Biology subjects

Zhao, Z.

Publications and source records attributed to Zhao, Z..

At least 127 records · Page 7Linked to original sources

bioRxiv2025

Mapping Human Survivability at Extreme Wet-Bulb Temperatures 32-35°C

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/677706v1_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@70b72dorg.highwire.dtl.DTLVardef@9fbfeorg.highwire.dtl.DTLVardef@17f05a5org.highwire.dtl.DTLVardef@4b977b_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO C_FIG HighlightsWe report the first empirical human survival map across wet-bulb temperatures of 32-35{degrees}C. HydraAon and behavior cooling extend survival beyond the 6-hour theoreAcal limit. Females consistently tolerate extreme humid heat longer than males in controlled trials. Our findings refine heat-health models and inform equitable climate adaptaAon strategies. In briefExtreme humid heat, measured by wet-bulb temperature (Tw), is nearing the threshold of human survivability under climate change. In controlled trials with 36 adults, we provide the first empirical survival map across Tw=32-35{degrees}C, showing that hydration and behavioral adaptations extend endurance beyond the theoretical 6-hour limit, with women tolerating heat longer than men. These findings refine human heat tolerance thresholds and inform equitable climate adaptation and heat-health strategies. SCIENCE FOR SOCIETYAs climate change accelerates, extreme humid heat events--measured by wet-bulb temperature (Tw)--pose a growing threat to human survival. This study provides the first empirical benchmarks of human survivability across Tw=32-35{degrees}C, clarifying how long healthy adults can endure these conditions under full hydration and minimal activity. We find that while the theoretical 6-hour survival limit at Tw=35{degrees}C is partly supported, both men and women can withstand these conditions for 7-8 hours when hydrated, with women demonstrating greater tolerance. At Tw=34{degrees}C, projected heatstroke onset extends beyond 12-16 hours, while conditions at Tw=32-33{degrees}C remain largely compensable for more than 30 hours. These results challenge prevailing heat tolerance models, underscore the role of behavioral and physiological adaptations, and provide critical data for designing equitable heat-health policies and early warning systems that account for sex differences and vulnerable populations. Climate change-driven extreme heat events increasingly threaten human health. Here, we provide the first empirical map of human survivability across four extreme wet-bulb temperatures (Tw=32- 35{degrees}C) in controlled trials. Thirty-six healthy young adults (20 males, 16 females) were exposed to twelve dry-bulb temperature and relative humidity combinations in shaded indoor settings under full hydration. Participants remained seated and performed light office tasks until core temperature (Tcore) reached 39{degrees}C or exposures lasted 8 hours. Clinical heatstroke threshold (Tcore=40.5{degrees}C) were projected by extrapolating from our direct Tcore measurements during controlled exposure trials. At Tw=35{degrees}C, males and females reached heatstroke in approximately 7.1-7.7 and 8.3-8.6 hours, respectively--partly supporting the 6-hour theoretical limit while revealing enhanced short-duration resilience when hydrated. At Tw=34{degrees}C, projected times are 12.5-12.9 hours (males) and 15.5-16.2 hours (females). Conditions at Tw=32{degrees}C and 33{degrees}C remained (quasi-)compensable, with projected heatstroke onset after 33-35 hours. Females consistently tolerated heat stress longer than males. These physiological benchmarks are critical for improving heat-health models and embedding sex-specific vulnerability in equitable adaptation strategies amid accelerating global warming.

physiology↗

bioRxiv2025

Cardiomyopathy-Associated and Basic Residue Mutations in Myopalladin Alter Actin Binding, Bundling, and Structural Stability

BACKGROUNDMyopalladin (MYPN) is a striated muscle-specific sarcomeric protein belonging to the immunoglobulin (Ig)-domain-containing family of actin regulators. The Ig3 domain of MYPN is indispensable for interacting with and stabilizing actin filaments, and an increasing number of cardiomyopathic (CM)-associated mutations have been identified within this domain, highlighting its importance for normal cardiac function. Despite their clinical significance, the molecular basis of how these variants disrupt MYPN activity remains poorly understood. METHODSTo identify actin-binding sites, we introduced charge-neutralizing alanine substitutions at basic residues in the Ig3 domain and assessed their impact using actin co-sedimentation assays. CM mutations were similarly evaluated for their effect on actin interaction. Wild-type (WT) and mutant MYPN were expressed in Drosophila cardiomyocytes and body wall muscles to assess localization and interaction with actin. The impact of these mutations on protein secondary structure and stability was determined by circular dichroism spectroscopy. In addition, the WT Ig3 domain was further analyzed in co-sedimentation and co-polymerization assays to quantify polymerization of monomeric actin, subsequent bundling and to assess how calcium and salt concentrations modulate MYPN Ig3-actin interactions. RESULTSMYPN-actin binding is electrostatically driven and mediated by conserved basic charge clusters within its Ig3 domain, a mechanism common among many actin-binding proteins. CM mutations disrupted the MYPN-actin interaction, leading to significant loss of actin bundling without broadly altering secondary structure, except in the destabilized P961L variant. Immunostaining and green fluorescent protein (GFP)-tagged constructs revealed that R955W and P961L mutants accumulated into clusters at the Z-discs. WT Ig3 domain assays further showed that MYPN promotes actin polymerization and crosslinking even under non-polymerizing conditions, and actin-binding activity was not modulated by calcium. CONCLUSIONSThese findings demonstrate that CM-linked MYPN Ig3 mutations compromise actin binding and organization, suggesting that disruption of the MYPN-actin interface is a key pathogenic mechanism in cardiomyopathy. What Is Known?O_LIMyopalladin (MYPN) is a striated muscle-specific sarcomeric protein. C_LIO_LIMYPN plays an important role in controlling skeletal muscle growth by regulating actin dynamics and modulating stress-responsive signal transduction and gene expression. C_LIO_LIMutations in MYPN are linked to various inherited cardiomyopathies (CMs), including dilated, hypertrophic, restrictive, and left ventricular noncompaction forms. C_LI What new information does this article contribute?O_LITwo conserved clusters of basic residues on the surface of Ig3 domain were identified as key mediators of actin binding and bundling. C_LIO_LISeveral cardiomyopathy (CM)-associated MYPN mutations localize to this region, where they impair actin interactions and nearly abolish actin bundling activity. C_LIO_LIMutants such as R955W and P961L mislocalize abnormally in Drosophila cardiomyocytes and body wall muscles. C_LI Novelty and SignificanceThe importance of the MYPN gene, which encodes the sarcomeric protein myopalladin (MYPN), was highlighted when 66 mutations in this gene were linked to cardiomyopathies (CMs). MYPN bundles and stabilizes actin filaments and is essential for the organization and maintenance of Z-disc integrity. The C-terminal immunoglobulin (Ig) domains mediate actin interactions, with the Ig3 domain representing the minimal actin-binding unit. Notably, many disease-associated mutations cluster within this actin-binding region and give rise to diverse clinical phenotypes. Despite its clinical relevance, the molecular mechanisms by which different MYPN mutations lead to CM remain poorly understood. Prior mechanistic studies focused mainly on mutations within the cardiac ankyrin repeat protein (CARP) interaction domain of MYPN, which disrupt signaling and nuclear shuttling. In contrast, our study addresses mutations localized to the actin-binding region of MYPN. The effects of these mutations on MYPN-actin interactions had not been previously explored. We show that CM-associated mutations within the Ig3 domain disrupt MYPN-actin interactions, reduce or abolish actin bundling, and induce abnormal subcellular localization patterns in Drosophila cardiac and skeletal muscles. This study provides the first direct evidence that disruption of MYPN-actin interactions represents a distinct and clinically relevant mechanism of cardiomyopathy.

biochemistry↗

bioRxiv2025

Transcranial Alternating Current Stimulation can disrupt or reestablish neural entrainment in a primate model of Parkinson's disease

Transcranial alternating current stimulation (tACS) is a non-invasive brain stimulation method which can affect brain oscillations by inducing neuronal entrainment through modulation of spike timings. tACS has high potential for clinical applications in many neurological disorders in which the stimulation can be delivered to modulate oscillatory activity and disrupt pathological brain oscillations. For instance, in Parkinsons Disease (PD), electrophysiological activity in the motor network often exhibits excessive and hyper-synchronized beta oscillations. However, the development of tACS as a therapeutic intervention for pathological oscillations requires the prior establishment of physiologically effective stimulation parameters. We recorded neuronal activity in the motor cortical area of three parkinsonian non-human primates and examined the influence of tACS-induced electric fields on neural firing patterns. We found that weak extracellular electric fields first disrupt beta-band spike timing patterns by changing the preferred spiking phase of neurons but eventually reestablish neural entrainment with altered phase preferences when electric fields are high. Additionally, we show that frequency-matched stimulation, when stimulation frequency corresponds to endogenous oscillatory activity, significantly enhances neural entrainment. Thus, tACS exhibits significant potential for controlling and modulating pathological oscillatory patterns in many neurological disorders such as PD.

neuroscience↗

bioRxiv2025

Structure of E. coli Twin-arginine translocase (Tat) complex with bound cargo

How the twin-arginine translocase (Tat) system transports fully folded substrate proteins across cellular membranes without disrupting membrane integrity has been a fundamental question in cell biology for decades. The Tat system recognizes cargo signal peptide via a conserved twin-arginine motif and is found in prokaryotes and plant organelles. Multi-subunit Tat complex facilitates proton motive force-dependent translocation process, yet its overall architecture remains unknown. Here, we present an atomic cryo-EM structure of a E. coli trimeric TatB3C3 complex bound to the substrate SufI. The complex adopts an unusual wide-open, bowl-shaped architecture with a polar inner cavity. Unexpectedly, the cargo is engaged in a dual-contact mode: while the signal peptide binds inside one TatBC unit, the folded domain docks tightly onto an adjacent unit. The structure offers a mechanistic framework for substrate engagement and translocation by the Tat system, suggesting a direct involvement of the entire Tat complex in substrate translocation.

biochemistry↗

bioRxiv2025

Resolving the estrogen paradox in hereditary retinal degeneration: Esr1 activation suppresses Tnf-α signaling as a photoreceptor self-protection mechanism

Retinitis pigmentosa (RP) is an inherited retinal degenerative disorder characterized by progressive photoreceptor loss and irreversible blindness. Increasing evidence implicates neuroinflammation as a contributor to photoreceptor degeneration extending beyond the initial genetic insult. Although estrogen has been reported to exert anti-inflammatory effects in the central nervous system, its role in RP remains controversial, with some studies suggesting a paradoxical exacerbation of retinal pathology. To address this discrepancy, we identify estrogen receptor alpha (Esr1) as a central immunoregulatory hub in RP. Transcriptomic analyses of rd1 and rd10 revealed upregulation of estrogen-responsive and inflammatory pathways, with Esr1 expression markedly elevated during degeneration. TUNEL assays demonstrated that systemic estradiol (E2) exerted divergent effects, protective in rd1 yet deleterious in rd10, whereas selective pharmacological activation of Esr1 with propyl pyrazole triol (PPT) consistently reduced photoreceptor death, preserved dark-adapted ERG responses, and downregulated inflammatory mediators including Tnf-, Cx3cl1/Cx3cr1, Cd68, and Iba1. Mechanistically, Esr1 activation repressed microglial Tnf transcription and disrupted a self-sustaining Cx3cl1/Cx3cr1-Tnf- signaling loop driving microglial recruitment, activation and neurotoxicity in the outer nuclear layer (ONL). Targeted interventions confirmed tumor necrosis factor receptor 1 (Tnfr1) as the principal mediator of Tnf-induced photoreceptor death: selective inhibition with R7050 conferred superior protection compared with broad-spectrum Tnf- inhibitors (etanercept, infliximab). Cx3cr1 blockade likewise suppressed microglial activation and improved visual outcomes. Collectively, our findings establish Esr1 activation as not merely an external intervention but the amplification of an intrinsic self-protective program, positioning Esr1, Tnfr1, and Cx3cr1 as actionable therapeutic targets to suppress neuroinflammation and preserve vision in RP and related retinal disorders.

neuroscience↗

bioRxiv2025

TREM2 macrophages are associated with enhanced response to PD-1 blockade in human hepatocellular carcinoma

Macrophages are known to dampen tumor immunity. However, identifying druggable targets that modulate these cells to improve existing immunotherapies has been limited by a dearth of studies identifying macrophages that associate with pathological response to immune checkpoint blockade. To fulfill this unmet clinical need, we leveraged transcriptional and spatial profiling of specimens collected from a Phase II clinical trial studying neoadjuvant PD-1 blockade in patients with hepatocellular carcinoma (HCC). We determined that the intratumoral abundance of TREM2-expressing macrophages and serological levels of soluble TREM2 are elevated in patients who responded to PD-1 blockade, compared to non-responders. We validated these findings in a second HCC cohort and in the IMbrave150 trial. These highlight the robust potential for TREM2 macrophages to predict therapeutic responses of HCC to immunotherapy. Therefore, our study provides a novel basis for the use of TREM2 macrophages to strategize treatment for patients with HCC to maximize therapeutic benefit.

immunology↗

bioRxiv2025

Simulation of evoked responses to transcranial magnetic stimulation using a multiscale cortical circuit model

BackgroundTranscranial magnetic stimulation (TMS) is a widely used non-invasive brain stimulation technique, but the neural circuits activated by TMS remain poorly understood. Previous modeling approaches have been limited to either simplified point-neuron networks or isolated single-cell models that lack synaptic connectivity. ObjectiveTo develop and validate a multiscale cortical circuit model that integrates morphologically-realistic neurons with accurate TMS-induced electric field distributions and to investigate mechanisms underlying cortical responses to stimulation. MethodsWe constructed a network model of a cortical column comprising 10,000 biophysically realistic neurons (excitatory pyramidal cells and inhibitory interneurons) across layers 2/3, 5, and 6 with over 10 million synaptic connections. The model incorporated thalamic and non-specific corticocortical inputs to generate physiological firing rates. TMS-induced electric fields were calculated using finite element modeling and coupled to individual neurons through the extracellular mechanism. We validated model predictions against experimental recordings of TMS-evoked local field potentials (LFPs) and multiunit activity. ResultsThe model reproduced key experimental observations including the dose-dependent N50 LFP component and multiphasic multi-unit responses, consisting of an (excitatory) increase followed by a (inhibitory) decrease in firing rates. The early excitatory response exhibited dual-peak dynamics reflecting distinct contributions from directly and indirectly activated neuronal populations, and the subsequent inhibitory phase reflected activation of feedback GABAergic circuits through both GABAA and GABAB conductances. Spatial analysis across 30 cortical columns distributed across the precentral gyrus revealed orientation-dependent evoked responses. ConclusionThis validated multiscale model provides mechanistic insights into TMS-evoked cortical dynamics, demonstrating how direct neuronal activation cascades through synaptic networks to generate characteristic population responses. The framework establishes a computational platform for optimizing stimulation protocols in research and clinical applications. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=124 SRC="FIGDIR/small/673790v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@1028145org.highwire.dtl.DTLVardef@1751714org.highwire.dtl.DTLVardef@3e2c06org.highwire.dtl.DTLVardef@1bc756e_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2025

MAESTRO uncovers tandem paralog dynamics as a core driver of fungal stress adaptation

Understanding how fungi adapt to diverse stresses is critical for mitigating emerging drug resistance and harnessing their robustness for biotechnology. Pichia kudriavzevii is a stress-tolerant and intrinsically drug-resistant yeast, with dual industrial and clinical importance. We analyzed 170 strains using MAESTRO, a machine-learning-assisted GWAS pipeline optimized for small cohorts. MAESTRO identified biologically meaningful features and revealed that copy number variation (CNV) of tandem paralogs (TPs) is a core mechanism of multi-stress adaptation. TPs were recurrently linked to tolerance of industrial inhibitors (HMF, phenolics, heat) and antifungal drugs (fluconazole, azoles), and deletion of the TP pair gene4260/gene4261 confirmed pleiotropic effects across stresses. These findings support a TP CNV model where recombination-driven TP CNVs and gene fusions enable rapid stress adaptation. Importantly, our results suggest that antifungal resistance can arise through co-option of mechanisms originally evolved for environmental stressors, raising a One Health concern about the environmental origins of drug-resistant pathogens.

genomics↗

bioRxiv2025

High-resolution spatial analysis reveals pregnenolone metabolism as a potential target for immuno-suppressive microenvironment of BRAF mutant colorectal cancer

BRAF mutant colorectal cancer (CRC) is highly aggressive and often resistant to traditional therapies, presenting a significant challenge in cancer immunotherapy. Understanding the tumor microenvironment (TME) and identifying novel immune checkpoints are crucial for improving treatment outcomes. Here, we employed high-resolution spatial analysis and single-cell RNA sequencing to map the TME of BRAF mutant CRC at single-cell resolution, focusing on the interactions between exhausted CD8+ T cells and C1QC+ macrophages. Mechanistically, we discovered that C1QC+ macrophages drive CD8+ T cell exhaustion through pregnenolone synthesis via the steroidogenic enzyme CYP11A1. Our findings identify pregnenolone metabolism as a previously unrecognized immunosuppressive pathway in BRAF mutant CRC, and suggest that steroid biosynthesis is a generalizable mechanism of immune evasion beyond endocrine-related malignancies. Targeting this pathway offers a promising therapeutic strategy for enhancing anti-tumor immunity in BRAF mutant CRC and potentially other cancers.

cancer biology↗

bioRxiv2025

Aminopeptidase N is a receptor for hedgehog merbecoviruses

Merbecoviruses, closely related to the highly pathogenetic Middle East Respiratory Syndrome Coronavirus (MERS-CoV), circulate in hedgehogs throughout Europe and Asia, raising concerns about zoonotic transmission to humans and domestic animals. Unfortunately, how these viruses enter host cells remains unknown, hindering experimental studies. Here, we tested known coronavirus receptor orthologues from European hedgehogs (Erinaceus europaeus) and identified Aminopeptidase N (APN) as an entry receptor for hedgehog merbecoviruses. We confirm this result with single-cycle pseudotype and replication-competent virus experiments as well as protein binding assays. A screen of 30 mammalian APN orthologues reveals restricted cross-species receptor use. Cryo-electron microscopy analysis of the viral glycoprotein-receptor complex shows a unique interface distinct from known coronavirus spike:APN interactions, providing a molecular basis for species barriers. These findings expand the known range of receptor use not only within merbecoviruses but also betacoronaviruses, improving our understanding of betacoronavirus receptors, and informing risk assessments for viral emergence.

microbiology↗

bioRxiv2025

Orthogonal and Robust Analytics Enable Reproducible and Scalable Manufacturing of High Purity Extracellular Vesicles Derived from Mesenchymal Stromal Cells

The development of extracellular vesicle (EV) based therapeutics requires robust analytical assays and scalable downstream processing (DSP) strategies to ensure product quality and reproducibility. In this study, we established an analytical toolbox comprising scattering and fluorescence mode nanoparticle tracking analysis (NTA), fluorescence-based flow cytometry (Fl-FC), and multi-detector analytical size exclusion chromatography. Liposomes, selected for their physicochemical similarity to EVs, were used as reference materials to optimize assay parameters, fluorescence labeling conditions, and dynamic range, minimizing artifacts such as photobleaching and masking effects. Using these optimized analytical tools, we designed a scalable DSP workflow for human bone marrow mesenchymal stromal cell (hBM-MSC) derived EVs, incorporating clarification, tangential flow filtration (TFF), ion exchange chromatography (IEX), buffer exchange, and sterile filtration. IEX chromatography resulted in the elution of two cell-secreted populations, with similar scattering signals while eluate 1 showed approximately 30 times higher absorbance signal compared to eluate 2. Transmission electron microscopy revealed that eluate 1 contained non-vesicular extracellular particles (NVEPs), and eluate 2 was enriched in EVs and showed higher expression of positive markers such as CD81 and CD73 using Simple Western. Additionally, the two IEX eluates showed different proteomic and lipidomic profiles. Then the analytical toolbox was utilized to monitor the DSP process in terms of particle recovery and impurity removal throughout the process determining the high purity level of the final EV preparation. Together, these results demonstrate that orthogonal analytics coupled to a scalable DSP yield reproducible MSC-EV preparations while depleting commonly co-isolated NVEPs. This practical framework advances process analytics of MSC-EV manufacturing and supports reporting of identity, purity, and function aligned with existing guidelines in the field. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=91 SRC="FIGDIR/small/672476v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@142ff8aorg.highwire.dtl.DTLVardef@196c0bforg.highwire.dtl.DTLVardef@485c36org.highwire.dtl.DTLVardef@9dff39_HPS_FORMAT_FIGEXP M_FIG C_FIG Liposomes were used as reference materials to optimize single particle analytics as well as multi detector analytical chromatography to assess particle recovery and impurity removal. This optimized analytical toolbox enabled accurate monitoring of downstream processing steps, establishing a robust workflow for reproducible and scalable mesenchymal stromal derived extracellular vesicles bioprocessing.

bioengineering↗

bioRxiv2025

Quantitative Analysis of Meloidogyne incognita Population Density Using Real-Time PCR and Its Correlation with Root-Knot Disease Index in Tabacco (Nicotiana tabacum)

Tobacco root-knot disease represents a significant threat to tobacco production, particularly in the western Henan region, where Meloidogyne incognita is the predominant species. This study collected samples of M. incognita and soil from Luoyang, Henan, and designed specific primers MiF and MiR based on the amplified 735 bp sequence of the ITS1-5.8S-ITS2 region. These primers exhibit mismatches with related species, including M. javanica, M. minor, M. hapla, and M. arenaria, showing 1, 6, 10, and 10 base differences in the forward direction, respectively, 3, 10, and 9 base mismatches in the reverse direction. Although the primers were used to detect corn-wheat soil samples, no amplification was observed. Additionally a real-time quantitative PCR curve for M. incognita in soil was constructed, revealing a negative correlation between the Ct value (Y) and the log-transformed number of nematodes (x) per 20 g of dry soil, represented by the equation y = -0.9757x + 35.565; (R2 = 0.9999, P < 0.01). According to the decomposition efficiency of nematode DNA in soil, the results showed that nematode DNA degrades rapidly in soil, with a degradation rate of approximately 87.3% at 3 days and 99.97% at 14 days. Furthermore, significant differences were observed in the real-time PCR detection efficiency among various nematode forms: the Ct value of J1 was significantly higher than that of J2, while abnormal eggs (empty eggs or internal bubble eggs) exhibited the highest Ct value. The proportion of abnormal eggs in the soil before planting was significantly higher at 64.08% compared to only 15.3% at harvest, indicating that the activity of nematodes in the soil is significantly reduced after harvesting in October and planting in March of the following year. A survey of 126 tobacco plants indicated a significant positive correlation between the root-knot index (RKI) and root-knot nematode density (r = 0.80, p < 0.01). The study identified a minimum disease threshold of 234 individuals / 20 g soil at harvest and revealed a nonlinear relationship between disease severity and nematode density. Specifically, a weak correlation was observed at low density (Low RKI: 0-1; nematode density<2000 individuals/20 g soil, r = 0.49, P<0.05), while a significant correlation was noted at moderate density (High RKI: nematode density>2000 individuals/20 g soil, r = 0.69, P<0.05). This study showed that as the RKI increases, the rate of increase in nematode density in the soil diminishes. These findings provide valuable insights for the development of effective scientific strategies for nematode control.

plant biology↗

bioRxiv2025

Cytoplasmic dynamics are overlooked in single nuclei RNA-seq but can be rescued by CytoRescue, a generative AI model to recover cytoplasm enriched gene

Single-nucleus RNA sequencing (snRNA-seq) generates single cell data from nuclei. It provides valuable compatibility with frozen or difficult-to-dissociate tissues while avoiding stress responses in fresh samples. However, cytoplasmic depletion inherently limits quantification of cytoplasm-enriched genes. Here, we present CytoRescue, a novel generative AI model designed to recover attenuated cytoplasmic signals in snRNA-seq data. Our results demonstrate that CytoRescue effectively restores expression of cytoplasm-enriched genes while preserving underlying gene expression signatures. Taking advantaging of the raw-in-raw-out design, CytoRescue can be easily integrated into the existing pipelines for single-cell sequencing analysis. Notably, CytoRescue successfully recovers EGF signaling pathway components, a critical cell-cell communication pathway in lung cancer, in an independent dataset. CytoRescue addresses a fundamental limitation of snRNA-seq technology, enhancing its utility for comprehensive transcriptomic profiling while maintaining the advantages of single nucleus-based approaches.

bioinformatics↗

bioRxiv2025

Temporal Clonal Tracing and Functional Perturbation Reveal Niche-Adaptive and Tumor-Intrinsic IFNγ Dependencies Driving Ovarian Cancer Metastasis

Metastasis is an emergent continuum driven by evolving reciprocal adaptations between disseminating tumor cells (DTCs) and specialized niches of different organs. The interplay between intrinsic and niche-driven mechanisms that enable DTCs to survive and home to distant organs in peritoneal ovarian cancer metastasis remains incompletely understood. Here, we present MetTag, a single-cell barcoding and transcriptome profiling approach with single cell clonality barcodes and time-stamped batch identifiers (BC.IDs) to resolve metastasis clonality and temporal dynamics of DTC colonization. Deep sequencing of MetTag barcodes revealed enrichment of early-disseminated clones across metastatic sites, and targeted depletion of pioneer DTCs diminished the outgrowth of subsequent arriving DTCs. Subsequent MetTag-coupled single cell RNA sequencing (scRNA-seq) on ascites and metastasis-bearing omenta revealed a distinct interferon-gamma (IFN{gamma})-centric transcriptional trajectory selectively enriched among pioneer clones. In vivo CRISPR/Cas9 screening of niche-specific signatures demonstrated that the tumor-intrinsic IFN{gamma} response is functionally required for peritoneal metastasis. Knockout of the IFN{gamma} receptor 1 (Ifngr1) in the initial pioneer DTCs significantly reduced total metastatic burden, revealing a critical window of time in which IFN{gamma} signaling shapes the post-seeding metastatic niche (PSMN) and subsequent metastatic evolution. Mechanistically, the tumor-intrinsic IFN{gamma} response induced Poly(ADP-Ribose) polymerase family member 14 (Parp14) and peritoneal macrophages cooperatively shield DTCs from anoikis by promoting pro-survival signaling. Our study defines the temporal clonal architecture of peritoneal niches and reveals a "first come, first served" adaptation principle, where pioneer colonizer fitness determines the success of subsequent colonizers.

cancer biology↗

bioRxiv2025

Structural assembly of maize CRY-GL2 photosignaling complex provides insights into its regulatory role in cuticular wax biosynthesis

Cryptochromes (CRYs) are blue-light photoreceptors in plants that regulate diverse physiological processes through oligomerization-dependent interactions with downstream effectors. While numerous light-inducible CRY-binding proteins have been identified, the structural basis for photoactivated CRY-effector assembly remains poorly understood. Here we report the crystal structure of a constitutively active maize CRY1c photolyase homology region (ZmCRY1c-PHRW368A) in complex with ZmGL2, a BAHD ptotein directing very-long-chain fatty acid (VLCFA) elongation for cuticular wax biosynthesis. Structural analysis reveals that light-activated ZmCRY1c forms a homotetrameric scaffold generating four symmetric binding sites at subunit interfaces. Each ZmCRY1c protomer engages one ZmGL2 molecule through conformational reorganization of its 16-17 loop and 17 helix, establishing a 4:4 hetero-octameric photosignaling complex validated by mutational profiling and optical design. Remarkably, structural alignment with the ZmCER6-ZmGL2 complex reveals substantial overlap (78%) in ZmGL2 binding interfaces. Biochemical quantification demonstrates that ZmCRY1c suppresses ZmCER6-GL2 enzyme activity in a dose dependent manner, unveiling a light-dependent regulatory switch controlling VLCFA elongation efficiency, though in vivo physiological investigation is awaited. Collectively, our work not only establishes the atomic model of light-activated plant CRY-effector assembly, but also uncovers a previously unrecognized spatial competition between photoreceptor and metabolic enzyme complexes that might function as a photoregulatory paradigm in cuticular wax biosynthesis.

plant biology↗

bioRxiv2025

ZmCRY1s interact with GL2 in a blue light dependent manner to regulate epidermal wax composition in Zea mays

Cryptochromes (CRYs) are photolyase-like blue-light / ultraviolet-A (UV-A) receptors that regulate diverse aspects of plant growth. Maize (Zea mays), a major crop often grown under high UV-B radiation, possesses four copies of CRY. However, it remains unclear whether the multiple copies of CRY in maize have evolved to improve UV tolerance or to acquire new functions. In this study, CRISPR-Cas9-engineered Zmcry mutants were used to investigate the functions of four cryptochromes (ZmCRYs) in maize. The findings revealed that ZmCRYs play a redundant role in mediating blue light signaling and in inhibiting the elongation of the mesocotyl. The results also demonstrated that ZmCRYs mediated blue light-enhanced UV-B stress tolerance in Zea mays by upregulating the expression of genes involved in UV-B stress tolerance-related metabolites such as phenylpropanoid, flavonoid, and fatty acid biosynthesis. Furthermore, blue light was found to influence both the accumulation and composition of epidermal waxes, suggesting that blue light enhances epidermal wax accumulation for UV-B stress tolerance. Additionally, it was discovered that ZmCRY1 directly interacted with GLOSSY2 (GL2) in a blue light dependent manner to mediate blue light promoted C32 aldehyde accumulation, shedding new light on the enigma of aldehyde-forming. These results highlight the critical roles of ZmCRY1s in mediating blue light regulated epidermal wax biosynthesis and UV-B tolerance in Zea mays.

plant biology↗

bioRxiv2025

Accurate detection and quantification of single-base m6A RNA modification using nanopore signals with multi-view deep learning

N6-methyladenosine (m6A) is a crucial epitranscriptomic mark. While Nanopore Direct RNA Sequencing (DRS) enables transcriptome-wide detection, most existing methods neglect or lack the capacity to effectively process the intrinsically variable-length raw signals generated by DRS reads.Here, we present MultiNano, a multi-view deep learning framework that converts variable-length raw signals into image-like feature representations, effectively resolving the length inconsistency problem. By integrating raw signal and basecalling features, MultiNano enables accurate and comprehensive transcriptome-wide detection of m6A modifications. Our model achieved state-of-the-art (SOTA) performance in various tasks, including site-level prediction, read-level prediction, cross-species transfer learning, and modification rate estimation. Furthermore, the false positive control strategy implemented in MultiNano significantly enhances the models robustness and predictive accuracy, offering a powerful alternative to traditional thresholding-based filtering algorithms. Collectively, our approach provides novel insights for the absolute quantification and single-base resolution of RNA modifications.

bioinformatics↗

bioRxiv2025

TRPM7 Annexin A1 Mechanosensitive Pathway Drives Capillary Infiltration by Circulating Tumor Cells

Successful metastatic dissemination requires tumor cells to overcome significant physical barriers. When circulating tumor cells (CTCs) become lodged in capillary beds, studies reveal that only a small subset can adapt to fluid shear stress (FSS) within the constricted vasculature. These rare, mechanically resilient cells may subsequently extravasate and form metastatic lesions, but details on this adaptation of the physical environment and its molecular basis are not well understood. Utilizing a microfluidics platform that mimics microcirculatory dynamics, we discover that only breast cancer cells with high metastatic potential maintain directional migration under physiologically relevant FSS conditions. This highlights the mechanical selection process of metastatic precursors during hematogenous dissemination. We identify the TRPM7-Annexin A1-actin signaling pathway as essential for overcoming physical barriers and regulating cell motility under capillary FSS. FSS prompts cytoskeletal reorganization and actin disassembly, restricting cell movement. Cancer cells respond to this by increasing mechanical loading, activating TRPM7 and triggering calcium influx, which then activates Annexin A1. This calcium-dependent protein, Annexin A1, interacts with the actin cortex to prevent FSS-induced actin disassembly, thus aiding migration. Experiments conducted in mouse liver capillaries validate the critical role of this pathway in cancer cell motility. Additionally, we propose a mechano-pharmacological strategy using FTY720 to target the TRPM7 pathway, highlighting its therapeutic potential to modify CTC receptor specificity and inhibit distant metastasis.

cancer biology↗