Biology subjects

Zhao, J.

Publications and source records attributed to Zhao, J..

At least 343 records · Page 19Linked to original sources

bioRxiv2023

Poly(A) tail length regulation by mRNA deadenylases is critical for suppression of transposable elements

Transposons are mobile genetic elements that can impair the host genome stability and integrity. In plants, suppression of transposons is thought to be mediated mainly by small RNAs; however, the role of RNA decay in posttranscriptional repression of transposons is unknown. Here we show that RNA deadenylation is critical for controlling transposons in Arabidopsis. Previously, we demonstrated that transposon RNAs often harbor structural aberrancy owing to its inherently suboptimal codon usage and ribosome stalling. Such RNA aberrancy is monitored and resolved by RNA decay which is initiated by removal of poly(A) tail or deadenylation. The CCR4-NOT complex is a primary RNA deadenylase in Arabidopsis, and we found that it is required for stable repression of transposons. Intriguingly, RNA deadenylation controls transposons that are not targeted by cytoplasmic secondary small RNAs, which implies a target-specific regulation of transposon by the host. Our study suggests a previously unknown mechanism for transposon repression mediated by RNA deadenylation and unveils a complex nature of the hosts strategy to maintain the genome integrity.

plant biology↗

bioRxiv2023

Artificial selection footprints in domestic chicken genomes

Accurate and low-cost next generation sequencing technologies make re-sequencing of large populations of a species possible. Although many studies related to artificial selection signatures of commercial and indigenous chickens have been carried out, quite a small number of genes have been found to be under selection. In this study, we re-sequenced 85 individuals of five indigenous chicken breeds with distinct traits from Yunnan, a southwest province of China. By analyzing these indigenous chickens together with 116 individuals of commercial chickens (broilers and layers) and 35 individuals of red jungle fowl, we find a substantially large number of selective sweeps and affected genes for each chicken breed using a rigorous statistic model than previously reported. We confirm most of previously identified selective sweeps and affected genes. Meanwhile the vast majority ([~]98.3%) of our identified selective sweeps overlap known chicken quantitative trait loci. Thus, our predictions are highly reliable. For each breed, we also identify candidate genes and selective sweeps that might be related to the unique traits of the chickens.

bioinformatics↗

bioRxiv2023

Epithelial galectin-3 induces mitochondrial complex inhibition and cell cycle arrest of CD8+ T Cells in severe/critical ill COVID-19

Several studies have identified the presence of functionally depleted CD8+ T cells in COVID-19 patients, and particularly abnormally reduced CD8+ T cells in severe/critical patients, which may be a major cause of disease progression and poor prognosis. In this study, a proliferating-depleted CD8+ T cell phenotype was observed in severe/critical COVID-19 patients through scRNA-seq and scTCR-seq analysis. These CD8+ T cells were subsequently found to be characterized by cell cycle arrest and downregulation of mitochondrial biogenesis and respiratory chain complex genes. Cellchat analysis revealed that the Galectin signaling pathways between infected lung epithelial cells and CD8+ T cells play the key role in inducing CD8+ T cell reduction and dysfunction in severe/critical COVID-19. We used SARS-COV-2 ORF3a to transfect A549 epithelial cells, and co-cultured with CD8+ T cells. The ex vivo experiments confirmed that galectin-3 inhibited the transcription of mitochondrial respiratory chain complex III/IV genes in CD8+ T cells by suppressing the nuclear translocation of nuclear respiratory factor 1 (NRF1). In addition, the regulatory effect of galectin-3 was correlated with the activation of ERK signaling and/or the inhibition of Akt signaling. Galectin-3 inhibitor, TD-139, promoted nuclear translocation of NRF1, and enhanced mitochondrial respiratory chain complex III/IV gene expression and mitochondrial biogenesis, then restore the expansion ability of CD8+ T cells. Our study improved the understanding the immunopathogenesis and provided new target for the prevention and treatment of severe/critical COVID-19.

immunology↗

bioRxiv2023

Endothelial cell-derived lactate triggers mesenchymal stem cell histone lactylation to attenuate osteoporosis

Blood vessels play a role in osteogenesis and osteoporosis; however, the role of vascular metabolism is unclear. The present study found that ovariectomized mice exhibit reductions in bone blood vessel density and expression of endothelial glycolytic regulator pyruvate kinase M2 (PKM2). Additional data showed that endothelial cell (EC)-specific deletion of Pkm2 impair osteogenesis and worsen osteoporosis in mice. This was attributed to the impaired differentiation ability toward osteoblast of bone mesenchymal stem cells (BMSCs). Mechanistically, EC-specific deletion of Pkm2 reduce serum lactate levels secreted by ECs, which affect histone lactylation of BMSCs. We identified collagen type I alpha 2 chain, cartilage oligomeric matrix protein, ectonucleotide pyrophosphatase/phosphodiesterase 1, and transcription factor 7 like 2 as histone H3K18 lactylation-regulated osteogenic genes using joint CUT&Tag and RNA-sequencing analyses. The overexpression of PKM2 in ECs, addition of lactate, and exercise were observed to restore the phenotype of endothelial Pkm2-deficient mice. Furthermore, metabolomics of the serum indicated that osteoporosis patients showed a relatively low lactate level. The histone lactylation and related osteogenic genes of BMSCs in osteoporosis patients also decreased. In conclusion, the glycolysis of ECs fuels the differentiation of BMSCs into osteoblasts through histone lactylation, and exercise partially ameliorates osteoporosis through increased serum lactate.

cell biology↗

bioRxiv2023

Broadly effective ACE2 decoy proteins protect mice from lethal SARS-CoV-2 infection

As SARS-CoV-2 variants have been causing increasingly serious drug resistance problem, development of broadly effective and hard-to-escape anti-SARS-CoV-2 agents is in urgent need. Here we describe further development and characterization of two SARS-CoV-2 receptor decoy proteins, ACE2-Ig-95 and ACE2-Ig-105/106. We found that both proteins had potent and robust in vitro neutralization activities against diverse SARS-CoV-2 variants including Omicron, with an average IC50 of up to 37 pM. In a stringent lethal SARS-CoV-2 infection mouse model, both proteins lowered lung viral load by up to [~]1000 fold, prevented the emergence of clinical signs in >75% animals, and increased animal survival rate from 0% (untreated) to >87.5% (treated). These results demonstrate that both proteins are good drug candidates for protecting animals from severe COVID-19. In a head-to-head comparison of these two proteins with five previously-described ACE2-Ig constructs, we found that two of these constructs, each carrying five surface mutations in the ACE2 region, had partial loss of neutralization potency against three SARS-CoV-2 variants. These data suggest that extensively mutating ACE2 residues near the RBD-binding interface should be avoided or performed with extra caution. Further, we found that both ACE2-Ig-95 and ACE2-Ig-105/106 could be produced to gram/liter level, demonstrating the developability of them as biologic drug candidates. Stress-condition stability test of them further suggests that more studies are required in the future to improve the stability of these proteins. These studies provide useful insight into critical factors for engineering and preclinical development of ACE2 decoys as broadly effective therapeutics against diverse ACE2-utilizing coronaviruses. Abstract ImportanceEngineering soluble ACE2 proteins that function as a receptor decoy to block SARS-CoV-2 infection is a very attractive approach to broadly effective and hard-to-escape anti-SARS-CoV-2 agents. This study here describes development of two antibody-like soluble ACE2 proteins that broadly block diverse SARS-CoV-2 variants including Omicron. In a stringent COVID-19 mouse model, both proteins successfully protected >87.5% animals from lethal SARS-CoV-2 infection. In addition, a head-to-head comparison of the two constructs developed in this study with five previously-described ACE2 decoy constructs were performed here. Two previously-described constructs with relatively more ACE2-surface mutations were found with less robust neutralization activities against diverse SARS-CoV-2 variants. Further, the developability of the two proteins as biologic drug candidates was also assessed here. This study provides two broadly anti-SARS-CoV-2 drug candidates and useful insight into critical factors for engineering and preclinical development of ACE2 decoy as broadly effective therapeutics against diverse ACE2-utilizing coronaviruses. TweetTwo antibody-like ACE2 decoy proteins could block diverse SARS-CoV-2 variants and prevent animals from severe COVID-19.

microbiology↗

bioRxiv2023

Multi-omics and Pharmacological Characterization of Patient-derived Glioma Cell Lines

Glioblastoma (GBM) is the most common brain tumor and is currently incurable. Primary GBM cultures are widely used tools for screening potentially therapeutic drugs; however, there is a lack of genomic and pharmacological characterization of these primary GBM cultures. Here, we collected 52 patient-derived glioma cell (PDGC) lines and characterized them through whole- genome sequencing (WGS), RNA-seq, and drug response screening. We identified three molecular subtypes among PDGCs: mesenchymal (MES), proneural (PN), and oxidative phosphorylation (OXPHOS). Upon profiling the responses of PDGCs to 214 drugs, we found that the PN subtype PDGCs were sensitive to tyrosine kinase inhibitors, whereas the OXPHOS subtype PDGCs were sensitive to histone deacetylase inhibitors, oxidative phosphorylation inhibitors, and HMG-CoA reductase inhibitors. PN and OXPHOS subtype PDGCs stably formed tumors in vivo upon intracranial transplantation into immunodeficient mice, while most MES subtype PDGCs were incapable of tumorigenesis in vivo. In addition, profiling and follow-up investigations showed that the serum-free culture system used for PDGCs enriched and propagated rare MYC/MYCN- amplified glioma cells. Our study provides a resource for understanding primary glioma cell cultures and aiding clinical translation. SignificanceOur study provides a resource for patient-derived glioma cell lines (PDGCs) on transcriptome, genome, drug response, and tumorigenic abilities. PDGCs are categorized into PN, MES, and OXPHOS subtypes, with MES-subtype PDGCs incapable of tumorigenesis in vivo. Notably, the serum-free culture system for PDGCs enriches glioma cells with MYC/MYCN amplification.

cancer biology↗

bioRxiv2023

Small-molecule targeting of GPCR-independent non-canonical G protein signaling inhibits cancer progression

Activation of heterotrimeric G-proteins (G{beta}{gamma}) by G-protein-coupled receptors (GPCRs) is a quintessential mechanism of cell signaling widely targeted by clinically-approved drugs. However, it has become evident that heterotrimeric G-proteins can also be activated via GPCR-independent mechanisms that remain untapped as pharmacological targets. GIV/Girdin has emerged as a prototypical non-GPCR activator of G proteins that promotes cancer metastasis. Here, we introduce IGGi-11, a first-in-class smallmolecule inhibitor of non-canonical activation of heterotrimeric G-protein signaling. IGGi-11 binding to G-protein -subunits (Gi) specifically disrupted their engagement with GIV/Girdin, thereby blocking non-canonical G-protein signaling in tumor cells, and inhibiting pro-invasive traits of metastatic cancer cells in vitro and in mice. In contrast, IGGi-11 did not interfere with canonical G-protein signaling mechanisms triggered by GPCRs. By revealing that small molecules can selectively disable non-canonical mechanisms of G-protein activation dysregulated in disease, these findings warrant the exploration of therapeutic modalities in G-protein signaling that go beyond targeting GPCRs.

biochemistry↗

bioRxiv2023

Fiberbots: Robotic fibers for high-precision minimally invasive surgery

Technologies that rely on the fundamental principle of thermal expansion have demonstrated high-precision, a growing demand in fields driven by miniaturization. However, scalable production of high aspect ratio devices that harness this capability while facilitating flexibility in design and functionality remains a challenge. We employed the high-throughput fiber thermal drawing technique to readily fabricate multimaterial fiberbots that can precisely and omnidirectionally move by asymmetric thermal expansion. These millimeter-scale fibers (< 2 mm) show excellent repeatability and linearity, negligible hysteresis, and can achieve micron-level resolution over four orders of magnitude motion range. By integrating these robotic fibers with medical devices that can perform cellular-level tissue imaging, diagnosis, and manipulation, we showcase their versatility through benchtop and preclinical animal studies and their overall potential impact on medicine, biomedical engineering, robotics, and beyond. One Sentence SummaryScalable manufacturing and integration of robotic fibers that deliver high-precision motion when heated.

bioengineering↗

bioRxiv2023

Ferroptosis is determined by chloride ions

Ferroptosis has emerged as a deliberate type of programmed cell death that manifests marked importance ubiquitously in health and diseases. However, after a decade of research, the mechanisms of ferroptosis execution remain unclear. Here we identify chloride ions (Cl-) as essential determinants of ferroptosis. Water homeostasis manipulated by extracellular solute concentration disrupts ferroptotic cell death. Hyperosmotic stress attenuates ferroptosis and endues cells with high lipid peroxidation. Analyses of a fluorescent chloride probe show that Cl- fluxes into the cytoplasm during ferroptosis, substantiating a role for Cl- to drive water flow. Depletion of extracellular chloride ions ([Cl-]o) from culture media congruously confers resistance to ferroptosis. The [Cl-]o-depleted ferroptotic cells fall into two populations: cells with low lipid peroxidation; cells with high lipid peroxidation but not cell swelling or cell rupture. Contrarily, solitary [Cl-]o overload is sufficient to elicit ferroptosis without canonical ferroptosis inducers. Further experiments show that ferroptotic cells depolarize and [Cl-]o is positively correlated with this process. Membrane depolarization upregulates the level of lipid peroxidation, suggesting that membrane potential may be a universal mechanism governing ferroptosis. Together, our findings reveal that ferroptosis is determined by chloride ions.

cell biology↗

bioRxiv2023

An Unexpected Role of Neutrophils in Clearing Apoptotic Hepatocytes In Vivo

Billions of apoptotic cells are removed daily in a human adult by professional phagocytes (e.g. macrophages) and neighboring nonprofessional phagocytes (e.g. stromal cells). Despite being a type of professional phagocyte, neutrophils are thought to be excluded from apoptotic sites to avoid tissue inflammation. Here we report a fundamental and unexpected role of neutrophils as the predominant phagocyte responsible for the clearance of apoptotic hepatic cells in the steady state. In contrast to the engulfment of dead cells by macrophages, neutrophils burrowed directly into apoptotic hepatocytes, a process we term perforocytosis, and ingested the effete cells from the inside. The depletion of neutrophils caused defective removal of apoptotic bodies, induced tissue injury in the mouse liver and led to the generation of autoantibodies. Human autoimmune liver disease showed similar defects in the neutrophil-mediated clearance of apoptotic hepatic cells. Hence, neutrophils possess a specialized immunologically silent mechanism for the clearance of apoptotic hepatocytes through perforocytosis, and defects in this key housekeeping function of neutrophils contribute to the genesis of autoimmune liver disease.

physiology↗

bioRxiv2023

Single-cell transcriptomic analyses reveal the cellular and genetic basis of aquatic locomotion in scyphozoan jellyfish

The authors have withdrawn their manuscript because the authors need to re-organize the data and writing. More experimental evidence from whole-mount ISH assay and RNAi assays will be added to support the viewpoints. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.

ecology↗

bioRxiv2023

IMPACT: a web server for exploring immunotherapeutic predictive and cancer prognostic biomarkers

Immune checkpoint inhibitors (ICIs) are a breakthrough in oncology treatment, and studies of screening predictive biomarkers of ICIs are emerging. We developed a web server named IMPACT (http://impact.brbiotech.com/) to thoroughly explore immunotherapeutic predictive or prognostic biomarkers. IMPACT contains a large dataset of 6,276 patients treated with ICIs and integrates 11 well-designed function modules, enabling an in-depth solution for biomarkers exploration. Compared with the existing tools, IMPACT was implemented with one exclusive module for interaction analysis and several optimized conventional functions for discovering novel biomarkers. Specifically, the interaction analysis of biomarker-treatment effect is essential to determine whether a biomarker is predictive and/or prognostic for ICIs. Moreover, several optimized functions allow complicated biomarker exploration, including customized selections of variant types in more detail, automatically screening meaningful co-mutations among multiple genes, and selecting cut-off values for gene expression biomarkers. In summary, IMPACT is a comprehensive analysis resource to facilitate biomarker research of ICIs.

bioinformatics↗

bioRxiv2023

Depletion-assisted multiplexing cell-free RNA sequencing reveals distinct human and microbial signatures in plasma versus extracellular vesicle

Cell-free long RNAs in human plasma and extracellular vesicles (EVs) have shown promise as biomarkers in liquid biopsy, despite their fragmented nature. To investigate these fragmented cell-free RNAs (cfRNAs), we developed a cost-effective cfRNA sequencing method called DETECTOR-seq (depletion-assisted multiplexed cell-free total RNA sequencing). DETECTOR-seq utilized a meticulously tailored set of customized guide RNAs to remove large amounts of unwanted RNAs (i.e., fragmented ribosomal and mitochondrial RNAs) in human plasma. Early barcoding strategy was implemented to reduce costs and minimize plasma requirements. Using DETECTOR-seq, we conducted a comprehensive analysis of cell-free transcriptomes in both whole human plasma and EVs. Our analysis revealed discernible distributions of RNA types in plasma and EVs. Plasma exhibited pronounced enrichment in structured circular RNAs, tRNAs, Y RNAs, and viral RNAs, while EVs showed enrichment in mRNAs and srpRNAs. Functional pathway analysis highlighted RNA splicing-related ribonucleoproteins (RNPs) and antimicrobial humoral response genes in plasma, while EVs demonstrated enrichment in transcriptional activity, cell migration, and antigen receptor-mediated immune signals. Our study indicates the comparable potential of cfRNAs from whole plasma and EVs in distinguishing cancer patients (i.e., colorectal and lung cancer) from healthy donors. And microbial cfRNAs in plasma showed potential in classifying specific cancer types. Our comprehensive analysis of total and EV cfRNAs in paired plasma samples provides valuable insights for determining the need for EV purification in cfRNA-based studies. We envision the cost-effectiveness and efficiency of DETECTOR-seq will empower transcriptome-wide investigations in the fields of extracellular vesicles and liquid biopsy.

molecular biology↗

bioRxiv2023

Construction of a 3D whole organism spatial atlas by joint modeling of multiple slices

Spatial transcriptomics (ST) technologies are revolutionizing the way that researchers explore the spatial architecture of tissues. Currently, ST data analysis is often restricted to 2D space within a single tissue slice, limiting our capacity to understand biological processes that take place in 3D space. Here, we present STitch3D, a unified computational framework that integrates multiple 2D tissue slices to reconstruct 3D cellular structures from the tissue level to the whole organism level. By jointly modeling multiple 2D tissue slices and integrating them with cell-type-specific expression profiles derived from single-cell RNA-sequencing data, STitch3D simultaneously identifies 3D spatial regions with coherent gene expression levels and reveals 3D distributions of cell types. STitch3D distinguishes biological variation among slices from batch effects, and effectively borrows shared information across slices to assemble powerful 3D models of tissues. Through comprehensive experiments using diverse datasets, we demonstrate the performance of STitch3D in building comprehensive 3D tissue architectures of the mouse brain, the human heart, and the Drosophila embryo, which allow 3D analysis in the entire tissue region or even the whole organism. To gain deeper biological insights, the outputs of STitch3D can be used for downstream tasks, such as inference of spatial trajectories, identification of spatially variable genes enriched in tissue regions or subregions, denoising or imputation of spatial gene expressions, as well as generation of virtual tissue slices.

bioinformatics↗

bioRxiv2023

P2X7 signaling mediates migraine-related augmentation of meningeal afferent mechanosensitivity

Cortical spreading depolarization (CSD) is a key pathophysiological event that underlies visual and sensory auras in migraine. CSD is also thought to drive the headache phase in migraine by promoting the activation and mechanical sensitization of trigeminal primary afferent nociceptive neurons that innervate the cranial meninges. The factors underlying meningeal nociception in the wake of CSD remain poorly understood but potentially involve the parenchymal release of algesic mediators and damage-associated molecular patterns, particularly ATP. Here, we explored the role of ATP-P2X purinergic receptor signaling in mediating CSD-evoked meningeal afferent activation and mechanical sensitization. Male rats were subjected to a single CSD episode. In vivo, extracellular single-unit recording was used to measure meningeal afferent ongoing activity changes. Quantitative mechanical stimuli using a servomotor force-controlled stimulator assessed changes in the afferents mechanosensitivity. Manipulation of meningeal P2X receptors was achieved via local administration of pharmacological agents. Broad-spectrum P2X receptor inhibition, selective blockade of the P2X7 receptor, and its related pannexin 1 channel suppressed CSD-evoked afferent mechanical sensitization but did not affect the accompanying afferent activation response. Surprisingly, inhibition of the pronociceptive P2X2/3 receptor did not affect the activation or sensitization of meningeal afferents post-CSD. P2X7 signaling underlying afferent mechanosensitization was localized to the meninges and did not affect CSD susceptibility. We propose that meningeal P2X7 and Pannexin 1 signaling, potentially in meningeal macrophages or neutrophils, mediates the mechanical sensitization of meningeal afferents, which contributes to migraine pain by exacerbating the headache during normally innocuous physical activities. Significance StatementActivation and sensitization of meningeal afferents play a key role in migraine headache, but the underlying mechanisms remain unclear. Here, using a rat model of migraine with aura involving cortical spreading depolarization (CSD), we demonstrate that meningeal purinergic P2X7 signaling and its related Pannexin 1 pore, but not nociceptive P2X2/3 receptors, mediate prolonged meningeal afferent sensitization. Additionally, we show that meningeal P2X signaling does not contribute to the increased afferent ongoing activity in the wake of CSD. Our finding points to meningeal P2X7 signaling as a critical mechanism underlying meningeal nociception in migraine, the presence of distinct mechanisms underlying the activation and sensitization of meningeal afferents in migraine, and highlight the need to target both processes for effective migraine therapy.

neuroscience↗

bioRxiv2023

Real-Time Quantification of Cell Mechanics and Functions by Double Resonator Piezoelectric Cytometry -Theory and Study of Cellular Adhesion of HUVECs

Cell mechanics is closely associated with cellular structure and function. However, the inability to measure both cellular force and viscoelasticity of statistically significant number of cells noninvasively remains a challenge for quantitative characterizations of various cellular functions and practical applications. Here a double resonator piezoelectric cytometry (DRPC), using AT and BT cut quartz crystals of the same frequency and surface morphology is developed to simultaneously quantify the cells-generated forces ({Delta}S) and viscoelastic moduli (G', G'') of a population of isolated single cells or cells with different degrees of cell-cell interactions in a non-invasive and real time manner. DRPC captures the dynamic mechanical parameters {Delta}S and G', G'' during the adhesions of human umbilical vein endothelial cells (HUVECs) under different ligand densities of adhesion molecules fibronectin or Arg-Gly-Asp (RGD) modified on the gold surfaces of 9 MHz AT and BT cut quartz crystals, and different seeding densities of HUVECs. It is found that both the ligand density and cell seeding density affect the magnitudes of {Delta}S and G', G'' and their correlations are revealed for the first time by DRPC. The validity of DRPC is further verified by mechanical changes of the cells in response to treatments with cytoskeleton regulators.

biophysics↗

bioRxiv2023

PI(3,5)P2 Controls the Signaling Activity of Class I PI3K

3-Phosphoinositides are ubiquitous cellular lipids that play pivotal regulatory roles in health and disease. Among 3-phosphoinositides, phosphatidylinositol-3,5-bisphosphate (PI(3,5)P2) remains the least understood species in terms of its spatiotemporal dynamics and physiological function due to the lack of a specific sensor that allows spatiotemporally resolved quantitative imaging of PI(3,5)P2. Using a newly developed ratiometric PI(3,5)P2 sensor engineered from the C-terminal SH2 domain of Class I phosphoinositide 3-kinases (PI3K)-p85 subunit we demonstrate that a unique pool of PI(3,5)P2 is generated on lysosomes and late endosomes in response to growth factor stimulation. This PI(3,5)P2, the formation of which is mediated sequentially by Class II PI3KC2{beta} and PIKfyve, plays a crucial role in terminating the activity of growth factor-stimulated Class I PI3K, one of the most frequently mutated proteins in cancer, via specific interaction with its regulatory p85 subunit. A small molecule inhibitor of p85-PI(3,5)P2 binding specifically blocks the feedback inhibition of Class I PI3K by PI(3,5)P2 and thus serves as a PI3K activator that promotes neurite growth. Furthermore, cancer-causing mutations of the Class I PI3K-p85 subunit inhibit p85-PI(3,5)P2 interaction and thereby induce sustained activation of Class I PI3K. Our results unravel a hitherto unknown spatiotemporally specific regulatory function of PI(3,5)P2 that links Class I and II PI3Ks and modulates the magnitude of PI3K-mediated growth factor signaling. These results also suggest new therapeutic possibilities for treating cancer patients with p85 mutations and promoting wound healing and tissue regeneration.

biochemistry↗

bioRxiv2023

Visualization and review of reads alignment on the graphical pan-genome with VAG

Recent advances in pangenomics have led to the rapid development of graph-based pangenomes that code genetic variant as nodes and edges preserve the contiguity of the sequence and structural variation between individuals. Data visualization is an essential component of genomic data analysis. However, the further application of the graph-based pangenome is still suffered from lacking bioinformatics tools to visualize graph format pangenomes and understand the reads alignment on graph pangenomes. In this research, we developed a novel bioinformatics platform, VAG (Visualizing read alignments in graph genomes), to overcome these challenges. VAG includes multifunctional modules integrated into a single command line and an online visualization platform supported through a web server. This tool can extract specific sequence regions from a graph pangenome and display read alignments on different paths of a graph pangenome. In addition, VAG provides population-level presence/absence variations frequency analysis and sequence path navigation to identify the population differentiation regions. To demonstrate the usage, we investigated genetic variations using a rice graph pangenome with population-level sequencing data to identify important genes and gene clusters underlying the indica-japonica differentiation with VAG. After investigating read alignments on the graph pangenome, we identified many false-positive alignments due to TE insertions. To reduce the impact of these misleading alignments, we developed a navigation module to determine and filter those false-positive alignments based on the pair-end alignment information. The utilization of mate-pair information in VAG provides a reliable reference for variation identification. Additionally, we developed a VAG web server to provide a user-friendly and interactive platform to visualize the read alignment data. VAG was also applied to SV discovery in the cucumber and soybean graph-based pangenome and details of VAG can be accessed by the following website (https://ricegenomichjx.xiaomy.net/VAG/sequenceextraction.php).

bioinformatics↗