Biology subjects

Jiang, S.

Publications and source records attributed to Jiang, S..

At least 55 records · Page 3Linked to original sources

bioRxiv2026

Single-Cell and Spatial Methods for Multimodal Functional Glycan Profiling in Tissues

Glycans regulate multiple physiological processes, including immune recognition and cancer progression. In disease, altered glycan landscapes are interpreted by human lectins. Functional glycan-lectin interactions are difficult to profile because glycans are not genome-encoded and their changes are poorly captured by existing multimodal methods. We present two platforms, single-cell outlining and transcriptome sequencing (scGOAT-seq) and GlycoScope, which use human lectins to enable functional glycan accessibility into single-cell and spatial multiomic measurements. ScGOAT-seq quantifies lectin-accessible glycan states with gene expression, while GlycoScope enables multiplexed in situ co-detection of glycans and proteins in tissues. Applying these approaches to immune cells, we identify stimulus-specific glycan remodeling and show that distinct Siglec-ligand-defined programs stratify immune activation states not captured by traditional methods; in follicular lymphoma, GlycoScope, resolves spatial glycan programs associated with malignant B cells and localized immune microenvironments. The presented methods provide a general framework for integrating functional glycan accessibility into single-cell and spatial multiomics.

molecular biology↗

bioRxiv2026

Pixel2Gene enables histology-guided reconstruction and prediction of spatial gene expression

Advances in spatial transcriptomics (ST) have fundamentally transformed our understanding of tissue biology by enabling gene expression profiling within intact spatial contexts and uncovering tissue organization and microenvironmental interactions. However, current high-resolution ST platforms remain constrained by high costs, limited tissue coverage, and technical artifacts, often yielding noisy, sparse, and incomplete data that compromise analytical accuracy, biological interpretation, and clinical utility. To address these challenges, we introduce Pixel2Gene, a deep learning framework that integrates co-registered histology images with ST data to enable histology-guided reconstruction and prediction of spatial gene expression. Pixel2Gene enhances existing expression measurements by denoising low-confidence data and reconstructing coherent expression patterns, while also predicting gene expression in unmeasured tissue regions and new samples lacking direct transcriptomic profiling. We systematically evaluated Pixel2Gene across multiple high-resolution ST platforms, including Visium HD, Xenium, and CosMx, spanning diverse tissue types and disease contexts using downsampling simulations and cross-platform comparisons in clinical samples. Across all settings, Pixel2Gene consistently improved data consistency, mitigated dropout effects, restored biologically meaningful spatial structure, and enabled accurate downstream analyses. By leveraging the scalability and ubiquity of routine histology, Pixel2Gene facilitates comprehensive, cost-effective ST profiling at whole-tissue scale, supporting large cohort studies, translational research, and next-generation biomarker discovery.

genomics↗

bioRxiv2026

Reconstructing multi-scale tissue spatial architecture from single-cell RNA-seq with REMAP

Understanding spatial organization of cells is critical for deciphering tissue function and disease. Single-cell RNA-sequencing (scRNA-seq) profiles transcriptomes at scale but loses spatial context, while spatial transcriptomics (ST) preserves spatial information but is constrained by cost and gene coverage. Here, we present REMAP, a deep learning framework that integrates gene expression with neighborhood-level gene-gene covariance to reconstruct multi-scale spatial organization of scRNA-seq data using one or multiple ST references. Across 2D and 3D mouse brain, human fetal cortex, and seven human cancer types, REMAP consistently outperformed existing approaches. Applied to a human multiple sclerosis atlas, REMAP resolved microglial neighborhood heterogeneity, and identified a rare pro-inflammatory microglia-astrocyte subpopulation. Across diverse cancers, REMAP recovered conserved spatially-defined cancer-associated fibroblast subtypes with known prognostic significance. By transforming cost-efficient single-cell datasets into spatially interpretable tissue maps, REMAP enables spatial hypothesis generation, microenvironment discovery, and population-scale inference of conserved and perturbed architectural principles in human disease.

genomics↗

bioRxiv2026

PlotGDP: an AI Agent for Bioinformatics Plotting

High-quality bioinformatics plotting is important for biology research, especially when preparing for publications. However, the long learning curve and complex coding environment configuration often appear as inevitable costs towards the creation of publication-ready plots. Here, we present PlotGDP (https://plotgdp.biogdp.com/), an AI agent-based web server for bioinformatics plotting. Built on large language models (LLMs), the intelligent plotting agent is designed to accommodate various types of bioinformatics plots, while offering easy usage with simple natural language commands from users. No coding experience or environment deployment is required, since all the user-uploaded data is processed by LLM-generated codes on our remote high-performance server. Additionally, all plotting sessions are based on curated template scripts to minimize the risk of hallucinations from the LLM. Aided by PlotGDP, we hope to contribute to the global biology research community by constructing an online platform for fast and high-quality bioinformatics visualization. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=78 SRC="FIGDIR/small/702995v3_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@73c77dorg.highwire.dtl.DTLVardef@12ddfc6org.highwire.dtl.DTLVardef@be5963org.highwire.dtl.DTLVardef@de90b6_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstract.C_FLOATNO PlotGDP: an AI Agent for Bioinformatics Plotting C_FIG

bioinformatics↗

bioRxiv2026

Synaptopodin enables directional mechanoadaptation of integrin-based adhesions

The attachment of cells to their substrate through adhesion complexes is fundamental to tissue architecture and function. These adhesions are inherently optimized to resist shear forces parallel to the substrate, yet certain specialized cells must also withstand substantial perpendicular forces. How cells adapt their adhesion machinery to resist forces in different directions has remained unclear. In the kidney, podocytes experience perpendicular forces from pressurized filtrate flow while maintaining attachment to the glomerular basement membrane through integrin-based adhesions. Here we show that synaptopodin converts adhesions from shear-resistant to perpendicular force-resistant structures through coordinated reorganization of the actin cytoskeleton and adhesion complexes. Using an inertial force application system, we demonstrate that synaptopodin triggers force-dependent redistribution of {beta}1-integrin to the cell periphery specifically in response to perpendicular loading, while synaptopodin-deficient cells lack this directional adaptation and detach. This mechanism operates in multiple cell types and is physiologically essential: synaptopodin-null mice subjected to elevated glomerular pressure develop significant proteinuria and podocyte foot process effacement. These findings reveal a molecular basis for directional mechanoadaptation, whereby a single protein enables cells to reconfigure their adhesion architecture in response to the direction of applied force.

biophysics↗

bioRxiv2026

Orthogonal Force Balance Between Contractility and Shear Stress Governs Podocyte Dynamics

Maintenance of tissue barriers under mechanical stress represents a fundamental biological challenge across organ systems. In the kidney, podocyte cells withstand highly variable hemodynamic forces while preserving a tensioned, nanostructured filtration barrier. Dysregulation of this barrier leads to significant pathology, but the mechanical principles underlying homeostasis of cells against flow of filtrates have not yet been identified. Here, we uncover a counterintuitive mechanical homeostasis mechanism whereby podocyte attachment depends on a dynamic balance between external fluid shear stress and internal cellular contractility. Integrated biomechanical modeling and experiment reveal a previously unrecognized mechanosensing circuit that optimizes integrin distribution at foot process peripheries. Our mathematical framework for cell-matrix adhesion stability reveals, surprisingly, that reducing blood pressure can worsen outcomes when cell contractility is impaired, contrary to clinical belief that lowering blood pressure universally benefits cellular adhesion and kidney function. We validated this principle through mouse models with manipulated blood pressure and myosin inhibition, demonstrating that concurrent reduction of both shear stress and contractility worsens podocyte injury and proteinuria. Super-resolution microscopy confirms our predicted integrin redistribution patterns under these mechanical perturbations. These findings establish a fundamental mechanobiological principle applicable beyond nephrology, and suggest potential treatment pathways targeting non-equilibrium steady states.

cell biology↗

bioRxiv2026

Phospholipase D1 and Phosphatidic Acid are required for MVE Fusion and Exosome Secretion

Extracellular vesicles (EVs) mediate critical intercellular communication, yet the molecular mechanisms that govern multivesicular endosome (MVE) fusion with the plasma membrane and exosome release remain poorly understood. Phospholipase D1 (PLD1) produces phosphatidic acid (PA), a lipid involved in membrane remodeling, but when and how PLD1 and PA act during exosome secretion has not been defined. Here, we used immunofluorescence and total internal reflection fluorescence microscopy (TIRFM) to track individual CD63+ MVEs together with fluorescent PLD1 or a PA reporter (GFP-PASS) in A549 cells. PLD1 localized to CD63+ MVEs during visiting, docking, and fusion. Inhibition or knockdown of PLD1 significantly reduced MVE fusion frequency and decreased the number of secreted small EVs, while causing only a modest reduction in vesicle availability at the plasma membrane. PLD1 inhibition also increased the number of membrane-proximal lysosomes, suggesting that MVEs are diverted toward degradation when fusion is impaired. Meanwhile, PA dynamics were stage-specific: PA remained low on visiting vesicles, gradually accumulated during docking, and exhibited a sharp spike followed by loss during fusion. PA has been reported to stabilize negative curvature and potentially play a role during fusion. To test whether PA influences fusion pore behavior, we quantified CD63 decay duration (t1/2) for individual fusion events. K-means clustering revealed that vesicles with longer decay durations had higher PA levels, whereas short-decay events showed minimal PA. The PA intensity correlated positively with decay duration, while cytosolic GFP did not, indicating a specific relationship between PA and exosome release rates. Furthermore, pharmacological activation of PLD1 increased the proportion of long-decay events. Together, these findings demonstrate that PLD1-generated PA regulates MVE fate at two levels: it promotes the docking-to-fusion transition and prolongs exosome release. This identifies a lipid-based mechanism that controls both the efficiency and kinetics of exosome secretion. SIGNIFICANCE STATEMENTExosome release is essential for cell-cell communication in processes such as development, immunity, and cancer. However, the mechanism that determines whether a multivesicular endosome fuses with the plasma membrane to release exosome cargo remains unclear. We identified a lipid-based mechanism that regulates both fusion and the rate of content release. We show that phospholipase D1 (PLD1) and its product phosphatidic acid (PA) act at late stages of exosome secretion: PLD1 promotes the docking-to-fusion transition and PA levels quantitatively correlate with content release rate, suggesting that PA stabilizes the pore. These findings demonstrate that lipids can control secretion frequency and kinetics, providing a new framework for understanding and modulating exosome output.

biophysics↗

bioRxiv2026

Rapid evolution in necromass use under resource limitation reduces persistence in producer-decomposer microbial biospheres

This study employs a synthetic microbial biosphere model to investigate system persistence under resource limitation. Escherichia coli (E. coli) strain MC4100 was incubated in matter-closed systems in an isotonic saline solution without added nutrients for 60 days in spatially homogeneous or heterogeneous conditions. We observed phenotypic adaptive changes in E. coli populations, including loss of motility, enhanced biofilm formation, and improved growth on various carbon sources, which were specific to the spatial structure conditions. While these changes were beneficial for E. coli persistence in monoculture, during co-culture with the alga Chlamydomonas reinhardtii the adapted populations exhibited a 25% reduction in persistence under spatially heterogeneous conditions compared to co-cultures containing non-adapted E. coli populations. Critically, adaptive changes in monoculture affected the dynamics of co-cultures, demonstrating that microbial evolution can constrain coexistence with other species and may be a fundamental determinant of ecosystem sustainability.

ecology↗

bioRxiv2026

Ac-FAPI-04 Promotes Anti-tumor Immunity via IL-27 Activation and Tnfaip3 Inhibition

BackgroundAccumulating evidence substantiates the capacity of targeted radionuclide therapy (TRT) to potentiate tumor immunotherapy responses. However, combining TRT and immunotherapies exhibits substantial varying efficacy, underscoring the importance of investigating mechanisms through which TRT remodels the tumor microenvironment (TME). Despite recent advancements, the specific signaling molecules mediated by TRT that regulate the functions of cytotoxic T lymphocytes (CTLs) remain unclear. MethodsGiven the demonstrated potential of -radionuclides targeting the fibroblast activation protein (FAP) in TRT, this study employed [225Ac]Ac-FAPI-04 to investigate the mechanisms of TRT-induced TME remodeling. The cytotoxic effects were first evaluated in the B16F10 cell line, followed by an assessment of its in vivo therapeutic efficacy in tumor-bearing mice. RNA-sequencing analysis was utilized to elucidate underlying signaling pathways involving immune responses. Results[225Ac]Ac-FAPI-04 significantly induced DNA double-strand breaks, further elevated ROS levels, and apoptosis in tumor cells. Transcriptomic profiling revealed that [225Ac]Ac-FAPI-04 activates the IL-27/JAK-STAT pathway to amplify interferon-mediated responses, indicating IL-27 serves as a critical regulatory signal for CTLs during TRT. A concomitant observation was the downregulation of Tnfaip3 following [225Ac]Ac-FAPI-04 treatment. Tnfaip3 is implicated in modulating CTLs behavior and intratumoral infiltration, while its downregulation was associated with the upregulation of cytotoxic effector molecules, as confirmed in this study. ConclusionThis study identifies a TRT-induced pattern of synergistic signaling mediated by IL-27 and Tnfaip3, which collectively enhances CTL recruitment and potentiates their cytotoxic function. These findings provide a detailed mechanistic rationale for strategies aimed at augmenting the therapeutic efficacy of TRT and immunotherapy combinations.

molecular biology↗

bioRxiv2025

FlashAffinity: Bridging the Accuracy-Speed Gap in Protein-Ligand Binding Affinity Prediction

Accurate prediction of protein-ligand interactions is central to computational drug discovery. Recent foundation models such as Boltz-2 have achieved remarkable accuracy in binding affinity prediction, yet their prohibitive computational cost remains a major barrier to large-scale virtual screening. Here we introduce FlashBind, a lightweight structure-based model that achieves a 50x speedup over Boltz-2 at inference time by replacing expensive structure prediction with a fast docking model and substituting costly PairFormer modules with a streamlined EGNN architecture. FlashBind attains early enrichment competitive with Boltz-2 on standard virtual screening benchmarks and demonstrates strong generalization to enzyme-substrate specificity prediction. To evaluate real-world applicability, we apply FlashBind to target-based antibiotic screening against the essential bacterial proteins in E. coli and show that FlashBind substantially outperforms Boltz-2 and other virtual screening baselines. Notably, several top-ranked candidates exhibit potent inhibition of DnaG and effective bacterial growth inhibition against E. coli in wet-lab validation. Together, these results demonstrate that FlashBind bridges the gap between accuracy and efficiency, enabling ultra-fast and accurate screening of massive chemical libraries for drug discovery.

molecular biology↗

bioRxiv2025

Adapting Epigenetic Clocks for Cell-Free DNA High-Throughput Sequencing Data

Background.Circulating cell-free DNA (cfDNA or ccfDNA) methylation sequencing holds promise for developing epigenetic aging clocks in minimally invasive aging assessment applications. However, current clocks--primarily trained on array-based data--do not readily generalize to methylation profiles of cfDNA, due to stochasticity and uncertainty arising from the limited input amounts of cfDNA and technical characteristics of high-throughput sequencing (HTS) platform. Due to lack of training to correct this uncertainty, direct application of legacy clocks to HTS data will inevitably produce unreliable estimates, introduce predictive discordance and undermine the trustworthiness of epigenetic biomarkers. Despite this urgent challenge, a systematic, model-agnostic framework for adapting existing epigenetic clocks to HTS-based cfDNA data remains lacking. Methods.Here, we generated a dedicated benchmark dataset comprising paired cfDNA and genomic DNA (gDNA), which was replicated and profiled across two methylation arrays and two targeted HTS platforms. We evaluated 53 epigenetic clocks for CpG coverage, reproducibility, cross-platform consistency, and age prediction accuracy. Further, we systematically explored and tested multiple adaptation strategies including depth filtering, beta-value imputation, and transfer learning via model distillation aiming at improving performance and clinical concordance of legacy epigenetic clocks on cfDNA HTS datasets. Results.Our results show that inherent technical noise of HTS platforms compromises diagnostic precision, presenting as a significant burden for applying legacy clocks on cfDNA HTS data. However, this technical limitation can be systematically neutralized. By enforcing stringent sequencing depth protocols ([≥]10x ideally 20x), employing robust algorithmic stabilization (L2-heavy clocks and imputation) and transfer learning, we can effectively isolate genuine physiological aging signals from technical artifacts. Ultimately, an adaptation pipeline incorporating all these methods effectively improved age prediction accuracy and, closed the gap between epigenetically induced aging status and clinically assessed realities. Besides, the pipeline demonstrated improved diagnostic sensitivity for neurodegenerative conditions such as amyotrophic lateral sclerosis, establishing a reliable foundation for non-invasive clinical monitoring. Conclusions.This study provided a comprehensive framework and practical guidelines for adapting epigenetic clocks to HTS-based cfDNA data, paved the path for more general application of cfDNA-based aging assessment and liquid biopsies.

bioinformatics↗

bioRxiv2025

Repurposed endogenous virus-like vesicles mediate dendritic cell long-range antigen presentation and T cell activation for enhanced cancer vaccination

Dendritic cells (DCs) release extracellular vesicles (DEVs) that amplify antigen presentation while incorporating patient-derived, rapidly evolving antigens. By enriching peptide-MHC and co-stimulatory ligands orders-of-magnitude above donor-cell levels, DEVs emerge as potent vesicle-vaccines, although efficacy remains limited by unclear mechanisms. We show that DCs repurpose viral components to generate endogenous virus-like vesicles (VLVs) that preferentially carry peptide-MHC and high-density co-stimulatory ligands, intensifying and extending antigen presentation. Upon antigen exposure, Arc partners with endogenous envelope proteins to assemble VLVs that directly engage T cells and trigger intrinsic adjuvanticity via viral mimicry. Arc-/- DEVs failed to prime antigen-specific T cell responses, whereas Arc overexpression with its 5'-UTR stem-loop shifted DEVs toward VLVs that trafficked to lymphoid organs, drove rapid CD4-assisted priming and durable CD8-biased memory, suppressed melanoma, and prolonged survival. These reveal a viral-mimicry mechanism enabling long-range immune activation and support Arc+ VLVs as an antigen-agnostic vaccine for cancer immunotherapy. HighlightsO_LIArc+ VLVs intensify and extend DC antigen presentation in vivo C_LIO_LIArc+ VLVs directly engage T cells and trigger viral-mimic adjuvanticity C_LIO_LIArc-/- DEVs fail to prime antigen-specific T cell responses C_LIO_LIEngineered Arc+ VLVs drive durable CD8-biased memory and tumor control C_LIO_LIArc+ VLV vaccination provides long-range and cross-tumor protection in vivo C_LI

bioengineering↗

bioRxiv2025

NSUN2-dependent 5-methylcytosine Modification Regulates Influenza A virus Gene Expression and Genomic Packaging

Emerging evidence indicates that methyltransferase NSUN2 catalyzes 5-methylcytosine (m5C) modifications on various viral RNAs and plays important roles in viral biology. However, the regulatory roles of NSUN2 in influenza A virus (IAV) replication have not been elucidated. Here, we revealed that NSUN2 negatively regulated the viral RNA transcription and protein production by knocking out and over-expressing NSUN2. By m5C MeRIP-seq and RNA-BisSeq, NSUN2-dependent m5C sites were identified on both the plus and minus viral RNA strands. In NSUN2-KO cells, the m5C modification on vRNAs was reduced, resulting in the production of a large number of deficient interfering particles (DIPs) which had lower vRNA content, imbalanced genome fragments, abnormal morphology and reduced pathogenicity. Mutation of m5C sites at the 5 and 3 ends of PB2 vRNA interfered with the selective packaging of the 8 vRNA segments into virus particles, resulting in the formation of a variety of abnormally packaged virus particles. PB2-vRNA mutants also had reduced replication ability and pathogenicity in mice. Overall, these data demonstrate that the m5C residues catalyzed by NSUN2 are required on vRNAs for the proper assembly of infectious viral particles, suggesting the depletion of m5C modification as a potential strategy that can be utilized to attenuate IAV strains. Significance StatementInfluenza A virus (IAV) infections pose a significant threat to global public health by causing substantial morbidity and mortality. The segmented nature of the IAV genome requires precise regulation of the genomic assembly to produce infectious progeny virus particles. In this study, we demonstrate that m5C modification is crucial for the proper assembly of infectious viral particles. The lowered level of m5C modifications on vRNAs leads to the generation of defective interfering particles with reduced replication capacity and pathogenicity. Our data shed new light on the selective packaging mechanism of IAV segmented genome and highlight a potential new strategy for attenuating IAV strains by targeting the m5C modification machinery.

microbiology↗

bioRxiv2025

Antiphospholipid syndrome (APS) is a platelet factor 4 (PF4)-centric immunothrombotic disorder

Antiphospholipid syndrome (APS) is an immunothrombotic disorder, frequently attributed to autoantibodies that bind {beta}2-glycoprotein I ({beta}2GPI). A study showed that the platelet-specific chemokine, platelet factor 4 (PF4), binds to {beta}2GPI, enhancing recognition of {beta}2GPI by APS antibodies. APS antibodies induce the release of neutrophil extracellular traps (NETs), webs of decondensed chromatin that bind both PF4 and {beta}2GPI. We propose that PF4 bridges {beta}2GPI to NETs (and other PF4-targeted polyanions), leading to the formation of prothrombotic PF4:{beta}2GPI:NET immunotargets in APS. Dynamic light-scattering studies of isolated IgGs from four patients with triple-positive APS show formation of PF4:{beta}2GPI:NET complexes that bind APS antibodies. NETs released in a microfluidic system bound {beta}2GPI, but only in the presence of PF4, forming a multimolecular APS antigenic target. Whole blood infused through a photochemically-injured, endothelium-lined microfluidic channel formed platelet-, fibrin-, and complement-rich thrombi that bound APS antibody only in the presence of PF4. Thrombi were reduced in size if either ADAMTS13 or DNase1 was infused. In a murine APS model, wildtype and transgenic mice expressing platelet human PF4 {+/-} Fc{gamma}RIIA developed more intense neutrophil rolling along veins, and more extensive thrombus formation following laser injury to cremaster arterioles and venules, whereas mice lacking PF4 did not. Three antigenically distinct anti-hPF4 monoclonal antibodies blocked thrombosis in vitro, and neutrophil rolling and thrombosis in vivo. Our studies provide new insights into the basis of APS that has mechanistic parallels to other known PF4 immunothrombotic disorders and offer potential diagnostic and non-anticoagulant therapeutic strategies for clinical management. Key pointsO_LIPF4 enhances {beta}2GPI binding to NETs and these complexes are central to APS immunothrombosis. C_LIO_LIAnti-PF4 monoclonal antibodies block APS immunothrombosis in microfluidic and murine studies. C_LI

cell biology↗

bioRxiv2025

Chicken IRF10 suppresses the cGAS-STING-IFN antiviral signaling pathway by targeting IRF7

The chicken (ch) innate immune cGAS-STING-IFN signaling pathway plays a crucial role in host antiviral defense, wherein chIRF7 serves as the key downstream transcription factor of chSTING responsible for initiating type I IFN transcription. While chicken IRF family encompasses 8 members, the potential involvement of other IRF members in chicken cGAS-STING-IFN signaling transduction and/or regulation remains poorly characterized. In this study, we identified chIRF10 as a potent suppressor of chicken cGAS-STING-IFN signaling against several viruses, by screening of chIRF family members. Mechanistically, the inhibitory activity of chIRF10 depended on its IRF associated domain (IAD) but not its DNA binding domain (DBD). Further, chIRF10 suppressed both cGAS-STING mediated IFN signaling and TBK1/IKK{varepsilon}/IRF7 activated IFN signaling. Importantly, chIRF10 could interact with chIRF7, and inhibit its dimerization and activation. In summary, our findings unveiled a novel function of chIRF10 as well as a novel regulatory mechanism governing chicken cGAS-STING-IFN antiviral signaling, thereby providing a scientific foundation for developing strategies against chicken viral diseases.

immunology↗

bioRxiv2025

Transcription profiling and functional analysis of spRNAs and their corresponding asRNAs in Methanosarcina mazei

Small proteins (sPs) encoded by small RNAs (sRNAs), the so-called small protein RNAs (spRNAs), and their corresponding antisense RNAs (asRNAs) have emerged as important regulatory elements in microbial stress adaptation, yet remain poorly characterized in archaea. In this study, we identified and characterized five sRNA/asRNA pairs in the methanoarchaeon Methanosarcina mazei. Expression analysis revealed condition-specific regulation of two of those pairs, spRNA23/asRNA93 and spRNA24/asRNA94, under oxidative, temperature, and osmotic stress. Overexpression of the corresponding small proteins, sP23 and sP24, revealed distinct physiological effects: both enhanced growth under high-salt conditions, while sP24 in addition supported cell size maintenance during osmotic stress, as confirmed by transmission electron microscopy. Consistently, overexpression of asRNA94 let to impaired growth under salt stress, suggesting its negative regulatory role towards sP24. The results also imply that the antisense RNA might be crucial for controlling sP24 expression under non-stress conditions. Bioinformatic analysis predicted one transmembrane domain in sP24. Membrane-association of sP24 was experimentally validated in vitro using a cell-free expression system, demonstrating sP24s integration into lipid bilayers. Conservation analysis showed that spRNA24 appears specific for M. mazei strains potentially reflecting niche adaptation, while spRNA23 is widespread within Methanosarcina. These findings demonstrate that archaeal sRNAs can encode functional small proteins, often encoded together with their respective regulatory asRNA, that contribute to environmental stress responses, overall offering new insights into the role of sRNA-derived small proteins in archaea.

microbiology↗

bioRxiv2025

Aphid-derived cross-kingdom RNA dynamics underpin maize resistance

Aphids are major pests of maize (Zea mays), yet the molecular mechanisms underlying their interactions with host plants remain poorly understood. Here, we identified and functionally characterized long non-coding RNAs (lncRNAs) from the aphid-specific Ya gene family in the cereal-specialist aphids Rhopalosiphum maidis and R. padi. We showed that Ya genes formed lineage-specific clusters and were transcriptionally active across both species. Multiple Ya transcripts secreted into maize tissues exhibited remarkable stability compared with rapidly degraded aphid mRNAs and migrated systemically, as visualized in planta using an RNA switch-controlled RNA-triggered fluorescence system. RNA interference of Ya genes significantly reduced aphid fecundity, while ectopic expression of Ya lncRNAs in maize enhanced aphid colonization. Importantly, maize genotypes differing in aphid resistance selectively influenced the persistence of aphid mRNAs but not Ya lncRNAs, indicating a decoupled fate of distinct aphid RNA classes in planta. These findings establish Ya lncRNAs as cross-kingdom effectors critical for aphid virulence, and suggest new molecular markers for breeding aphid-resistant maize.

plant biology↗

bioRxiv2025

Small Molecule Approach to RNA Targeting Binder Discovery (SMARTBind) Using Deep Learning Without Structural Input

Accurate identification of small molecule binders to RNA is critical for chemical probes and therapeutics. Computational approaches offer a cost-effective strategy to identify small molecules targeting RNA but are often limited by poor predictive accuracy and high computational demands. Here, we introduce Small Molecule Approaches to RNA Targeting Binder Discovery (SMARTBind), a structure-agnostic ligand discovery framework that combines an RNA large language model with contrastive learning and a ligand-specific decoy enhancement strategy. The RNA language model, pre-trained on millions of RNA sequences, together with the decoy enhancement strategy, addresses data scarcity and improves model generalizability. SMARTBind uses only RNA primary sequence to identify small molecule binders and their binding sites accurately. Across multiple benchmarks and case studies, SMARTBind outperforms existing data-driven and docking-based methods while significantly reducing computational cost. In a real- world application, SMARTBind identified novel small molecules targeting the precursor of oncogenic microRNA-21, validated by in vitro and cellular assays. These results highlight SMARTBinds potential as a scalable, accurate, and structure-independent platform for RNA-targeted small molecule discovery.

bioinformatics↗