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Wei, L.

Publications and source records attributed to Wei, L..

At least 163 records · Page 9Linked to original sources

Antibody Upstream Sequences Diversity and Its Biological Implications Revealed by Repertoire Sequencing

The sequence upstream of antibody variable region (Antibody Upstream Sequence, or AUS) consists of 5 untranslated region (5 UTR) and two leader regions, L-PART1 and L-PART2. The sequence variations in AUS affect the efficiency of PCR amplification, mRNA translation, and subsequent PCR-based antibody quantification as well as antibody engineering. Despite their importance, the diversity of AUSs has long been neglected. Utilizing the rapid amplification of cDNA ends (5RACE) and high-throughput antibody repertoire sequencing (Rep-Seq) technique, we acquired full-length AUSs for human, rhesus macaque (RM), cynomolgus macaque (CM), mouse, and rat. We designed a bioinformatics pipeline and discovered 2,957 unique AUSs, corresponding to 2,786 and 1,159 unique sequences for 5 UTR and leader, respectively. Comparing with the leader records in the international ImMunoGeneTics (IMGT), while 529 were identical, 313 were with single nucleotide polymorphisms (SNPs), 280 were totally new, and 37 updated the incomplete records. The diversity of AUSs impact on related antibody biology was also probed. Taken together, our findings would facilitate Rep-Seq primer design for capturing antibodies comprehensively and efficiently as well as provide a valuable resource for antibody engineering and the studies of antibody at the molecular level.

bioinformatics↗

A Genome-wide CRISPR-Cas9 Screen Identifies Host Factors Essential for Optimal Plasmodium Liver Stage Development.

Prior to initiating symptomatic malaria, a single Plasmodium sporozoite infects a hepatocyte and develops into thousands of merozoites, in part by scavenging host resources. We show that host microtubules dynamically reorganize around the developing liver stage (LS) parasite. Using a genome-wide CRISPR-Cas9 screen, we identified host regulators of cytoskeleton organization, vesicle trafficking, ER/Golgi stress and lipid biogenesis that regulate Plasmodium LS development. These novel regulators of infection, including Centromere Protein J (CENPJ), led us to interrogate how microtubule organizing centers (MTOCs) are regulated during infection. Foci of {gamma}-tubulin localized to the parasite periphery; depletion of CENPJ exacerbated this re-localization and increased infection. Further, we show that the Golgi acts as a non-centrosomal MTOC by organizing {gamma}-tubulin and stimulating microtubule nucleation at the parasite periphery. Collectively, we show that the Plasmodium LS recruits the host Golgi to form MT mediated conduits along which host organelles are recruited to the PVM, to support liver stage development. Our findings suggest many host-targeted pharmacological inhibitors may inhibit LS infection.

cell biology↗

SARS-Cov-2-, HIV-1-, Ebola-neutralizing and anti-PD1 clones are predisposed

Antibody repertoire refers to the totality of the superbly diversified antibodies within an individual to cope with the vast array of possible pathogens. Despite this extreme diversity, antibodies of the same clonotype, namely public clones, have been discovered among individuals. Although some public clones could be explained by antibody convergence, public clones in naive repertoire or virus-neutralizing clones from not infected people were also discovered. All these findings indicated that public clones might not occur by random and they might exert essential functions. However, the frequencies and functions of public clones in a population have never been studied. Here, we integrated 2,449 Rep-seq datasets from 767 donors and discovered 5.07 million public clones - ~10% of the repertoire are public in population. We found 38 therapeutic clones out of 3,390 annotated public clones including anti-PD1 clones in healthy people. Moreover, we also revealed clones neutralizing SARS-CoV-2, Ebola, and HIV-1 viruses in healthy individuals. Our result demonstrated that these clones are predisposed in the human antibody repertoire and may exert critical functions during particular immunological stimuli and consequently benefit the donors. We also implemented RAPID - a Rep-seq Analysis Platform with Integrated Databases, which may serve as a useful tool for others in the field.

immunology↗

A system-level gene regulatory network model for Plasmodium falciparum

Many of the gene regulatory processes of Plasmodium falciparum, the deadliest malaria parasite, remain poorly understood. To develop a comprehensive guide for exploring this organisms gene regulatory network, we generated a system-level model of Plasmodium falciparum gene regulation using a well-validated, machine-learning approach for predicting interactions between transcription regulators and their targets. The resulting network accurately predicts expression levels of transcriptionally coherent gene regulatory programs in independent transcriptomic data sets from parasites collected by different research groups in diverse laboratory and field settings. Thus, our results indicate that our gene regulatory model has predictive power and utility as a hypothesis-generating tool for illuminating clinically relevant gene regulatory mechanisms within Plasmodium falciparum. Using the set of regulatory programs we identified, we also investigated correlates of artemisinin resistance based on gene expression coherence. We report that resistance is associated with incoherent expression across many regulatory programs, including those controlling genes associated with erythrocyte-host engagement. These results suggest that parasite populations with reduced artemisinin sensitivity are more transcriptionally heterogenous. This pattern is consistent with a model where the parasite utilizes bet-hedging strategies to diversify the population, rendering a subpopulation more able to navigate drug treatment.

systems biology↗

MicroPhenoDB associates metagenomic data with pathogenic microbes, microbial core genes, and human disease phenotypes

Microbes play important roles in human health and disease. The interaction between microbes and hosts is a reciprocal relationship, which remains largely under-explored. Current computational resources lack manually and consistently curated data to connect metagenomic data to pathogenic microbes, microbial core genes, and disease phenotypes. We developed the MicroPhenoDB database by manually curating and consistently integrating microbe-disease association data. MicroPhenoDB provides 5677 non-redundant associations between 1781 microbes and 542 human disease phenotypes across more than 22 human body sites. MicroPhenoDB also provides 696,934 relationships between 27,277 unique clade-specific core genes and 685 microbes. Disease phenotypes are classified and described using the Experimental Factor Ontology (EFO). A refined score model was developed to prioritize the associations based on evidential metrics. The sequence search option in MicroPhenoDB enables rapid identification of existing pathogenic microbes in samples without running the usual metagenomic data processing and assembly. MicroPhenoDB offers data browsing, searching and visualization through user-friendly web interfaces and web service application programming interfaces. MicroPhenoDB is the first database platform to detail the relationships between pathogenic microbes, core genes, and disease phenotypes. It will accelerate metagenomic data analysis and assist studies in decoding microbes related to human diseases. MicroPhenoDB is available through http://www.liwzlab.cn/microphenodb and http://lilab2.sysu.edu.cn/microphenodb.

bioinformatics↗

TASK-3 Two-Pore Potassium Channels drive neuronal excitability of the circadian clock and entrainment to challenging light environments

The suprachiasmatic nucleus (SCN), the mammalian circadian clock, is a heterogenous structure made of several neuron types that generate a circadian electrical activity profile. However, it is unclear how such regulation in endogenous neuronal excitability is maintained. Background two-pore domain potassium channels (K2P), such as TASK-3, play an important role in inhibiting neuronal activity. Here, we utilize a TASK-3 KO mouse model to unravel the role played by this channel in SCN circadian neuronal regulation and behavioral photoentrainment. Our results reveal that TASK-3 is needed to adapt to challenging lighting conditions, such as those experienced through seasonal changes and jet lag. From our investigations this appears to be very distinct from pathways that drive acute, one-off adjustments in clock phase, in response to single pulses of light. These findings provide crucial information on the intricate pathways linking clock output to behavioral adaptation to light-dark cycles.

neuroscience↗

ScRNA-seq discover cell cluster change under OAB: ACE2 expression reveal possible alternation of 2019-nCoV infectious pathway

ObjectivePrevious study indicated that bladder cells which express ACE2 were a potential infection route of 2019-nCov. This study observed some differences of bladder cell cluster and their ACE2 expression between OAB mice and healthy mice, indicating the change of infectious possibility and pathway under overactive bladder (OAB) circumstance. Material and methodPubic dataset acquisition was used to get ACE2 expression in normal human bladder and mice bladder (GSE129845). We built up over OAB model and studied the impact on cell typing and ACE2 expression. By way of using single-cell RNA sequencing (scRNA-seq) technique, bladder cell clustering and ACE2 expression in various cell types were measured respectively. ResultIn pubic database (healthy human and mice bladder), ACE2 expression in humans and mice is concentrated in bladder epithelial cells. The disappearance of umbrella cells, a component of bladder epithelial, was found in our OAB model. In the two mouse bladder samples, ACE2 expression of epithelial cells is 34.1%, also the highest of all cell types. ConclusionThe disappearance of umbrella cell may alternate the infection pathway of 2019-nCov and relate to the onset and progression of OAB.

cell biology↗

Microfluidics-enabled 96-well perfusion system for high-throughput tissue engineering and long-term all-optical electrophysiology

This work demonstrates a novel high-throughput (HT) microfluidics-enabled uninterrupted perfusion system (HT-UPS) and validates its use with chronic all-optical electrophysiology in human excitable cells. HT-UPS consists of a soft multichannel microfluidic plate cover which could button on a commercial HT 96-well plate. Herein, we demonstrate the manufacturing process of the system and its usages in acute and chronic all-optical electrophysiological studies of human induced pluripotent stem-cell-derived cardiomyocytes (iPSC-CM) and engineered excitable (Spiking HEK) cells. HT-UPS perfusion maintained functional voltage and calcium responses in iPSC-CM and Spiking HEK cells under spontaneous conditions and under optogenetic pacing. Long-term culture with HT-UPS improved cell viability and optogenetically-tracked calcium responses in Spiking HEK cells. The scalability and simplicity of this design and its compatibility with HT all-optical electrophysiology can empower cell-based assays for personalized medicine using patient-derived cells.

bioengineering↗

Potential microenvironment of SARS-CoV-2 infection in airway epithelial cells revealed by Human Protein Atlas database analysis

The outbreak of COVID-19 has caused serious epidemic events in China and other countries. With the rapid spread of COVID-19, it is urgent to explore the pathogenesis of this novel coronavirus. However, the foundational research of COVID-19 is very weak. Although angiotensin converting enzyme 2 (ACE2) is the reported receptor of SARS-CoV-2, information about SARS-CoV-2 invading airway epithelial cells is very limited. Based on the analysis of the Human Protein Atlas database, we compared the virus-related receptors of epithelial-derived cells from different organs and found potential key molecules in the local microenvironment for SARS-CoV-2 entering airway epithelial cells. In addition, we found that these proteins were associated with virus reactive proteins in host airway epithelial cells, which may promote the activation of the immune system and the release of inflammatory factors. Our findings provide a new research direction for understanding the potential microenvironment required by SARS-CoV-2 infection in airway epithelial, which may assist in the discovery of potential drug targets against SARS-CoV-2 infection.

cell biology↗

Obstructed tear duct causes epiphora and precocious eyelid opening due to disruption of Prickle 1-mediated Wnt/PCP signaling

The tear drainage apparatus evolved in terrestrial animals serving as conduits for tear flow. Obstruction of tear drainage causes a range of ocular surface disorders. Hitherto, genetics of tear duct development and obstruction has been scarcely explored. Here we report that a severe Prickle 1 hypomorph mouse line exhibited epiphora. This phenotype was due to blockage of the tear drainage by the incompletely formed nasolacrimal duct (NLD) and lacrimal canaliculi (CL). Further analysis revealed that the precocious eyelid opening, previously observed in the same type of Prickle 1 mutants, is also caused by tear duct dysplasia. A comparison of wild type, the Prickle 1 hypomorph and null mutants revealed a dose-dependent requirement of Prickle 1 for tear duct outgrowth. As a key component of a set of six Wnt/PCP core proteins, Prickle 1 usually works together with other PCP components. An investigation of expression of Wnt/PCP core genes demonstrated three of the six PCP components in tear duct, supporting the notion of context-dependent organization of PCP protein complexes. Furthermore, expression of Fgfr2/Fgf10 and p63 genes, mutations of which are associated with NLD and CL hypoplasia in human, were not altered in Prickle 1 mutant mice. Lastly, we showed that Prickle 1 expression in developing tear drainage system is conserved between mouse and human despite anatomical differences. Altogether, the study uncovered how obstruction of the tear drainage could lead to a complex ocular surface disorder, which may have genetic implications in human ocular health.

genetics↗

Genomic Architecture of Cells in Tissues (GeACT): Study of Human Mid-gestation Fetus

By circumventing cellular heterogeneity, single cell omics have now been widely utilized for cell typing in human tissues, culminating with the undertaking of human cell atlas aimed at characterizing all human cell types. However, more important are the probing of gene regulatory networks, underlying chromatin architecture and critical transcription factors for each cell type. Here we report the Genomic Architecture of Cells in Tissues (GeACT), a comprehensive genomic data base that collectively address the above needs with the goal of understanding the functional genome in action. GeACT was made possible by our novel single-cell RNA-seq (MALBAC-DT) and ATAC-seq (METATAC) methods of high detectability and precision. We exemplified GeACT by first studying representative organs in human mid-gestation fetus. In particular, correlated gene modules (CGMs) are observed and found to be cell-type-dependent. We linked gene expression profiles to the underlying chromatin states, and found the key transcription factors for representative CGMs. HighlightsO_LIGenomic Architecture of Cells in Tissues (GeACT) data for human mid-gestation fetus C_LIO_LIDetermining correlated gene modules (CGMs) in different cell types by MALBAC-DT C_LIO_LIMeasuring chromatin open regions in single cells with high detectability by METATAC C_LIO_LIIntegrating transcriptomics and chromatin accessibility to reveal key TFs for a CGM C_LI

genomics↗

miRNAs and neural alternative polyadenylation specify the virgin behavioral state

How are diverse regulatory strategies integrated to impose appropriately patterned gene expression that underlie in vivo phenotypes? Here, we reveal how coordinated miRNA regulation and neural-specific alternative polyadenylation (APA) of a single locus controls complex behaviors. Our entry was the unexpected observation that deletion of Bithorax-Complex (BX-C) miRNAs converts virgin female flies into a subjective post-mated behavioral state, normally induced by seminal proteins following copulation. Strikingly, this behavioral switch is directly attributable to misregulation of homothorax (hth). We localize specific CNS abdominal neurons where de-repressed Hth compromises virgin behavior in BX-C miRNA mutants. Moreover, we use genome engineering to demonstrate that precise mutation of hth 3UTR sites for BX-C miRNAs, or deletion of its neural 3 UTR extension containing most of these sites, both induce post-mated behaviors in virgins. Thus, facilitation of miRNA-mediated repression by neural APA is required for virgin females to execute behaviors appropriate to their internal state.

neuroscience↗

MosaicBase: A Knowledgebase of Postzygotic Mosaic Variants in Noncancer Diseases and Asymptomatic Human Individuals

Mosaic variants resulting from postzygotic mutations are prevalent in the human genome and play important roles in human diseases. However, except for cancer-related variant collections, there are no collections of mosaic variants in noncancer diseases and asymptomatic individuals. Here, we present MosaicBase (http://mosaicbase.cbi.pku.edu.cn/ or http://49.4.21.8:8000/), a comprehensive database that includes 6,698 mosaic variants related to 269 noncancer diseases and 27,991 mosaic variants identified in 422 asymptomatic individuals. The genomic and phenotypic information for each variant was manually extracted and curated from 383 publications. MosaicBase supports the query of variants with Online Mendelian Inheritance in Man (OMIM) entries, genomic coordinates, gene symbols, or Entrez IDs. We also provide an integrated genome browser for users to easily access mosaic variants and their related annotations within any genomic region. By analyzing the variants collected in MosaicBase, we found that mosaic variants that directly contribute to disease phenotype showed features distinct from those of variants in individuals with a mild or no phenotype in terms of their genomic distribution, mutation signatures, and fraction of mutant cells. MosaicBase will not only assist clinicians in genetic counseling and diagnosis but also provide a useful resource to understand the genomic baseline of postzygotic mutations in the general human population.

bioinformatics↗

Ultra-deep sequencing differentiates patterns of skin clonogenic mutations associated with sun-exposure status

Non-melanoma skin cancer is the most common human malignancy and is primarily caused by exposure to ultraviolet (UV) radiation. The earliest detectable precursor of UV-mediated skin cancer is the growth of cell groups harboring clonal mutation (CM) in clinically normal appearing skin. Systematic evaluation of CMs is crucial to understand early photo-carcinogenesis. Previous studies confirmed the presence of CMs in sun-exposed skin. However, the relationship between UV-exposure and the accumulation of CMs, and the correlation of CMs with skin cancer risk remain poorly understood. To elucidate the exact molecular and clinical effects of long-term UV-exposure on skin, we performed targeted ultra-deep sequencing in 450 individual-matched sun-exposed (SE) and non-sun-exposed (NE) epidermal punch biopsies obtained from clinically normal skin from 13 donors. A total of 638 CMs were identified, including 298 UV-signature mutations (USMs). The numbers of USMs per sample were three times higher in the SE samples and were associated with significantly higher variant allele frequencies (VAFs), compared with the NE samples. We identified genomic regions in TP53, NOTCH1 and GRM3 where mutation burden was significantly associated with UV-exposure. Six mutations were almost exclusively present in SE epidermis and accounted for 42% of the overall difference between SE and NE mutation burden. We defined Cumulative Relative Clonal Area (CRCA), a single metric of UV-damage calculated by the overall relative percentage of the sampled skin area affected by CMs. The CRCA was dramatically elevated by a median of 11.2 fold in SE compared to NE samples. In an extended cohort of SE normal skin samples from patients with a high- or low-burden of cutaneous squamous cell carcinoma (cSCC), the SE samples in high-cSCC patients contained significantly more USMs than SE samples in low-cSCC patients, with the difference mostly conferred by mutations from low-frequency clones (defined by VAF≤1%) but not expanded clones (VAF>1%). Our studies of differential mutational features in normal skin between paired SE/NE body sites and high/low-cSCC patients provide novel insights into the carcinogenic effect of UV exposure, and suggest CMs might be used to develop novel biomarkers for predicting cancer risk.Significance statement In UV radiation exposed skin, mutations fuel clonal cell growth. We established a sequencing-based method to objectively assess the mutational differences between sun-exposed (SE) and non-sun-exposed (NE) areas of normal human skin. Striking differences, in both the numbers of mutations and variant allele frequencies, were found between SE and NE areas. Furthermore, we identified specific genomic regions where mutation burden is significantly associated with UV-exposure status. These findings revealed previously unknown mutational patterns associated with UV-exposure, providing important insights into UV radiation’s early carcinogenic effects. Additionally, in an extended cohort, we identified preliminary association between normal skin mutation burden and cancer risk. These findings pave the road for future development of quantitative measurement of subclinical UV damage and skin cancer risk.Competing Interest StatementThe authors have declared no competing interest.Common AbbreviationsUVUltravioletCMClonogenic mutationNMSCNonmelanoma skin cancerSESun-exposedNENon-sun-exposedUSMUV-signature mutationNUSMNon-UV-signature mutationCRCACumulative Relative Clonal AreacSCCCutaneous squamous cell carcinomaAKActinic keratosisSNVSingle nucleotide variant Indels – Insertions/deletionsDNVDinucleotide variantCSNVCluster of single nucleotide variantMACMulti-Nucleotide Variant Annotation CorrectorVAFVariant allele frequencyView Full Text

cancer biology↗

Live-Cell Imaging and Quantification of PolyQ Aggregates by Stimulated Raman Scattering of Selective Deuterium Labeling

Huntingtons disease, a major neurodegenerative disorder, involves deposition of aggregation-prone proteins with long polyglutamine (polyQ) expansions. The ability to non-perturbatively visualize the formation of aggregates could offer new molecular insight for their pathologic roles. Here, we propose stimulated Raman scattering imaging of deuterium-labeled glutamine to investigate native polyQ aggregates in live cells with subcellular resolution. Through the enrichment of deuterated glutamine in the polyQ sequence of mutant Huntingtin (mHtt) proteins, we first achieved sensitive and specific SRS imaging of carbon-deuterium bonds (C-D) from aggregates without GFP labeling. These aggregates become 1.8-fold denser compared to those with GFP. Second, we performed ratiometric quantification, which revealed a dependence of protein compositions on aggregation sizes. Moreover, we calculated the absolute concentrations for sequestered mHtt and non-mHtt proteins within the same aggregates. Our method may readily reveal new features of polyQ aggregates and could be suited for in vivo investigations on multicellular organisms.

biophysics↗

HPV-positive head and neck cancer derived exosomal miR-9 induces M1 macrophage polarization and increases tumor radiosensitivity

Human papillomavirus (HPV) is an etiological risk factor for a subset of head and neck squamous cell carcinoma (HNSCC). HPV+ HNSCC is significant more radiosensitive than HPV-HNSCC, but the underlying mechanism is still unknown. Tumor microenvironment can affect tumor response to radiation therapy. Cancer secreted exosomes are emerging as crosstalk mediators between tumor cells and the tumor microenvironment. The main objectives of this study were to determine the role of HPV+ HNSCC-derived exosomes in increased radiation sensitivity. Here, we found that exosomes derived from HPV+ HNSCC cells activate macrophages into the M1 phenotype, which then increases the radiosensitivity of HNSCC cells. miR-9 was enriched in exosomes released from HPV+ HNSCC cells and it could be transported to macrophages, leading to altered cellular functions. Overexpression of miR-9 in macrophages induced polarization into the M1 phenotype via downregulation of PPAR{delta}. Increased radiosensitivity was observed for HNSCC cells co-cultured with macrophages in which miR-9 was upregulated or treated with M1 macrophages. These observations suggest that HPV+ HNSCC cells secrete miR-9-rich exosomes, which then polarize macrophages into M1 phenotype and lead to increased radiosensitivity of HNSCC cells. Hence, miR-9 may be a potential treatment strategy for HNSCC.\n\nStatement of significanceHPV+ HNSCC through the release of miR-9-rich exosomes polarize macrophages into M1 phenotype and lead to increased radiosensitivity of HNSCC.

cancer biology↗

Large-scale Analysis of 2,152 dataset reveals key features of B cell biology and the antibody repertoire

Antibody repertoire sequencing (Ig-seq) has been widely used in studying humoral responses, with promising results. However, the promise of Ig-seq has not yet been fully realized, and key features of the antibody repertoire remain elusive or controversial. To clarify these key features, we analyzed 2,152 high-quality heavy chain antibody repertoires, representing 582 donors and a total of 360 million clones. Our study revealed that individuals exhibit very similar gene usage patterns for germline V, D, and J genes and that 53 core V genes contribute to more than 99% of the heavy chain repertoire. We further found that genetic background is sufficient but not necessary to determine usage of V, D, and J genes. Although gene usage pattern is not affected by age, we observed a significant sex preference for 24 V genes, 9 D genes and 5 J genes, but found no positional bias for V-D and D-J recombination. In addition, we found that the number of observed clones that were shared between any two repertoires followed a linear model and noted that the mutability of hot/cold spots and single nucleotides within antibody genes suggested a strand-specific somatic hypermutation mechanism. This population-level analysis resolves some critical characteristics of the antibody repertoire and thus may serve as a reference for research aiming to unravel B cell-related biology or diseases. The metrics revealed here will be of significant value to the large cadre of scientists who study the antibody repertoire.

immunology↗

Bootstrapping promotes the RSFC-behavior associations: an application of individual cognitive traits prediction

Resting state functional connectivity records enormous functional interaction information between any pair of brain nodes, which enriches the prediction of individual phenotypes. To reduce the high dimensional features in prediction, correlation analysis is a common way for feature selection. However, rs-fMRI signal exhibits typically low signal-to-noise ratio and correlation analysis is sensitive to outliers and data distribution, which may bring unstable and uninformative features to subsequent prediction. To alleviate this problem, a bootstrapping-based feature selection framework was proposed and applied on three widely used regression models: connectome-based predictive model (CPM), support vector regression (SVR) and least absolute shrinkage and selection operator (LASSO). A large open-source dataset from Human Connectome Project (HCP) was adopted in the study and a series of cognitive traits were acted as the prediction targets. To systematically investigate the influences of different parameter settings on the bootstrapping-based framework, a total of 216 parameter combinations were evaluated through the R value between the predicted and real cognitive traits, and the best identified performance among them was chosen out as the final prediction accuracy for each cognitive trait. By using bootstrapping without replacement, the best performances of CPM with positive and negative feature sets, SVR and LASSO averagely increased by 28.0%, 33.2%, 11.6% and 24.3% in R values in contrast to the baseline method without bootstrapping. By using bootstrapping with replacement, these best performances increased by 22.1%, 22.9%, 9.4% and 19.6%. Furthermore, the bootstrapping-based feature selection methods could effectively refine the original feature sets obtained from correlation analysis, which thus retained the more stable and informative feature sets. The results demonstrate that bootstrapping-based feature selection is an easy-to-use and effective method to improve RSFC prediction of cognitive traits and is highly recommended in future RSFC prediction studies.

neuroscience↗