Biology subjects

Qiu, X.

Publications and source records attributed to Qiu, X..

At least 91 records · Page 5Linked to original sources

bioRxiv2023

Coronavirus endoribonuclease nsp15 induces host cellular protein synthesis shutoff

The endoribonuclease (EndoU) nsp15 of coronaviruses plays an important role in evasion of host innate immune responses by reducing the abundance of viral double-stranded RNA, whereas less is known about potential host cellular targets of nsp15. In this study, we show that cellular protein synthesis is inhibited upon over-expression of nsp15 from four genera of coronaviruses and this is accompanied by nuclear retention of the poly(A) binding protein cytoplasmic 1 (PABPC1). We also show that the EndoU activity of nsp15 is indispensable for both, inhibition of protein synthesis and PABPC1 nuclear relocation. FISH analysis using oligo-dT probes, revealed an overlap between the localization of cellular mRNA and that of overexpressed nsp15 in some cells, suggesting that, when expressed alone, nsp15 may target host mRNA. When investigating the association of nsp15 on protein shut off in the context of a viral infection, we observed that the{gamma} -coronavirus infectious bronchitis virus (IBV), induced host translation shutoff in an p-eIF2-independent manner and mainly retained PABPC1 in the cytoplasm, whereas the nsp15 EndoU-deficient IBV accumulated viral dsRNA and caused p-PKR-p-eIF2-dependent host protein translation shutoff, accompanied with PABPC1 nuclear relocation or stress granule (SG) localization. This phenomenon suggests that during infection with wild type IBV, although the cellular translation is inhibited, initiation of viral mRNA translation leads to PABPC1 binding to viral mRNA, thereby preventing its nuclear entry; during infection with nsp15 EndoU-deficient IBV however, the eIF2-dependent host protein translation shutoff prevents both host and viral mRNA translation initiation, releasing PABPC1 from binding to cytosolic and viral mRNA, thereby relocating it to the nucleus or to SG. Altogether, this study reveals unique yet conserved mechanisms of host protein shutoff that add to our understanding of how coronaviruses regulate host protein expression through a mechanism that involves catalytically active nsp15 EndoU, and describes how nsp15 may target both, viral and host mRNA. Author summaryIt has been reported that coronavirus infection suppresses host protein translation, - and {beta}- coronavirus nsp1 is responsible for inhibition of host gene expression. However, for{gamma} - and{delta} -coronavirus, there is no nsp1 and the underlying mechanisms by which virus regulates host translation are not well characterized. Here, we show that coronavirus endoribonuclease nsp15 is responsible for the inhibition of host translation by targeting to host factors, meanwhile it helps virus bypass the PKR-eIF2 mediated host translation shutoff, which is harmful for virus gene expression, by reducing the accumulation of viral dsRNA. This novel finding gives insight how does nsp15 targets to both host factors and viral RNA, to facilitate virus replication. Moreover, the novel function of nsp15 is found to be conserved among coronaviruses, revealing the essential role of this endoribonuclease in hijacking host translation machinery for virus replication.

microbiology↗

bioRxiv2023

Stem cell factor and granulocyte colony-stimulating factor promote remyelination in the chronic phase of severe traumatic brain injury

Severe traumatic brain injury (TBI) causes long-term disability and death in young adults. White matter is vulnerable to TBI damage. Demyelination is a major pathological change of white matter injury after TBI. Demyelination which is characterized by myelin sheath disruption and oligodendrocyte cell death leads to long-term neurological function deficits. Stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) treatments have shown neuroprotective and neurorestorative effects in the subacute and chronic phases of experimental TBI. Our previous study has revealed that combined SCF and G-CSF treatment (SCF+G-CSF) enhances myelin repair in the chronic phase of TBI. However, the long-term effect and mechanism of SCF+G-CSF-enhanced myelin repair remain unclear. In this study, we uncovered persistent and progressive myelin loss in the chronic phase of severe TBI. SCF+G-CSF treatment in the chronic phase of severe TBI enhanced remyelination in the ipsilateral external capsule and striatum. The SCF+G-CSF-enhanced myelin repair is positively correlated with the proliferation of oligodendrocyte progenitor cells in the subventricular zone. These findings reveal the therapeutic potential of SCF+G-CSF in myelin repair in the chronic phase of severe TBI and shed light on the mechanism underlying SCF+G-CSF-enhanced remyelination in chronic TBI.

neuroscience↗

bioRxiv2023

TGFB1 Induces Fetal Reprogramming and Enhances Intestinal Regeneration

The adult gut epithelium has a remarkable ability to recover from damage. To achieve cellular therapies aimed at restoring and/or replacing defective gastrointestinal tissue, it is important to understand the natural mechanisms of tissue regeneration. We employed a combination of high throughput sequencing approaches, mouse genetic models, and murine and human organoid models, and identified a role for TGFB signaling during intestinal regeneration following injury. At 2 days following irradiation (IR)-induced damage of intestinal crypts, a surge in TGFB1 expression is mediated by monocyte/macrophage cells at the location of damage. Depletion of macrophages or genetic disruption of TGFB-signaling significantly impaired the regenerative response following irradiation. Murine intestinal regeneration is also characterized by a process where a fetal transcriptional signature is induced during repair. In organoid culture, TGFB1-treatment was necessary and sufficient to induce a transcriptomic shift to the fetal-like/regenerative state. The regenerative response was enhanced by the function of mesenchymal cells, which are also primed for regeneration by TGFB1. Mechanistically, integration of ATAC-seq, scRNA-seq, and ChIP-seq suggest that a regenerative YAP-SOX9 transcriptional circuit is activated in epithelium exposed to TGFB1. Finally, pre-treatment with TGFB1 enhanced the ability of primary epithelial cultures to engraft into damaged murine colon, suggesting promise for the application of the TGFB-induced regenerative circuit in cellular therapy. GRAPHIC ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/523825v1_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@147f3b7org.highwire.dtl.DTLVardef@d1a64dorg.highwire.dtl.DTLVardef@1fd0dc0org.highwire.dtl.DTLVardef@91f7d8_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2023

Sexual dimorphism in obesity is governed by RELMα regulation of adipose macrophages and eosinophils

Obesity incidence is increasing worldwide with the urgent need to identify new therapeutics. Sex differences in immune cell activation drive obesity-mediated pathologies where males are more susceptible to obesity co-morbidities and exacerbated inflammation. Here, we demonstrate that the macrophage-secreted protein RELM critically protects females against high fat diet-induced obesity. Compared to male mice, RELM levels were elevated in both control and high fat dietfed females and correlated with adipose macrophages and eosinophils. RELM-deficient females gained more weight and had pro-inflammatory macrophage accumulation and eosinophil loss, while both RELM treatment and eosinophil transfer rescued this phenotype. Single cell RNA-sequencing of the adipose stromal vascular fraction was performed and identified sex and RELM-dependent changes. Genes involved in oxygen sensing and iron homeostasis, including hemoglobin and lncRNA Gm47283, correlated with increased obesity, while eosinophil chemotaxis and response to amyloid-beta were protective. Monocyte-to-macrophage transition was also dysregulated in RELM-deficient animals. Collectively, these studies implicate a RELM-macrophage-eosinophil axis in sex-specific protection against obesity and uncover new therapeutic targets for obesity.

immunology↗

bioRxiv2022

Spateo: multidimensional spatiotemporal modeling of single-cell spatial transcriptomics

Cells do not live in a vacuum, but in a milieu defined by cell-cell communication that can be measured via emerging high-resolution spatial transcriptomics approaches. However, analytical tools that fully leverage such data for kinetic modeling remain lacking. Here we present Spateo (aristoteleo/spateo-release), a general framework for quantitative spatiotemporal modeling of single-cell resolution spatial transcriptomics. Spateo delivers novel methods for digitizing spatial layers/columns to identify spatially-polar genes, and develops a comprehensive framework of cell-cell interaction to reveal spatial effects of niche factors and cell type-specific ligand-receptor interactions. Furthermore, Spateo reconstructs 3D models of whole embryos, and performs 3D morphometric analyses. Lastly, Spateo introduces the concept of "morphometric vector field" of cell migrations, and integrates spatial differential geometry to unveil regulatory programs underlying various organogenesis patterns of Drosophila. Thus, Spateo enables the study of the ecology of organs at a molecular level in 3D space, beyond isolated single cells.

systems biology↗

bioRxiv2022

Targeting TR4-induced RCC cells-derived exosomally initiated signaling increases Sunitinib efficacy

Sunitinib is the first-line therapy for metastatic clear cell renal cell carcinoma (RCC) via suppressing neoangiogenesis and tumor growth. The detailed mechanisms, especially whether and how RCC cells can impact endothelial cells sensitivity to Sunitinib, remain unclear. Here, we found that TR4 was commonly upregulated in RCC tissue and relative to tumor angiogenesis. Tube formation and Mouse aortic ring assay showed that the overexpression or knockdown of TR4 in RCC cells enhanced or reduced the angiogenesis of endothelial cells and their resistance to Sunitinib in vitro. Mechanistically, We found that TR4 transcriptionally increase ADAM15 expression, as a consequence, exosomes carrying relatively large amounts of ADAM15 secreted from RCC cell resulted in activating the EGFR phosphorylation and reducing the efficacy of Sunitinib in endothelial cells. Targeting the TR4-induced renal cancers-derived exosomelly initiated signaling with a small molecular, CRM197, increases sunitinib efficacy in vitro and xenograft tumor models. Taken together, our findings indicate a novel function of TR4 in RCC blunted the efficacy of sunitinib via exosomal ADAM15-induced activation of EGFR signaling pathway in endothelial cells.

cancer biology↗

bioRxiv2022

Structural basis for BIRC6 to balance apoptosis and autophagy

Caspase-9 is the initiator caspase for the intrinsic apoptotic cell death pathway, and is critical to the activation of effector caspases during apoptosis, but how its levels and activities are maintained remains unclear. The gigantic Inhibitor of Apoptosis Protein (IAP) BIRC6/BRUCE/Apollon not only inhibits apoptosis, but also promotes ubiquitination of the key autophagic protein LC3 and inhibits autophagy. Here we show that BIRC6 forms an anti-parallel U-shaped dimer in a 3.6-[A] cryo-EM structure with multiple previously unannotated domains, including a ubiquitin-like domain, and discover that the mitochondria-derived pro-apoptotic factor Smac/DIABLO binds BIRC6 by interacting with one BIR domain, two carbohydrate-binding modules and two helices in the central cavity. Notably, Smac outcompetes the effector caspase 3 and the pro-apoptotic protease HtrA2, but not caspase 9, for binding BIRC6. BIRC6 strongly inhibits cellular activity of caspase 9, but weakly suppresses that of caspase 3. Meanwhile, BIRC6 binds LC3 through an LC3-interacting region, probably following dimer disruption of this BIRC6 region. Deficiency in LC3 ubiquitination promotes autophagy and autophagic degradation of BIRC6, and inhibits apoptosis. Moreover, induction of autophagy promotes autophagic degradation of both procaspase-9 and active caspase-9, but not of effector caspases. These results are important to understand how the balance between apoptosis and autophagy is regulated under pathophysiological conditions.

cell biology↗

bioRxiv2022

Targeting the BAG-1 family of co-chaperones in lethal prostate cancer.

Therapies that abrogate persistent androgen receptor (AR) signaling in castration resistant prostate cancer (CRPC) remain an unmet clinical need. The N-terminal domain (NTD) of the AR drives transcriptional activity in CRPC but is intrinsically disordered and remains a challenging therapeutic target. Therefore, inhibiting critical co-chaperones, such as BAG-1L, is an attractive alternative strategy. We performed druggability analyses demonstrating the BAG domain to be a challenging drug target. Thio-2, a tool compound, has been reported to bind the BAG domain of BAG-1L and inhibit BAG-1L-mediated AR transactivation. However, despite these data, the mechanism of action of Thio-2 is poorly understood and the BAG domain which is present in all BAG-1 isoforms has not been validated as a therapeutic target. Herein, we demonstrate growth inhibiting activity of Thio-2 in CRPC cell lines and patient derived models with decreased AR genomic binding and AR signaling independent of BAG-1 isoform function. Furthermore, genomic abrogation of BAG-1 isoforms did not recapitulate the described Thio-2 phenotype, and NMR studies suggest that Thio-2 may bind the AR NTD, uncovering a potential alternative mechanism of action, although in the context of low compound solubility. Furthermore, BAG-1 isoform knockout mice are viable and fertile, in contrast to previous studies, and when crossed with prostate cancer mouse models, BAG-1 deletion does not significantly impact prostate cancer development and growth. Overall, these data demonstrate that Thio-2 inhibits AR signaling and growth in CRPC independent of BAG-1 isoforms, and unlike previous studies of the activated AR, therapeutic targeting of the BAG domain requires further validation before being considered a therapeutic strategy for the treatment of CRPC.

molecular biology↗

bioRxiv2022

A bioinformatics approach to systematically analyze the molecular patterns of monkeypox virus-host cell interactions

Monkeypox has been spreading worldwide since May 2022, when the World Health Organization (WHO) declared the outbreak of a "Public health emergency of international concern". The spread of monkeypox has posed a serious threat to the health of people all over the world but few studies have been carried out on it, and the molecular mechanism of monkeypox after infection remains unclear. We therefore implemented a transcriptome analysis to identify signaling pathways and biomarkers in monkeypox-infected cells to help understand monkeypox-host cell interactions. In this study, the dataset GSE36854 and GSE11234 were obtained from GEO. Among them, 84 significantly different genes were identified in the dataset GSE36854, followed by KEGG, GO analysis protein-protein interaction (PPI) construction and Hub gene extraction. We also analyzed the expression regulation patterns of hub genes and screened the drugs targeting the hub genes. The results showed that monkeypox-infected cells significantly activated the cellular immune response, and induced inflammatory response. IER3, IFIT2, IL11, ZC3H12A, EREG, IER2, NFKBIE, FST, IFIT1 and AREG were the top 10 hub genes, in which anti-viral gene IFIT1 and IFIT2 were significantly suppressed. AP-26113 and itraconazole promoting the expression of IFIT1 and IFIT2 may be used as new candidates for the treatment of monkeypox viral infection. Our results provide a new entry point for understanding the mode of interaction between monkeypox virus and its host.

microbiology↗

bioRxiv2022

Spatiotemporally resolved transcriptomics reveals subcellular RNA kinetic landscape

Spatiotemporal regulation of the cellular transcriptome is crucial for proper protein expression and cellular function1. However, the intricate subcellular dynamics of RNA synthesis, decay, export, and translocation remain obscured due to the limitations of existing transcriptomics methods2-8. Here, we report a spatiotemporally resolved RNA mapping method (TEMPOmap) to uncover subcellular RNA profiles across time and space at the single-cell level in heterogeneous cell populations. TEMPOmap integrates pulse-chase metabolic labeling of the transcriptome with highly multiplexed three-dimensional (3D) in situ sequencing to simultaneously profile the age and location of individual RNA molecules. Using TEMPOmap, we constructed the subcellular RNA kinetic landscape of 991 genes in human HeLa cells from upstream transcription to downstream subcellular translocation. Clustering analysis of critical RNA kinetic parameters across single cells revealed kinetic gene clusters whose expression patterns were shaped by multi-step kinetic sculpting. Importantly, these kinetic gene clusters are functionally segregated, suggesting that subcellular RNA kinetics are differentially regulated to serve molecular and cellular functions in cell-cycle dependent manner. Together, these single-cell spatiotemporally resolved transcriptomics measurements provide us the gateway to uncover new gene regulation principles and understand how kinetic strategies enable precise RNA expression in time and space.

genomics↗

bioRxiv2022

Systematic functional interrogation of SARS-CoV-2 host factors using Perturb-seq

Genomic and proteomic screens have identified numerous host factors of SARS-CoV-2, but efficient delineation of their molecular roles during infection remains a challenge. Here we use Perturb-seq, combining genetic perturbations with a single-cell readout, to investigate how inactivation of host factors changes the course of SARS-CoV-2 infection and the host response in human lung epithelial cells. Our high-dimensional data resolve complex phenotypes such as shifts in the stages of infection and modulations of the interferon response. However, only a small percentage of host factors showed such phenotypes upon perturbation. We further identified the NF-{kappa}B inhibitor I{kappa}B (NFKBIA), as well as the translation factors EIF4E2 and EIF4H as strong host dependency factors acting early in infection. Overall, our study provides massively parallel functional characterization of host factors of SARS-CoV-2 and quantitatively defines their roles both in virus-infected and bystander cells.

systems biology↗

bioRxiv2022

Human iPSC-derived neuron of 16p11.2 deletion reveals haplotype-specific expression of MAPK3 and its contribution to variable NDD phenotypes

Recurrent proximal 16p11.2 deletion (16p11.2del) is risk factor of diverse neurodevelopmental disorders (NDDs) with variable penetrance. Although previous human induced pluripotent stem cell (hiPSC) models of 16p11.2del confirmed disrupted neuron development, it is not known which gene(s) at this interval are mainly responsible for the abnormal cellular phenotypes and how the NDD penetrance is regulated. After haplotype phasing of 16p11.2 region, we generated hiPSCs for two 16p11.2del families with distinct residual haplotypes and variable NDD phenotypes. We also differentiated the hiPSCs to cortical neural cells and demonstrated MAPK3 as a driver signal of 16p11.2 region contributing to the dysfunctions in multiple pathways related to neuron development, which leads to altered morphological or electrophysiological properties in neuron cells. Furthermore, residual haplotype-specific MAPK3 expression was identified in 16p11.2del neuron cells, associating MAPK3 down-expression with the minor allele of the residual haplotype. Ten SNPs of the residual haplotype are mapped as enhancer SNPs (enSNPs) of MAPK3, eight enSNPs were functionally validated by luciferase assays, implying enSNPs contribute to residual haplotype-specific MAPK3 expression via cis-regulation. Finally, the analyses of three different patient cohorts showed that the residual haplotype of 16p11.2del is associated with variable NDD phenotypes.

genetics↗

bioRxiv2022

Modification Mechanism of Phenol-formaldehyde Resin with Crude Bio-oil by Model Compound Method

To clarify the modification mechanism of bio-oil for phenol-formaldehyde resin with crude bio-oil (BPF), the bio-oil compounds were simplified by model compound method according to the component distribution. The phenol-formaldehyde resin with bio-oil model compounds (BMPF) were prepared and their basic performance, bonding strength and aging characteristics were determined. The changes on the microstructure and chemical bonds of BMPF were also analyzed by scanning electron microscope, Fourier transform infrared spectroscopy, and nuclear magnetic resonance analysis. Results showed that the components of bio-oil had different influence on the performance and microstructure of BMPF, especially phenols. Structural analysis indicated that the phenols and ketones of bio-oil had positive effects on the synthesis of BMPF, while the aldehydes and acids had negative effects. But all components of bio-oil could improve the aging resistance of BMPF inordinately. These results could provide a basis for the modification of BPF.

ecology↗

bioRxiv2022

Decisive Roles of Sequence Distributions in the Generalizability of de novo Deep Learning Models for RNA Secondary Structure Prediction

Taking sequences as the only inputs, the class of de novo deep learning (DL) models for RNA secondary structure prediction has achieved far superior performances than traditional algorithms. However, key questions remain over the statistical underpinning of such models that make no use of physical laws or co-evolutionary information. We present a quantitative study of the capacity and generalizability of a series of de novo DL models, with a minimal two-module architecture and no post-processing, under varied distributions of the seen and unseen sequences. Our DL models outperform existing methods on commonly used benchmark datasets and demonstrate excellent learning capacities under all sequence distributions. These DL models generalize well over non-identical unseen sequences, but the generalizability degrades rapidly as the sequence distributions of the seen and unseen datasets become dissimilar. Examinations of RNA family-specific behaviors manifest not only disparate familydependent performances but substantial generalization gaps within the same family. We further determine how model generalization decreases with the decrease of sequence similarity via pairwise sequence alignment, providing quantitative insights into the limitations of statistical learning. Model generalizability thus poses a major hurdle for practical uses of de novo DL models and several tenable avenues for future advances are discussed.

bioinformatics↗

bioRxiv2022

Expanding adult tubular microvessels on stiff substrates with endothelial cells and pericytes from the same tissue

Microvessels are essential for tissue engineering and regeneration. In current methods, endothelial cells are usually cultured in commercially available media and form a monolayer of cell sheets on stiff substrates and a tubular structure when cultured with soft hydrogels. To mimic the microvessels in vivo, researchers usually coculture the endothelial cells and pericytes from different adult tissues or derived from pluripotent stem cells in a three-dimensional hydrogel. However, there is a challenge for these models to reflect tissue-specific characteristics due to the vascular heterogeneity throughout the body. Here, we established a culture model for expanding adult tubular microvessels on stiff substrates with endothelial cells and pericytes derived from the same tissue. We isolated microvessels from adult rat subcutaneous soft connective tissue and cultured them on regular plastic dishes. We performed a series of screenings and formulated a custom-made medium (Medium-X), containing mainly antioxidants and three small molecules, Chir99021, A83-01, and Y27632. Medium-X significantly promoted adult microvessel growth while maintaining their characteristic tubular morphology up to 8 weeks in vitro, contrary to the monolayer of endothelial cell sheets in the commercially available medium EGM2MV. Transcriptomic analysis showed that Medium-X maintained the tubular morphology of microvessels by promoting angiogenesis and vascular remodeling while suppressing oxidation and lipid metabolic pathways. The model presented in this study can be applied to other organs for expanding organ-specific microvessels for tissue engineering and vascular regeneration.

bioengineering↗

bioRxiv2022

Genetic Variation and Disease Severity of Respiratory Syncytial Viruses

Respiratory Syncytial Virus (RSV) disease in newborns ranges from mild symptoms to severe disease requiring hospitalization. RSV is classified into two subtypes (RSVA and RSVB) based on antigenic and genetic differences. The role these genomic variations play in disease severity remains unknown. Genome sequences were obtained using next-generation RNA sequencing on archived frozen nasal swabs of young children (< 8 months-old) infected with RSV in Rochester, NY between 1977-1998. Samples were chosen from both children hospitalized with severe RSV disease (inpatient) and those presenting with mild symptoms (outpatient) during their first cold-season. Both A and B subtypes demonstrated significant differences in the phylogeny and primary-protein structure during this time period. We found a significant association between RSV phylogeny over this time period and disease severity. For both subtypes, the G-protein demonstrated the greatest amino acid substitutions, although the number of amino acid substitutions was higher in the RSVA subtype. We found a significant association between G-protein variation and disease severity for RSVA, but not RSVB. For both subtypes, variation in the M2-2 protein was significantly associated with disease severity. These results suggest that the genetic variability of RSV proteins may contribute to disease severity in humans. ImportanceEach cold-season Respiratory Syncytial Virus (RSV) infects thousands of children in the US. Some will display mild cold symptoms while others develop severe disease, sometimes resulting in lifelong lung problems or fatality. RSV initiates infection and replicates in the nasopharynx. Substitutions in the RSV genome can be found in clinically isolated nasal-swab samples of RSV infected children. Whether these genome variations contribute to severe disease is unknown. Here we found a statistically significant association between RSV phylogeny and disease severity. Furthermore, we found specific RSV proteins (G and M2-2) whose amino acid variation was statistically associated with severe disease, although which protein was associated depended on subtype. Taken together, our results suggest that RSV genotype contributed to disease severity over this time period.

microbiology↗

bioRxiv2021

Homology mediated end joining enables efficient non-viral targeted integration of large DNA templates in primary human T cells

Adoptive cellular therapy using genetically engineered immune cells holds tremendous promise for the treatment of advanced cancers. While the number of available receptors targeting tumor specific antigens continues to grow, the current reliance on viral vectors for clinical production of engineered immune cells remains a significant bottleneck limiting translation of promising new therapies. Here, we describe an optimized methodology for efficient CRISPR-Cas9 based, non-viral engineering of primary human T cells that overcomes key limitations of previous approaches. By synergizing temporal optimization of reagent delivery, reagent composition, and integration mechanism, we achieve targeted integration of large DNA cargo at efficiencies nearing those of viral vector platforms with minimal toxicity. CAR-T cells generated using our approach are highly functional and elicit potent anti-tumor cytotoxicity in vitro and in vivo. Importantly, our method is readily adaptable to cGMP compliant manufacturing and clinical scale-up, offering a near-term alternative to the use of viral vectors for production of genetically engineered T cells for cancer immunotherapy.

bioengineering↗

bioRxiv2021

Genotyping of structural variation using PacBio high-fidelity sequencing

BackgroundStructural variations (SVs) pervade the genome and contribute substantially to the phenotypic diversity of species. However, most SVs were ineffectively assayed because of the complexity of plant genomes and the limitations of sequencing technologies. Recent advancement of third-generation sequencing technologies, particularly the PacBio high-fidelity (HiFi) sequencing, which generates both long and highly accurate reads, offers an unprecedented opportunity to characterize SVs and reveal their functionality. Since HiFi sequencing is new, it is crucial to evaluate HiFi reads in SV detection before applying the technology at scale. ResultsWe sequenced wheat genomes using HiFi, then conducted a comprehensive evaluation of SV detection using mainstream long-read aligners and SV callers. The results showed the accuracy of SV discovery depends more on aligners rather than callers. For aligners, pbmm2 and NGMLR provided the most accurate results while detecting deletion and insertion, respectively. Likewise, cuteSV and SVIM achieved the best performance across all SV callers. We demonstrated that the combination of the aligners and callers mentioned above is optimal for SV detection. Furthermore, we evaluated the impact of sequencing depth on the accuracy of SV detection. The results showed that low-coverage HiFi sequencing is capable of generating high-quality SV genotyping. ConclusionsThis study provides a robust benchmark of SV discovery with HiFi reads, showing the remarkable potential of long-read sequencing to investigate structural variations in plant genomes. The high accuracy SV discovery from low-coverage HiFi sequencing indicates that skim HiFi sequencing is an ideal approach to study structural variations at the population level.

bioinformatics↗