

bioRxiv2025

Molecular basis of high-torque transmission of the Vibrio polar flagellar motor

The bacterial flagellar motors are huge protein nanomachines that drive rotation of the flagellum for bacterial motility, and have considerable diversity in structure among bacterial species, which enables the transmission of different torques to the flagellar filaments to propel bacteria and renders various swimming abilities. Relative to the bacterial peritrichous flagellar motors, the polar flagellar motors are the faster rotational machines and transmit high torque to drive bacterial high-speed motility in liquid and empower swimming in viscous environments. However, the structural basis of high-torque transmission of the polar flagellar motors is still unclear. Here we present an atomic-resolution cryo-electron microscope structure of the polar flagellar motor in complex with the hook from Vibrio alginolyticus, comprising 295 subunits from 18 proteins. Extensive inter-subunit interactions and additional phospholipids generate the higher rigidity of the rod. The LP ring utilizes more electrostatic charges on the inner surface and less physical contacts to facility the higher-speed rotation of the rod. The additional HT ring tightly binds to the outer surface of the LP ring to enhance the LP-ring stability. A previously function-unknown component FlrP enhances the interactions of 15 FliF peptides of the MS ring with the rod and stabilizes the LPHT and MS rings. The hook has two different states, L- and R-state, which provides structural flexibility of the hook to drive the unique forward-reverse-flick motility of the bacteria in the flicking process. This study provides unprecedented molecular insights into evolution and structural adaptions of the bacterial polar flagellar motors for high-torque transmission.

microbiology↗

bioRxiv2025

Design principles of neuromorphic computing using genetic circuits

Cells have evolved to sense a wide range of input combinations and integrate those signals through signaling pathways to produce context-specific responses, such as differentiation, cell-type specification, and patterning. To replicate this information-processing capacity, synthetic biology has developed large-scale circuitry inspired by the fundamental principles of computer science. Within this framework, neuromorphic computing implemented using genetic circuits offers the opportunity to significantly enhance the computational capabilities of single cells. In this work, we establish design principles for implementing neuromorphic computing in living cells by identifying the key feature that enables a chemical reaction network to function as a perceptron: an input-output mapping with a tunable threshold. We demonstrate that four ubiquitous chemical reaction networks, namely molecular sequestration, catalytic degradation, competitive binding, and activation/deactivation cycles, all satisfy this requirement and can be engineered as perceptrons. By layering these perceptrons into multi-layer architectures, we then show how to construct both linear and nonlinear decision boundaries through rational tuning of production rates that encode network weights. As proof of principle, we apply this framework to design neural networks capable of discriminating between healthy and cancer cells based on gene expression data from 19 tissue types. Together, this work formalizes the design principles for engineering genetic circuits as neural networks and establishes a foundation for implementing next-generation cellular computation.

synthetic biology↗

bioRxiv2025

Single-cell transcriptomic dynamics exposes hidden survival trajectory under antibiotic treatment

Heterogeneity plays a major role in bacterial resistance to antibiotic treatment while the mechanism at single-cell level is largely unknown. Here, we employed a robust integration of bulk and bacterial single-cell RNA-seq (scRNA-seq) to uncover how individual cells of Acinetobacter baumannii reorganise the heterogenous transcriptome in response to antibiotic. Using polymyxin as a representative, bulk RNA-seq showed canonical envelope- and efflux-centred responses but obscured underlying heterogeneity. Single-cell profiling resolved these averages into discrete subpopulations whose abundances shifted with concentration and time. Specifically, in early time an envelope-stress programme predominated survival at the low concentration, whereas an outer-membrane repair/efflux programme dominated survival at the high concentration. These patterns revealed structured and time-resolved heterogeneity, highlighting an ingenious bacterial stress responsive strategy. Through trajectory inference, we further revealed a concentration-dependent bifurcation of cell fates: low-concentration treated cells detoured through a transient stress state and rejoined growth, whereas high-concentration treated survivors diverted into a slow-growing, tolerant branch. Perturbing marker genes from these programmes altered fitness eventually and rapidly increased permeability, depolarisation and reactive-oxygen burden, linking state to survival capability. Collectively, we map a dynamic, concentration-structured landscape of antibiotic responses at the single-cell level, revealing diverse survival trajectories that are obscured in conventional population-averaged analyses. Our developed single-cell based framework revealed tolerant bacterial cells with rewiring of the transcriptional landscape, causing emergence of antibiotic resistance. Importantly, these findings urge precision antimicrobial therapy in patients to minimise emergence of antibiotic resistance.

microbiology↗

bioRxiv2025

BioLACE: unifying spatial geometry and marker priors for cohesive cell-type clustering in spatial transcriptomics

Spatial transcriptomics (ST) provides high-dimensional gene expression profiles together with spatial coordinates, enabling the reconstruction of tissue architecture at cellular resolution. While recent graph-based deep learning methods have advanced spatial clustering in ST, many rely on complex architectures that obscure interpretability and rarely integrate biological priors such as marker gene information. We introduce BioLACE, a scalable framework that unifies spatial structure, transcriptomic variation, and curated marker gene profiles within a shared Variational Autoencoder (VAE) latent space. BioLACEjointly optimizes three complementary objectives: (1) a VAE reconstruction loss to preserve transcriptional structure, (2) a graph Laplacian regularizer to enforce spatial smoothness, and (3) a temperature-scaled InfoNCE contrastive loss guided by marker-informed similarities. Applied to MERFISH hypothalamus, mouse spinal cord, and Slide-seq mouse cerebellum datasets, BioLACE achieves superior cell type clustering accuracy, well-defined biologically consistent boundaries, and interpretable latent representations, highlighting its generality and scalability for modern ST analysis. The source code and tutorials for BioLACE are publicly available at https://github.com/maiziezhoulab/BioLACE.

genomics↗

bioRxiv2025

Associations Between Age, Heart Rate Variability, and BOLD fMRI Signal Variability

Numerous studies report that BOLD fMRI signal variance (SDBOLD) decreases with age. However, these associations may partly reflect cardiovascular contributions to the BOLD signal. For example, heart rate variability (HRV) has been positively associated with Resting State Fluctuation Amplitude (RSFA), which captures low frequency components of BOLD fMRI variability. HRV is also negatively associated with age, which could potentially confound age-SDBOLD associations. Yet, limited research has examined HRV-SDBOLD associations or tested within-person HRV-SDBOLD coupling using sliding window analyses of simultaneous HRV and SDBOLD. We analyzed resting-state fMRI data from two independent Midlife in the United States (MIDUS) samples: Core at M3 (n=115) and Refresher at MR1 (n=101). Partial Least Squares (PLS) analyses revealed significant positive HRV-SDBOLD associations (Core: permutation p=0.018; Refresher: permutation p<0.001). Whole brain age-SDBOLD PLS associations were non-significant via permutation tests across several models (Core: permutation p=0.201; Refresher: permutation p=0.121). We found age-related decreases in SDBOLD across [~]70% of voxels in both samples. Concordance analyses showed 67-69% of brain voxels exhibited negative age-SDBOLD but positive HRV-SDBOLD relationships, suggesting that regions showing age-related decreases in SDBOLD also showed HRV-related increases in SDBOLD. Sliding-window analyses demonstrated robust positive within-person associations between person-centered HRV and SDBOLD via different HRV metrics: SDNN (Core: p < 0.001; Refresher: p < 0.001), RMSSD (Core: p = 0.072; Refresher: p = 0.009), and low frequency (Core: p < 0.001; Refresher: p < 0.001), with non-significant effects of high frequency (Core: p = 0.516; Refresher: p = 0.12) HRV. Thus, regardless of baseline levels, windows with higher HRV corresponded to higher SDBOLD, suggesting that cardiovascular factors partially explain age-SDBOLD associations and HRV may mechanistically influence SDBOLD. These results suggest that controlling for HRV, especially low-frequency HRV or SDNN, may be necessary when analyzing SDBOLD to isolate neural effects.

neuroscience↗

bioRxiv2025

Ataxin-2 knockdown is neuroprotective via cell-autonomous and non-cell-autonomous mechanisms

Ataxin-2 (ATXN2) is a genetic modifier of TDP-43 toxicity and a promising therapeutic target in amyotrophic lateral sclerosis (ALS). However, the mechanisms underlying its neuroprotective effects remain poorly understood. Here we show that ataxin-2 reduction confers neuroprotection by engaging adaptive metabolic programs in both neuronal and glial cells. Using global proteomic profiling in yeast and mouse TDP-43 models, we establish that pbp1/ataxin-2 (pbp1 is the yeast ataxin-2 ortholog) activates orthogonal stress-adaptive programs that enable alternative energy production and augment trophic support, rather than simply reversing TDP-43-induced damage. In neurons, ataxin-2 downregulation activates glycolysis and reductive glutamine carboxylation driven by IDH1, restoring ATP production independently of impaired mitochondria. In astrocytes, ataxin-2 downregulation upregulates cholesterol biosynthesis via HMGCS1, enhancing trophic support to neurons in a non-cell-autonomous manner. Full neuroprotection requires both mechanisms: neuronal survival is only completely rescued when ataxin-2 is reduced in the context of neuron-astrocyte co-cultures. Importantly, ataxin-2 downregulation protects against both TDP-43 gain-of-function and loss-of-function toxicity, broadening its therapeutic relevance. Collectively, our findings reveal novel mechanisms by which ataxin-2 orchestrates adaptive metabolic resilience and establish ataxin-2 as a regulator of stress-adaptive response in brain cells under the conditions of stress.

neuroscience↗

bioRxiv2025

Fluctuating flooding motivates the facilitation of a wetland invader: a probe into the stress gradient hypothesis

Stress gradient hypothesis proposes that the plant interaction tends to convert from competition to facilitation when facing accelerating stress. The present study aims to explore the interaction between two co-existing wetland plant invaders - Alternanthera philoxeroides and Myriophyllum aquaticum with exposure to flooding stress. Different plant assemblages including monospecific (single individual per species), homospecific (two individuals of same species) and heterospecific (two individuals of different species) treatments were subjected to different flooding regimes including mesic habitat, constant flooding and fluctuating flooding. The plant interaction was evaluated by biomass accumulation, biomass allocation and relative competition intensity index (RCI). We found that the biomass accumulation and allocation of A. philoxeroides were consistent across different treatments of community assemblage and flooding regime, while the root mass, biomass and root mass allocation of M. aquaticum significantly increased at the presence of A. philoxeroides. Moreover, the RCI of M. aquaticum decreased at the presence of A. philoxeroides. Especially, the RCI of M. aquaticum was 0.06 in the homospecific community, while that of M. aquaticum was -0.82 in the heterospecific treatment under fluctuating flooding condition. Overall, Alternanthera philoxeroides facilitated M. aquaticum, especially under fluctuating flooding condition, which complies with the stress gradient hypothesis. The water level drawdown can benefit the impedance of the growth of M. aquaticum and the deceleration of the co-invasion of A. philoxeroides and M. aquaticum.

ecology↗

bioRxiv2025

3-Phenylpropionic acid, a microbiota-derived polyphenol metabolite linked to hippuric acid, ameliorates colitis as a colon-enriched PPAR-γ agonist

Polyphenols are promising therapeutics for Crohns disease (CD), yet their bioavailability is largely dependent on the gut microbiome. The bioactive metabolites, metabolic pathways, and anti-inflammatory mechanisms underlying their effects remain poorly defined. We identified hippuric acid (HIPA) as a biomarker of polyphenol-microbiome interaction. HIPA is a meta-organismal metabolite produced through a cross-organ pathway that involves synthesis from gut microbial polyphenol metabolism via 3-phenylpropionic acid (3-PPA), followed by renal clearance. We found that CD is characterised by impaired polyphenol metabolism, resulting in deficient colonic 3-PPA and reduced serum hippuric acid relative to healthy individuals. Furthermore, we demonstrate that 3-PPA is a colon-enriched, mild PPAR-{gamma} agonist that ameliorates colitis in murine model. Systemic bioinformatic profiling of metagenomic data suggested the existence of undiscovered 3-PPA producers. Through targeted microbial incubation with polyphenols, we identified the probiotic Bifidobacterium breve and a lab isolate Escherichia coli FAH as novel 3-PPA producers. Collectively, our findings establish 3-PPA as a diet-dependent, colon-produced, mild PPAR-{gamma} agonist with the potential to minimise adverse effects associated with traditional PPAR-{gamma} agonists. This work establishes a foundation for developing microbiota-targeted, polyphenol-based therapeutic strategies for CD patients.

microbiology↗

bioRxiv2025

Uncovering structural determinants of peptide recognition by public and private T-cell receptors

Public T cell receptors (TCRs) recurrently emerge across individuals in response to common pathogens, yet the structural and biophysical basis distinguishing public from private clonotypes remains incompletely defined. Here, we combine epitope mapping, single-cell TCR sequencing, and single-particle cryo-electron microscopy to dissect CD8+ T cell responses to the immunodominant SARS-CoV-2 ORF3a(207-215) epitope presented by HLA-A*01:01. Among responding clonotypes, we identify a shared public TCR (TCRpub) and an individual-specific private TCR (TCRpriv) that use nearly identical TRBV5-1 {beta} chains but distinct chains. Both clonotypes exhibit comparable micromolar affinity and functional avidity, yet their structures reveal different antigen-recognition modes. We determined cryo-EM structures of the TCRpub and TCRpriv in complex with ORF3a(207-215)/ HLA-A*01:01 at [~]3 [A] resolution. Despite targeting the same epitope, the two receptors engaged the peptide-MHC complex with distinct CDR-loop orientations and contact footprints: TCRpub engages the peptide through a peptide-centric AGDL CDR3{beta} motif and focuses interactions on the MHC 2-helix, whereas TCRpriv distributes contacts across both MHC -helices via a canonical CDR3{beta} configuration. These findings illustrate how near-identical {beta} chains can yield divergent recognition strategies to recognise the same pMHC ligand through alternative -chain pairing. More broadly, this work establishes cryo-EM as a robust approach for resolving physiological-affinity TCR/pMHC complexes, providing mechanistic insight into how public TCRs emerge and persist in antiviral immunity.

immunology↗

bioRxiv2025

Lighting and circadian cues shape locomotor strategies for balance and navigation in larval zebrafish

Most fish are inherently unstable and must swim to stabilize posture. How diurnal fish reduce activity at night while maintaining postural control remains unclear. We defined distinct locomotor strategies that larval zebrafish (Danio rerio) use to control posture and navigate the water column in response to light and circadian cues. In the dark, larvae maintain balance by swimming in long bouts with large nose-up rotations, compensating for nose-down drift accrued during prolonged inactivity. Effective postural compensation requires vestibular sensation from the utricle. By contrast, in the light, larvae navigate with short, frequent, and variable bouts. While lighting exerts a dominant, masking effect on the locomotor strategies, circadian rhythms modulate swim kinematics. Our results reveal distinct light-dark locomotor strategies and disentangle how ambient light and the internal clock jointly shape balance control and navigation. This work lays the foundation for understanding how external and internal cues interact to govern locomotor activity in freely moving diurnal animals.

animal behavior and cognition↗

bioRxiv2025

A unified language model bridging de novo and fragment-based 3D molecule design delivers potent CBL-B inhibitors for cancer treatment

The rational design of small molecules is central to drug discovery, yet current artificial intelligence (AI) methodologies for generating three-dimensional (3D) molecules are often siloed, focusing on either de novo design or fragment-based design. The lack of a holistic framework limits AIs application across the complex and multi-step pipeline spanning from novel scaffold identification to lead compound optimization, and prevents AI from effectively learning from the entire process. Here, we introduce UniLingo3DMol, a language model for 3D molecular generation, empowered by fragment permutation-capable molecular representation alongside multi-stage and multi-task training strategy. This integrated design enables UniLingo3DMol to seamlessly span both de novo and fragment-retained molecular design, demonstrating superior performance over existing generation models in in silico evaluations across more than 100 diverse biological targets. We further leveraged UniLingo3DMol in the design of inhibitors targeting CBL-B, a crucial immune E3 ubiquitin ligase and attractive immunotherapy target. This strategy led to a lead compound demonstrating excellent in vitro activity and robust in vivo anti-tumor efficacy. Our findings establish UniLingo3DMol as a generalized and powerful platform, showing the strong potential to advance AI-driven drug discovery.

bioinformatics↗

bioRxiv2025

Bruch's Membrane Contributes to the Structural Integrity of the Human Eye

PurposeTo investigate the contribution of the Bruchs membrane and sclera tissues to the overall structural integrity of the ocular wall. MethodsTwenty-three human globes were subjected to biomechanical testing. A piece of sclera measuring 5 x 5 mm was carefully removed at the nasal region, 2 mm away from the optic nerve head. The intraocular pressure was increased at approximately 1 mmHg/s until Bruchs membrane-uvea-retina-tissue layer (BMUR) ruptured. Next, strips of sclera and Bruchs membrane-choriocapillaris (BMC) complex were isolated from the superior fundus region. Uniaxial tension tests were performed at a strain rate of 0.01/s and sampling rate of 15 Hz. The tangent moduli of the BMC and sclera at 0.01, 0.02 and 0.03 strains were compared. ResultsThe rupture pressure of the BMUR was 98.1 {+/-} 21.4 mmHg. The tangent moduli of the BMC at 0.01, 0.02 and 0.03 strains were 2.96 {+/-} 1.44 MPa, 7.68 {+/-} 1.78 MPa and 9.43 {+/-} 2.11 MPa, respectively, and the tangent moduli of the sclera at 0.01, 0.02 and 0.03 strains were 1.09 {+/-} 0.80 MPa, 2.72 {+/-} 1.67 MPa and 5.69 {+/-} 3.27 MPa, respectively. ConclusionThe BMUR was able to sustain relatively high IOP before rupturing. The uniaxial tensile tests showed that the BMC tangent moduli were about 3 times of those of the sclera at strains of 0.01 and 0.02. Although the sclera is approximately 47 times thicker, the BMC is still likely to make a significant contribution (3.51% to 7.42% at strain <0.03) to the overall structural strength of the ocular wall.

bioengineering↗

bioRxiv2025

Single-Islet Proteomics Maps Pseudo-Temporal Islet Immune Responses and Dysfunction in Stage 1 Type 1 Diabetes

Progressive {beta}-cell dysfunction precedes the onset of type 1 diabetes (T1D), yet the molecular mechanisms driving early T1D development remain poorly understood. Although single-cell RNA-sequencing has uncovered transcript-level changes in human islet cells, it offers limited insight into the heterogeneity of distinct islet microenvironments. Here, we applied a single-islet proteomics workflow to profile intra-donor islet heterogeneity in three stage 1 T1D cases with matched non-diabetic controls and define in situ protein signatures of pseudo-temporal islet dysfunction. Intra-donor analyses of [~]100 individual islets per donor revealed highly consistent proteomic patterns reflecting pseudo-time progression of islet immune responses and {beta}-cell dysfunction. Several pathways, including extracellular matrix remodeling and mRNA processing, were identified as closely associated with progressive islet immune activation and loss of {beta}-cell function. These findings provide robust proteome-wide evidence of the progression of islet dysfunction, offer a valuable resource for investigating early mechanisms of T1D pathogenesis-- including novel candidates for functional studies--and underscore the utility of single-islet spatial proteomics for examining islet heterogeneity in T1D.

systems biology↗

bioRxiv2025

An EEG-based Multi-Scale Hybrid Attention and Squeeze Network for objective taste assessment

Taste perception is central to flavor experiences. Electroencephalography (EEG) signals carry rich information about taste perception. Because these neural data are independent of verbal reports and bypassing conscious filtering, EEG holds promise as a neural-sensing platform for objective taste representation. However, taste-evoked EEG signals exhibit complex spatiotemporal and spectral dynamics that require advanced computational approaches to decode effectively. This study developed an EEG-based Multi-Scale Hybrid Attention and Squeeze Network (MHASNet) for taste evaluation. EEG signals evoked by distinct taste stimuli were recorded, a dedicated taste-EEG dataset was compiled, and a novel deep learning architecture, MHASNet, was designed to classify these signals. MHASNet synergistically integrates multi-scale convolutions to capture temporal dynamics across different time scales, dual-attention mechanisms to localize discriminative brain regions and electrode positions, and a squeeze-and-excitation module to optimize frequency-band contributions-collectively enabling precise extraction of taste-specific neural signatures. Results showed that the proposed model achieved superior performance across five taste categories (sweet, sour, salty, bitter, and tasteless), with 94.33% accuracy, 91.37% F1-score, 91.89% precision, and 92.07% recall. These results surpass benchmark models while maintaining millisecond-level inference latency suitable for real-time applications. By complementing subjective evaluations and instrumental analyses, the model offers an objective, neurophysiology-based solution for taste evaluation.

neuroscience↗

bioRxiv2025

Identification of Protein Cargo in Extracellular Vesicles from Macrophages in Progressing and Regressing Tumors

Exosomes are extracellular vesicles (EVs) that carry bioactive molecules from a cell of origin and may alter gene expression of the acceptor cell via binding to cell surface receptors and/or delivering their cargo into the cell. Tumor-associated macrophages (TAMs) can increase or decrease tumor growth through various mechanisms, but the impact of TAM-EVs on tumors has been difficult to elucidate due to challenges in isolating and culturing TAMs for EV purification. In this study, we set out to uncover the protein identities within EVs from TAMs in progressing and regressing tumors, thereby uncovering EV proteins roles in TAM-tumor crosstalk. TAMs were purified from tumors using magnetic bead isolation and cultured for up to 9 days. TAMs from regressing tumors were found to be more M1-like by high expression of major histocompatibility complex (MHC) class II, while TAMs from progressing tumors are more M2-like (low expression of MHC class II). Long-term in vitro culture of TAMs resulted in reduced expression of MHC class II. EVs were harvested from plated TAMs, and EV proteins were identified by mass spectrometry and compared to EV proteins from bone marrow-derived macrophages (BMDMs). Unsupervised hierarchical clustering studies revealed that proteins from TAM-EVs converge based on their in vitro culture duration rather than their tumor origin. Signature proteins in TAM-EVs were characterized, with the identification of galectin-3-binding protein as a signature protein present in EVs from M1-like BMDMs and regressor macrophages. Finally, immunostimulatory pathways were identified in progressor-TAM-EVs through protein-protein interaction network analysis.

immunology↗