Biology subjects

Zhao, M.

Publications and source records attributed to Zhao, M..

At least 91 records · Page 5Linked to original sources

bioRxiv2025

Infections in the Atlantic Blue Crab Callinectes sapidus (Rathbun, 1896) in coastal Adriatic and Aegean sea, and Atlantic Iberian coast

In the Mediterranean Sea, the abundance of the invasive portunid crab, Callinectes sapidus, has dramatically increased in recent years. This raises concerns about damage to ecosystems, but also offers opportunities for exploitation of a new fishery. Newly invasive species may escape from pathogens in their native range, may introduce new pathogens, or can become host to endemic pathogens. Understanding these factors is important for predicting or managing natural resources in the invaded range. This study investigated the prevalence of two pathogens common in C. sapidus in its home range of North America: the reovirus CsRV1 and the protozoan parasite Hematodinium perezi. In crabs collected from Aegean, Adriatic, and Atlantic waters, the CsRV1 virus was not detected. In contrast, the parasite H. perezi was found in crabs from all areas except the Aegean Sea. Sequence analysis of the H. perezi ITS1 gene indicated that the strains observed are most related to genotypes already described in Europe and the Mediterranean, and not to strains from the Americas or Asia. The arrival of new species and new potential pathogens is ongoing through transfer of ballast water to the Atlantic and Mediterranean. Although systems are in place to exchange or inactivate ballast water, it is advisable to continue and expand surveillance for pathogens in introduced species, to inform management of movement of these species between regions.

ecology↗

bioRxiv2025

EPSD 2.0: An Updated Database of Protein Phosphorylation Sites across Eukaryotic Species

As one of the most crucial post-translational modifications (PTMs), protein phosphorylation regulates a broad range of biological processes in eukaryotes. Biocuration, integration and annotation of reported phosphorylation events will deliver a valuable resource for the community. Here, we present an updated database, the eukaryotic phosphorylation site database 2.0 (EPSD 2.0), which includes 2,769,163 experimentally identified phosphorylation sites (p-sites) in 362,707 phosphoproteins from 223 eukaryotes. From the literature, 873,718 new p-sites identified through high-throughput phosphoproteomic research were first collected, and 1,078,888 original phosphopeptides together with primary references were reserved. Then, this dataset was merged into EPSD 1.0, comprising 1,616,804 p-sites within 209,326 proteins across 68 eukaryotic organisms [1]. We also integrated 362,190 additional known p-sites from 10 public databases. After redundancy clearance, we manually re-checked each p-site and annotated 88,074 functional events for 32,762 p-sites, covering 58 types of downstream effects on phosphoproteins, and regulatory impacts on 107 biological processes. In addition, phosphoproteins and p-sites in 8 model organisms were meticulously annotated utilizing information supplied by 100 external platforms encompassing 15 areas. These areas included kinase/phosphatase, transcription regulators, three-dimensional structures, physicochemical characteristics, genomic variations, functional descriptions, protein domains, molecular interactions, drug-target associations, disease-related data, orthologs, transcript expression levels, proteomics, subcellular localization, and regulatory pathways. We expect that EPSD 2.0 will become a useful database supporting comprehensive studies on phosphorylation in eukaryotes. The EPSD 2.0 database is freely accessible online at https://epsd.biocuckoo.cn/.

bioinformatics↗

bioRxiv2025

NAMPT activity plays a key role in driving autoimmune processes that characterize type 1 diabetes development in mice

Type 1 diabetes (T1D) is characterised by destruction of pancreatic beta cells by islet-infiltrating cytotoxic lymphocytes, and elevated intra-islet secretion of pro-inflammatory cytokines. However, the underlying pathophysiological mechanisms remain incompletely understood. We hypothesised that abnormal elevation of islet NAD, via activation of NAMPT, plays a key role in driving islet autoimmune processes in T1D. Here, we report that NAMPT inhibition protects against pro-inflammatory cytokine (IL-1{beta}, TNF and IFN{gamma}) mediated beta-cell dysfunction and apoptosis in isolated mouse and human islets. RNAseq revealed that NAMPT inhibition blocked cytokine-mediated gene expression linked to pro-inflammatory responses and leukocyte migration. In vivo, diabetes was induced in CD1 mice via multiple low dose streptozotocin (MLDS) injection. MLDS mice were administered the NAMPT inhibitor FK866 (10 mg/kg; IP) or saline equivalent for 16 days. These experiments demonstrated that NAMPT inhibition improved glycaemic control and beta-cell function and insulin content in MLDS mice. FK866 also reduced proportions of islet-residing TNF-producing CD4+T-cells and F4/80+macrophages, proliferation of spleen-derived CD4+ and CD8+T-cells, and proliferation of islet-derived CD4+T-cells and F4/80+macrophages. Finally, we report that NAMPT inhibition was able to block pro-inflammatory cytokine-mediated migration of cytotoxic CD8+T-cells into isolated islets, using an in vitro transwell platform. This data supports a key immunomodulatory role for NAMPT in islet autoimmunity. NAMPT inhibition may represent a novel therapeutic approach for T1D. The effects of increased NAD levels on islet inflammation require in-depth characterisation, and caution should be exercised with regard to use of NAD boosting supplements, particularly in individuals at risk of developing T1D.

cell biology↗

bioRxiv2025

Dimerization and dynamics of angiotensin-I converting enzyme revealed by cryoEM and MD simulations

Angiotensin-I converting enzyme (ACE) regulates the levels of disparate bioactive peptides, notably converting angiotensin-I to angiotensin-II and degrading amyloid beta. ACE is a heavily glycosylated dimer, containing 4 analogous catalytic sites, and exists in membrane bound and soluble (sACE) forms. ACE inhibition is a frontline, FDA-approved, therapy for cardiovascular diseases yet is associated with significant side effects, including higher rates of lung cancer. To date, structural studies have been confined to individual domains or partially denatured cryo-EM structures. Here we report the cryo-EM structure of the glycosylated full sACE dimer. We resolved four structural states at 2.99 to 3.65 [A] resolution which are primarily differentiated by varying degrees of solvent accessibility to the active sites and reveal the full dimerization interface. We also employed all-atom molecular dynamics (MD) simulations and heterogeneity analysis in cryoSPARC, cryoDRGN, and RECOVAR to elucidate the conformational dynamics of sACE and identify key regions mediating conformational change. We identify differences in the mechanisms governing the conformational dynamics of individual domains that have implications for the design of domain-specific sACE modulators.

biophysics↗

bioRxiv2025

Permissive central tolerance plus defective peripheral checkpoints licence pathogenic memory B cells in CASPR2-antibody encephalitis

Autoimmunity affects 10% of the population. Within this umbrella, autoantibody-mediated diseases targeting one autoantigen provide a unique opportunity to comprehensively understand the developmental pathway of disease-causing B cells and autoantibodies. While such autoreactivities are believed to be generated during germinal centre reactions, the roles of earlier immune checkpoints in autoantigen-specific B cell tolerance are poorly understood. We address this concept in patients with CASPR2-autoantibody encephalitis and healthy controls. In both groups, comparable and high ([~]0.5%) frequencies of unmutated CASPR2-reactive naive B cells were identified. By contrast, CASPR2-reactive memory B cells were exclusive to patients, and their B cell receptors demonstrated affinity-enhancing somatic mutations with heterogenous binding kinetics. These effector molecules possessed epitope-dependent pathogenic effects in vitro neuronal cultures and in vivo. The unmutated common ancestors of these memory B cells showed a distinctive balance between strong CASPR2 reactivity and very limited binding across the remaining human proteome. Our results are the first to propose mechanisms underlying autoantigen-specific tolerance in humans. We identify permissive central tolerance, defective peripheral tolerance and heterogenous autoantibody binding properties as sequential pathogenic steps which licence CASPR2-directed pathology. By leveraging the basic immunobiology, we rationally direct tolerance-restoring approaches in CASPR2-antibody diseases. This paradigm is applicable across autoimmune conditions.

immunology↗

bioRxiv2025

Competing signaling pathways controls electrotaxis

Understanding how cells migrate following exogenous cues is one of the most fundamental questions for biology, medicine, and bioengineering. Growing evidence shows that electrotaxis, the directed cell migration toward electric potential gradients, represents a precise and programmable method to control cell migration. Most data suggest that the polarization of membrane components and the following downstream signaling are central to electrotaxis. Unfortunately, how these multiple mechanisms coordinate with the motile machinery of the cell to respond to an electric field is still poorly understood. Here, we develop a mechanistic model that explains and recapitulate electrotaxis across different cell types. Using the zebrafish proteome, we identify membrane proteins directly related to migration signaling pathways that polarize anodally and cathodally. Further, we show that simultaneous and asymmetric distribution of charged membrane receptors towards the anode and the cathode establishes multiple cooperative and competing stimuli for downstream signaling pathways. The resulting polarization of signals controls the actomyosin network dynamics, directing the anodal and cathodal migration of the cell. Our theoretical framework rationalizes the physical processes that determine electrotaxis across cell types and provides a physical framework to test multiple electrotactic pathways. These results together show us how to control cell migration to, e.g., enhance, cancel, or switch directed cell migration, which opens up new avenues not only to promote tissue regeneration or arrest tumor progression but also to design better biomimetic-engineered tissue constructs.

biophysics↗

bioRxiv2025

Molecular basis for the activation of Pseudomonas aeruginosa MsbA by Zn2+

Proteins involved in the biogenesis of lipopolysaccharide (LPS), a lipid exclusive to Gram-negative bacteria, are promising candidates for drug discovery. Specifically, the ABC transporter MsbA plays a crucial role in translocating an LPS precursor from the cytoplasmic to periplasmic facing leaflet of the inner membrane, and small molecules that inhibit its function exhibit bactericidal activity. Here, we use native mass spectrometry (MS) to determine lipid binding affinities of MsbA from P. aeruginosa (PaMsbA), a Gram-negative bacteria associated with hospital-acquired infections, in different conformations. We show the ATPase activity of the transporter is stimulated by Zn2+ and successfully trapping the protein with vanadate requires Zn2+ not Mg2+, which is necessary to trap MsbA from E. coli. We also present cryogenic-electron microscopy structures of PaMsbA in occluded and open outward-facing conformations determined to a resolution of 2.98 and 2.72 angstroms, respectively. The structures reveal a triad of histidine residues and mutation of these residues abolishes Zn2+ stimulation of PaMsbA activity. Together our studies provide detailed insight into PaMsbA structure, lipid binding preferences, and uncover a mechanism through which Zn2+ promotes the dimerization of the transporter, resulting in enhanced ATPase activity.

biochemistry↗

bioRxiv2024

Afadin Sorts Different Retinal Neuron Types into Accurate Cellular Layers

Neurons use cell-adhesion molecules (CAMs) to interact with other neurons and the extracellular environment: the combination of CAMs specifies migration patterns, neuronal morphologies, and synaptic connections across diverse neuron types. Yet little is known regarding the intracellular signaling cascade mediating the CAM recognitions at the cell surface across different neuron types. In this study, we investigated the neural developmental role of Afadin1-4, a cytosolic adapter protein that connects multiple CAM families to intracellular F-actin. We introduced the conditional Afadin mutant5 to an embryonic retinal Cre, Six3-Cre6-8. We reported that the mutants lead to the scrambled retinal neuron distribution, including Bipolar Cells (BCs), Amacrine Cells (ACs), and retinal ganglion cells (RGCs), across three cellular layers of the retina. This scrambled pattern was first reported here at neuron-type resolution. Importantly, the mutants do not display deficits for BCs, ACs, or RGCs in terms of neural fate specifications or survival. Additionally, the displayed RGC types still maintain synaptic partners with putative AC types, indicating that other molecular determinants instruct synaptic choices independent of Afadin. Lastly, there is a significant decline in visual function and mis-targeting of RGC axons to incorrect zones of the superior colliculus, one of the major retinorecipient areas. Collectively, our study uncovers a unique cellular role of Afadin in sorting retinal neuron types into proper cellular layers as the structural basis for orderly visual processing.

neuroscience↗

bioRxiv2024

Non-invasive diagnosis of early chronic colitis cancerization via amplified sensing of miRNA-21 in NIR-IIb window

Early diagnosis of the colitis-associated colorectal cancer (CRC) is of great significance for improving prognosis and survival rates. However, the clinical used colonoscopy and biopsy methods are invasive and lack of sensitivity at early stage of cancerization. Herein, we present an amplified sensing strategy in the second near-infrared b (NIR-IIb, 1500-1700 nm) window for non-invasive in situ visualization of early cancerization biomarker miRNA-21. A CRC nanosensor composed of Er3+ doped lanthanide nanoparticles, DNAzyme, and IR820 dye is designed as a NIR-IIb ratiometric luminescence reporter, providing rapid feedback and high sensitive miRNA-21 detection with limit of detection (LOD) of 1.26 pM. This strategy enables a non-invasive detection of colitis-associated cancerization up to [~]4 weeks ahead of the clinically used biopsy analysis, providing a promising alternative for early diagnosis of chronic colitis cancerization and guidance for therapeutic management.

bioengineering↗

bioRxiv2024

Electron-density informed effective and reliable de novo molecular design and lead optimization with ED2Mol

Generative drug design opens new avenues for discovering novel compounds within the vast chemical space rather than conventional screening against limited compound libraries. However, the practical utility of the generated molecules is frequently constrained, as many designs prioritize a narrow range of pharmacological properties while neglecting physical reliability, which hinders the success rate of subsequent wet-lab evaluations. To address this, we propose ED2Mol, a deep learning-based approach that leverages fundamental electron density information to improve de novo molecular generation and lead optimization. The extensive evaluations across multiple benchmarks demonstrate that ED2Mol surpasses existing methods in terms of generation success rate and >97% physical reliability. It also facilitates automated lead optimization that is not fully implemented by other methods using fragment-based strategies. Furthermore, ED2Mol exhibits generalizability to more challenging, unseen allosteric pocket benchmarks, attaining consistent performance in both de novo molecule generation and lead optimization. More importantly, ED2Mol has been applied to various real-world essential targets, successfully identifying wet-lab validated bioactive compounds, ranging from FGFR3 orthosteric inhibitors to CDC42 allosteric inhibitors and GCK allosteric activators. The directly generated binding modes of these compounds with target proteins are close to predictions through molecular docking and further validated via the X-ray co-crystal structure. All these results highlight ED2Mols potential as a useful tool in realistic drug design with enhanced effectiveness, physical reliability, and practical applicability.

bioinformatics↗

bioRxiv2024

Remote-Controlled Wireless Bioelectronics for Fluoxetine Therapy to Promote Wound Healing in a Porcine Model

Wound healing presents a significant challenge in biomedical science, requiring precise therapeutic delivery and real-time monitoring. Bioelectronic systems offer a promising solution but remain largely unexplored for wound care, particularly in large animal models that reflect human healing dynamics. This study introduces a remote controlled wireless bioelectronic platform equipped with an iontophoretic pump to deliver fluoxetine, a selective serotonin reuptake inhibitor that promotes wound repair. In vitro and ex-vivo testing validated efficient on demand fluoxetine delivery. In vivo experiments in a porcine wound model demonstrated clear therapeutic efficacy over 3-day and 7-day periods. The system enhanced healing outcomes, increasing re-epithelialization by 37% (H&E staining), reducing the M1/M2 macrophage ratio by 33%, and stimulating neuronal growth at the wound site. This bioelectronic platform delivers fluoxetine in a controlled, remotely-controlled manner while allowing for wound direct wound imaging that can be used to monitor wound healing progress. Additionally, it allows precise dose and temporal delivery of treatment to enhance the outcome of future large animal wound healing studies.

bioengineering↗

bioRxiv2024

A molecularly defined brain circuit module for regulating the panic-like defensive state

Panic is an episode of strong defensive state, characterized by intense fear and severe physical symptoms such as elevated cardiorespiratory activities. How the brain generates panic state remains poorly understood. Here, we developed a robot-based experimental paradigm to evoke panic-like defensive state in mice. When stimulated by the robot, the mice exhibited jumping escapes and elevated cardiorespiratory activities. With this paradigm, we identified Cbln2-expressing (Cbln2+) neurons in the posterior hypothalamic nucleus (PHN) as a key neuronal population essential for the induction of panic-like defensive state. Activation of Cbln2+ PHN neurons induced behavioral and physical symptoms of panic-like defensive state. These neurons were strongly activated by noxious mechanical stimuli and encode jumping escape vigor. They were synaptically innervated by anxiety-associated brain areas and provoked panic-like defensive state via their projection to the periaqueductal gray. Together, our results reveal a molecularly defined circuit module that regulates the panic-like defensive state in mice.

neuroscience↗

bioRxiv2024

Wireless bioelectronic device for wound healing

Wireless bioelectronic actuators have been developed to deliver targeted treatments over multiple days while continuously monitoring delivery, thereby improving wound healing. Specifically, these devices can deliver charged biomolecules such as fluoxetine cations (Flx+) and electric field (EF) in freely moving pigs. Treatments can be controlled and monitored in real time via WiFi, with options for both user-specified delivery rates and durations, as well as automated closed-loop (CL) control. The devices are engineered to handle various failure scenarios that may arise in dynamic, real-world experiments--such as communication or power interruptions--ensuring that valuable experimental data is collected with minimal disruption. The ion pump features eight drug reservoirs and channels arranged around a 20 mm diameter-wound, with a central ground electrode (0 V). When voltages above 0 V are applied to the outer channels, currents flow from the reservoirs and channels into the wound, delivering Flx+ and/or EF depending on the reservoir solution. The device records applied voltages and currents locally to a microSD card at a high sampling rate, while simultaneously transmitting real-time measurements via a local WiFi network to a wound healing algorithm running on a nearby laptop. CL control of current/delivery rate is performed by an onboard microcontroller unit (MCU) and current-source microchips, based on instructions received from the wound healing algorithm. A graphical user interface (GUI) provides intuitive user control and real-time data visualization, with support for multiple devices. In vivo studies over seven days showed that Flx+-treated wounds had a 20% lower M1/M2 macrophage ratio and 41.67% greater re-epithelialization compared to controls (standard-of-care), demonstrating the actuators potential to enhance wound healing.

biochemistry↗

bioRxiv2024

Molecular and spatial analysis of ganglion cells on retinal flatmounts: diversity, topography, and perivascularity

Diverse retinal ganglion cells (RGCs) transmit distinct visual features from the eye to the brain. Recent studies have categorized RGCs into 45 types in mice based on transcriptomic profiles, showing strong alignment with morphological and electrophysiological properties. However, little is known about how these types are spatially arranged on the two-dimensional retinal surface--an organization that influences visual encoding--and how their local microenvironments impact development and neurodegenerative responses. To address this gap, we optimized a workflow combining imaging-based spatial transcriptomics (MERFISH) and immunohistochemical co-staining on thin flatmount retinal sections. We used computational methods to register en face somata distributions of all molecularly defined RGC types. More than 75% (34/45) of types exhibited non-uniform distributions, likely reflecting adaptations of the retinas anatomy to the animals visual environment. By analyzing the local neighborhoods of each cell, we identified perivascular RGCs located near blood vessels. Seven RGC types are enriched in the perivascular niche, including members of intrinsically photosensitive RGC (ipRGC) and direction-selective RGC (DSGC) subclasses. Orthologous human RGC counterparts of perivascular types - Melanopsin-enriched ipRGCs and ON DSGCs - were also proximal to blood vessels, suggesting their perivascularity may be evolutionarily conserved. Following optic nerve crush in mice, the perivascular M1-ipRGCs and ON DSGCs showed preferential survival, suggesting that proximity to blood vessels may render cell-extrinsic neuroprotection to RGCs through an mTOR-independent mechanism. Overall, our work offers a resource characterizing the spatial profiles of RGC types, enabling future studies of retinal development, physiology, and neurodegeneration at individual neuron type resolution across the two-dimensional space.

neuroscience↗

bioRxiv2024

A female-specific RdDM-associated element guides de novo DNA methylation during rice gametophyte development

DNA methylation is crucial for regulating gene expression and silencing transposons. While the spatiotemporal specificity of DNA methylation controls organ formation and development, the molecular mechanisms underlying locus- or tissue-specific DNA methylation remain unclear. Previously, locus- or tissue-specific DNA methylation was thought to depend solely on the recognition of histone methylations. Here we show that high expression of 24-nucleotide small interfering RNAs (24-nt siRNAs) leads to incremental CHH methylation at specific loci via the RNA-directed DNA methylation (RdDM) pathway during female gametophyte development in rice. We also identify a DNA motif, the Female-specific RdDM-associated Element (FRE), which guides methylation to these loci. Furthermore, we show that the female-preferentially expressed protein OsCLSY3 directly interacts with FRE to regulate CHH methylation specifically in female reproductive tissues. These findings uncover a novel molecular mechanism necessary for tissue- and locus-specific de novo DNA methylation that might be conserved across diverse organisms including animals.

plant biology↗

bioRxiv2024

SS-31 protects against bleomycin-induced lung injury and fibrosis

ObjectiveThe aim of this research was to investigate if the mitochondria-targeting peptide SS-31 could serve as a protective measure against bleomycin-induced pulmonary fibrosis in mice. MethodMice were split into four groups named CON group, SS-31 group, BLM group, and the BLM+ SS-31 group. SS-31 was administered daily from the day prior to the experiment for the control and model groups. Mice were euthanized after 28 days of the experiment, following which blood, bronchoalveolar lavage fluid, and lung tissue were collected for analysis. ResultsThe study demonstrated that SS-31 could potentially mitigate the reduction in mice. It was observed through HE and Masson staining, immunohistochemistry, hydroxyproline detection, and fibrosis index measurement via Western blot that SS-31 could alleviate pulmonary fibrosis caused by BLM. Electron microscopy and ATP detection further suggested that SS-31 might help protect mitochondrial structure and function. It was also found that SS-31 could reduce reactive oxygen species and myeloperoxidase, thereby alleviating the reduction of antioxidant factor MPO and SOD, as well as diminishing the inflammatory factors TNF-, IL-1 {beta}, and IL-6. ConclusionThe mitochondria-targeting drug SS-31 exhibited potential in mitigating bleomycin-induced pulmonary fibrosis, improving mitochondrial structural and functional damage, stabilizing the balance between oxidative and antioxidant systems, reducing inflammatory factor expression, and improving apoptosis in lung tissue.

pharmacology and toxicology↗

bioRxiv2024

Molecular characterization of the archaic HLA-B*73:01 allele reveals presentation of a unique peptidome and skewed engagement by KIR2DL2

HLA class I alleles of archaic origin may have been retained in modern humans because they provide immunity against diseases to which archaic humans had evolved resistance. According to this model, archaic introgressed alleles were somehow distinct from those that evolved in African populations. Here we show that HLA-B*73:01, a rare allotype with putative archaic origins, has a relatively rare peptide binding motif with an unusually long-tailed peptide length distribution. We also find that HLA-B*73:01 combines a restricted and unique peptidome with high-cell surface expression, characteristics that make it well-suited to combat one or a number of closely-related pathogens. Furthermore, a crystal structure of HLA-B*73:01 in complex with KIR2DL2 highlights differences from previously solved structures with HLA-C molecules. These molecular characteristics distinguish HLA-B*73:01 from other HLA class I alleles previously investigated and may have provided early modern human migrants that inherited this allele with a selective advantage as they colonized Europe and Asia.

biochemistry↗

bioRxiv2024

A Practical and Safe Model of Nitrogen Mustard Injury in Cornea

PurposeSulfur mustard (SM) is an alkylating agent used in warfare and terrorism that inflicts devastating ocular injuries. Although the clinical symptoms are well described, the underlying mechanisms are not fully understood, hindering the development of effective treatments. One major roadblock is the lack of a suitable model due to the extremely hazardous nature of SM, which requires strict safety measures. As a safe and practical alternative, we report a novel model that uses mechlorethamine (nitrogen mustard) gel, an FDA-approved topical chemotherapeutic administered by patients at home. Here we demonstrate its suitability to induce mustard corneal injury in any laboratory. MethodsEx vivo porcine corneas were injured with mechlorethamine gel. Hematoxylin-eosin staining, and immunohistochemistry were performed to evaluate histopathology of SM-like corneal injuries: epithelium thickness and stromal separation, keratocyte and inflammatory cell counts, and expression of inflammation and fibrosis markers. ResultsThis model showed the characteristic histopathology and expression of cyclooxygenase-2 (inflammation) and fibronectin-1 (fibrosis), which were consistent with other well-established SM-like corneal injury models. ConclusionGiven its ease of implementation and safety, this mechlorethamine model could be used to study the full course of mustard corneal injuries. This model would greatly facilitate mustard injury research, shedding light on new knowledge that would increase our understanding of mustard ocular injuries while investigating novel therapeutics. Translational relevancethis model will allow safe evaluation of SM-like corneal injuries within 24 hours, facilitating the identification of early/new molecules that might help to develop novel treatments which could be readily translated into the clinic.

pharmacology and toxicology↗