Biology subjects

Wang, D.

Publications and source records attributed to Wang, D..

At least 721 records · Page 40Linked to original sources

bioRxiv2019

M-GWAS for the gut microbiome in Chinese adults illuminates on complex diseases

The gut microbiome has been established as a key environmental factor to health. Genetic influences on the gut microbiome have been reported, yet, doubts remain as to the significance of genetic associations. Here, we provide shotgun data for whole genome and whole metagenome from a Chinese cohort, identifying no less than 20% genetic contribution to the gut microbiota. Using common variants-, rare variants- and copy number variations (CNVs)-based association analyses, we identified abundant signals associated with the gut microbiome especially in metabolic, neurological and immunological functions. The controversial concept of enterotypes may have a genetic attribute, with the top 2 loci explaining 11% of the Prevotella-Bacteroides variances. Stratification according to gender led to the identification of differential associations in males and females. Genetically encoded responses to ectopic presence of oral bacteria in the gut appear to be a common theme in a number of diseases investigated by MWAS (Metagenome-wide association studies). Our two-stage M-GWAS (Microbiome genome-wide association studies) on a total of 1295 individuals unequivocally illustrates that neither microbiome nor GWAS studies could overlook one another in our quest for a better understanding of human health and diseases.\n\nHighlightsM-GWAS using high-depth whole genome identifies contributions from rare variants and CNVs.\n\nGut microbial modules such as butyrate, amino acids, mucin degradation show genetic associations.\n\nGender differential M-GWAS underscores differences in metabolic and psychological predispositions.\n\nSome of the MWAS markers for colorectal cancer and cardiometabolic diseases show genetic associations.

genetics↗

bioRxiv2019

Widespread transcriptional responses to the thermal stresses are prewired in human 3D genome

Temperature changes is one of the most common environmental stress that consequences with massive phenotypic responses for almost all the life forms. The dysregulation of heat shock (HS) response genes had been found associated with various severe diseases, including cancer. Although the HS response has been well studied in animal cells, it remains elusive whether or not the cells response to cold shock (CS) similarly. Here, we comprehensively compared the changes of gene expression, epigenetic marks (H3K4me3 and H3K27ac), binding of genome architecture proteins (CTCF, SMC3 and Pol II) and chromatin conformation after HS and CS in human cells. Widespread expression change was observed after both HS and CS. Remarkably, we identified distinguished characters in those thermal stress responded genes at nearly all levels of chromatin architecture, i.e, the compartment, topological associated domain, chromatin loops and transcription elongation regulators, in the normal condition. However, the global chromatin architecture remains largely stable after both CS and HS. Interestingly, the thermal stresses responded genes are prone to spatial clustering even before the temperature changes. Our data suggested that the transcriptional response to the thermal stresses maybe independent to the changes of the high-level chromatin architecture, e.g., compartments and TAD, while it may be more dependent on the precondition of the chromatin and epigenetic settings at the normal condition.

genomics↗

bioRxiv2019

Characterization of TAG-63 and its role on axonal transport in C. elegans

Model organisms are increasingly used to study and understand how neurofilament (NF)-based neurological diseases develop. However, whether a NF homolog exists in C. elegans remains unclear. We characterize TAG-63 as a NF-like protein with sequence homologies to human NEFH carrying various coiled coils as well as clustered phosphorylation sites. TAG-63 also exhibits features of NFL such as a molecular weight of around 70 kD, the lack of KSP repeats and the ability to form 10 nm filamentous structures in transmission electron micrographs. An anti-NEFH antibody detects a band at the predicted molecular weight of TAG-63 in Western blots of whole worm lysates and this band cannot be detected in tag-63 knockout worms. A transcriptional tag-63 reporter expresses in a broad range of neurons, and various anti-NFH antibodies stain worm neurons with an overlapping expression of axonal vesicle transporter UNC-104(KIF1A). Cultured neurons grow shorter axons when incubating with drugs known to disintegrate the NF network and rhodamine-labeled in vitro reconstituted TAG-63 filaments disintegrate upon drug exposure. Speeds of UNC-104 motors are diminished in tag-63 mutant worms with visibly increased accumulations of motors along axons. UNC-104/TAG-63 and SNB-1/TAG-63 not only co-localize in neurons but also revealed positive BiFC (bimolecular fluorescence assay) signals.

cell biology↗

bioRxiv2019

Enhance genome editing efficiency and specificity by a quick CRISPR/Cas9 system

CRISPR/Cas9 is a powerful genome editing tool that has been successfully applied to a variety of species, including zebrafish. However, targeting efficiencies vary greatly at different genomic loci, the underlying causes of which were still elusive. Here we report a quick CRISPR/Cas9 system, designated as qCas9, which exhibits accelerated turnover of Cas9 protein in zebrafish. Our data showed that qCas9 significantly improved targeting efficiency, including both knock-out and knock-in in F0 embryos, and yielded higher germline transmission rate in founder screen. Importantly, qCas9 showed little to no off-target editing in zebrafish and profoundly reduced off-target effect in HEK293T cell line. In summary, our findings demonstrate that qCas9 is a simple, economic and highly effective method to improve genome editing efficiency in zebrafish embryos and also holds great potential in reducing off-target effect in mammalian cell lines.

genetics↗

bioRxiv2019

An integrative ENCODE resource for cancer genomics

ENCODE comprises thousands of functional genomics datasets, and the encyclopedia covers hundreds of cell types, providing a universal annotation for genome interpretation. However, for particular applications, it may be advantageous to use a customized annotation. Here, we develop such a custom annotation by leveraging advanced assays, such as eCLIP, Hi-C, and whole-genome STARR-seq on a number of data-rich ENCODE cell types. A key aspect of this annotation is comprehensive and experimentally derived networks of both transcription factors and RNA-binding proteins (TFs and RBPs). Cancer, a disease of system-wide dysregulation, is an ideal application for such a network-based annotation. Specifically, for cancer-associated cell types, we put regulators into hierarchies and measure their network change (rewiring) during oncogenesis. We also extensively survey TF-RBP crosstalk, highlighting how SUB1, a previously uncharacterized RBP, drives aberrant tumor expression and amplifies the effect of MYC, a well-known oncogenic TF. Furthermore, we show how our annotation allows us to place oncogenic transformations in the context of a broad cell space; here, many normal-to-tumor transitions move towards a stem-like state, while oncogene knockdowns show an opposing trend. Finally, we organize the resource into a coherent workflow to prioritize key elements and variants, in addition to regulators. We showcase the application of this prioritization to somatic burdening, cancer differential expression and GWAS. Targeted validations of the prioritized regulators, elements and variants using siRNA knockdowns, CRISPR-based editing, and luciferase assays demonstrate the value of the ENCODE resource.

genomics↗

bioRxiv2019

The C. elegans RIG-I homolog drh-1 mediates the Intracellular Pathogen Response upon viral infection

Mammalian RIG-I-like receptors detect viral dsRNA and 5 triphosphorylated RNA to activate transcription of interferon genes and promote antiviral defense. The C. elegans RIG-I-like receptor DRH-1 promotes defense through antiviral RNA interference, but less is known about its role in regulating transcription. Here we describe a role for drh-1 in directing a transcriptional response in C. elegans called the Intracellular Pathogen Response (IPR), which is associated with increased pathogen resistance. The IPR includes a set of genes induced by diverse stimuli including intracellular infection and proteotoxic stress. Previous work suggested that the proteotoxic stress caused by intracellular infections might be the common trigger of the IPR, but here we demonstrate that different stimuli act through distinct pathways. Specifically, we demonstrate that DRH-1/RIG-I is required for inducing the IPR in response to Orsay virus infection, but not in response to other triggers like microsporidian infection or proteotoxic stress. Furthermore, drh-1 appears to be acting independently of its known role in RNAi. Interestingly, expression of the replication competent Orsay virus RNA1 segment alone is sufficient to induce most of the IPR genes in a manner dependent on RNA dependent RNA polymerase activity and on drh-1. Altogether, these results suggest that DRH-1 is a pattern-recognition receptor that detects viral replication products to activate the IPR stress/immune program in C. elegans.\n\nImportanceC. elegans lacks homologs of most mammalian pattern recognition receptors, and how nematodes detect pathogens is poorly understood. We show that the C. elegans RIG-I homolog drh-1 mediates induction of the Intracellular Pathogen Response (IPR), a novel transcriptional defense program, in response to infection by the natural C. elegans viral pathogen Orsay virus. drh-1 appears to act as a pattern-recognition receptor to induce the IPR transcriptional defense program by sensing the products of viral RNA-dependent RNA polymerase activity. Interestingly, this signaling role of drh-1 is separable from its previously known role in antiviral RNAi. In addition, we show that there are multiple host pathways for inducing the IPR, shedding light on the regulation of this novel transcriptional immune response.

microbiology↗

bioRxiv2019

Dirac mixture distributions for the approximation of mixed effects models

Mixed effect modeling is widely used to study cell-to-cell and patient-to-patient variability. The population statistics of mixed effect models is usually approximated using Dirac mixture distributions obtained using Monte-Carlo, quasi Monte-Carlo, and sigma point methods. Here, we propose the use of a method based on the Cramer-von Mises Distance, which has been introduced in the context of filtering. We assess the accuracy of the different methods using several problems and provide the first scalability study for the Cramer-von Mises Distance method. Our results indicate that for a given number of points, the method based on the modified Cramer-von Mises Distance method tends to achieve a better approximation accuracy than Monte-Carlo and quasi Monte-Carlo methods. In contrast to sigma-point methods, the method based on the modified Cramer-von Mises Distance allows for a flexible number of points and a more accurate approximation for nonlinear problems.

systems biology↗

bioRxiv2019

Maintenance of cell type-specific connectivity and circuit function requires Tao kinase

Sensory circuits are typically established during early development, yet how circuit specificity and function are maintained during organismal growth has not been elucidated. To gain insight we quantitatively investigated synaptic growth and connectivity in the Drosophila nociceptive network during larval development. We show that connectivity between primary nociceptors and their downstream neurons scales with animal size. We further identified the conserved Ste20-like kinase Tao as a negative regulator of synaptic growth required for maintenance of circuit specificity and connectivity. Loss of Tao kinase resulted in exuberant postsynaptic specializations and aberrant connectivity during larval growth. Using functional imaging and behavioral analysis we show that loss of Tao-induced ectopic synapses with inappropriate partner neurons are functional and alter behavioral responses in a connection-specific manner. Our data show that fine-tuning of synaptic growth by Tao kinase is required for maintaining specificity and behavioral output of the neuronal network during animal growth.

neuroscience↗

bioRxiv2019

ALI-1, candidate gene of B1 locus, is associated with awn length and grain weight in common wheat

Awn plays a vital role in the photosynthesis, grain production and drought tolerance of common wheat; however, works on the systematic identification or cloning of genes controlling wheat awn length (AL) were seldom reported. Here, we conducted the Genome-wide association study (GWAS) in 364 wheat accessions and identified 25 loci involved in the AL, including dominant awn suppressors B1, B2 and four homologs of awn controlling genes in rice and barley. Furthermore, the B1 locus was mapped to a 125-kb physical interval harboring two genes on chromosome 5AL through map-based cloning. As the candidate gene for B1 locus, a C2H2 zinc finger gene Awn Length Inhibitor 1 (ALI-1) expressed predominantly in the developing spike of awnless individuals and suppresses downstream genes transcriptionally. ALI-1 reduces cytokinin content and simultaneously restrains cytokinin signal transduction, which leads to a stagnation of cell proliferation and reduction of cell number in awn. Noteworthily, ali-1 was the first awn controlling locus that observed increasing grain length in wheat, which is a valuable supplemental attribution of awn on grain weight besides photosynthesis. Thus, ALI-1 pleiotropically regulates awn and grain development, and this work provides a strategy to achieve improved grain yield and address future extreme climate.\n\nHighlightALI-1, candidate gene of awn suppressing B1 locus, associates with awn length and grain length, providing a reacquaint of the effect of wheat awn on grain production.

plant biology↗

bioRxiv2019

Dysfunction of ventral tegmental area GABA neurons causes mania-like behavior

The ventral tegmental area (VTA), an important source of dopamine, regulates goal- and reward-directed and social behaviors, wakefulness and sleep. Hyperactivation of dopamine neurons generates behavioral pathologies. But any roles of non-dopamine VTA neurons in psychiatric illness have been little explored. Lesioning or chemogenetically inhibiting VTA GABAergic (VTAVgat) neurons generated persistent wakefulness with mania-like qualities: locomotor activity was increased; sensitivity to D-amphetamine was heightened; immobility times decreased on the tail suspension and forced swim tests; and sucrose preference increased. Furthermore, after sleep deprivation, mice with lesioned VTAVgat neurons did not catch up on the lost NREM sleep, even though they were starting from an already highly sleep-deprived baseline, suggesting that the sleep homeostasis process was bypassed. The mania-like behaviors, including the sleep loss, were reversed by the mood-stabilizing drug valproate, and re-emerged when valproate treatment was stopped. Lithium salts, however, had no effect. The mania like-behaviors partially depended on dopamine, because giving D1/D2/D3 receptor antagonists partially restored the behaviors, but also on VTAVgat projections to the lateral hypothalamus (LH). Optically or chemogenetically inhibiting VTAVgat terminals in the LH elevated locomotion and decreased immobility time during the tail suspension and forced swimming tests. VTAVgat neurons are centrally positioned to help set an animals (and humans) level of mental and physical activity. Inputs that inhibit VTAVgat neurons intensify wakefulness (increased activity, enhanced alertness and motivation), qualities useful for acute survival. Taken to the extreme, however, decreased or failed inhibition from VTAVgat neurons produces mania-like qualities (hyperactivity, hedonia, decreased sleep).

neuroscience↗

bioRxiv2019

Genomic loci susceptible to systematic sequencing bias in clinical whole genomes

Accurate massively parallel sequencing (MPS) of genetic variants is key to many areas of science and medicine, such as cataloguing population genetic variation and diagnosing genetic diseases. Certain genomic positions can be prone to higher rates of systematic sequencing and alignment bias that limit accuracy, resulting in false positive variant calls. Current standard practices to differentiate between loci that can and cannot be sequenced with high confidence utilise consensus between different sequencing methods as a proxy for sequencing confidence. These practices have significant limitations and alternative methods are required to overcome these. We have developed a novel statistical method based on summarising sequenced reads from whole genome clinical samples and cataloguing them in "Incremental Databases" that maintain individual confidentiality. Allele statistics were catalogued for each genomic position that consistently showed systematic biases with the corresponding MPS sequencing pipeline. We found systematic biases present at [~]1-3% of the human autosomal genome across five patient cohorts. We identified which genomic regions were more or less prone to systematic biases, including large homopolymer flanks (odds ratio=23.29-33.69) and the NIST high confidence genomic regions (odds ratio=0.154-0.191). We confirmed our predictions on a gold-standard reference genome and showed that these systematic biases can lead to suspect variant calls within clinical panels. Our results recommend increased caution to address systematic biases in whole genome sequencing and alignment. This study provides the implementation of a simple statistical approach to enhance quality control of clinically sequenced samples by flagging variants at suspect loci for further analysis or exclusion.

bioinformatics↗

bioRxiv2019

Diverse noncanonical PAMs recognized by SpCas9 in human cells

The CRISPR/Cas9 system derived from Streptococcus pyogenes (SpCas9) provides unprecedented genome editing capabilities, but the potential for off-target mutations limits its application. In addition to NGG protospacer adjacent motif (PAM), off-target mutations are also associated with noncanonical PAMs, which have not yet been systematically evaluated. Here, we developed a highly sensitive approach that allows systematically analyzing PAM sequences in human cells, and identified multiple alternative PAMs recognized by SpCas9.

bioengineering↗

bioRxiv2019

Mesenchymal stem cells restore local microenvironment and systemically suppress leukemia via reprogramming macrophages

Bone marrow (BM) mesenchymal stem cells (MSCs) are critical components of the BM microenvironment and play an essential role in supporting hematopoiesis. Dysfunction of MSCs is associated with the impaired BM microenvironment that promotes leukemia development. However, whether and how restoration of the impaired BM microenvironment can inhibit leukemia development remain unknown. Using an established leukemia model and the RNA-seq analysis, we discovered functional degeneration of MSCs during leukemia progression. Importantly, intra-BM instead of systemic transfusion of donor healthy MSCs restored the BM microenvironment, thus systemically altering cytokine expression patterns, improving normal hematopoiesis, reducing tumor burden, and ultimately prolonging survival of the leukemia-bearing mice. Donor MSC treatment restored the function of host MSCs and reprogrammed host macrophages to fulfill tissue-repair function. Transfusion of MSC-reprogrammed macrophages largely recapitulated the therapeutic effects of MSCs. Further, we found that donor MSCs reprogrammed macrophages to reduce leukemia burden through autocrine of IL-6. Taken together, our study reveals that donor MSCs reprogram host macrophages to restore the BM microenvironment and inhibit leukemia development, thus offering rationales for local MSC administration as a potentially effective therapy for leukemia.\n\nKey PointsKey Point 1: Intra-BM transfusion of MSCs restores the BM microenvironment, improves thrombopoiesis, and suppresses MDS/MPN initiated by Nras mutation.\n\nKey Point 2: Donor MSCs reprogram macrophages to restore the BM microenvironment, improve thrombopoiesis, and suppress leukemia.

cancer biology↗

bioRxiv2019

Pneumococcal Colonisation is an Asymptomatic Event in Healthy Adults using an Experimental Human Colonisation Model.

IntroductionPneumococcal colonisation is regarded as a pre-requisite for developing pneumococcal disease. In children previous studies have reported colonisation to be a symptomatic event and described a relationship between symptom severity/frequency and colonisation density. The evidence for this in adults is lacking in the literature. This study uses an experimental human pneumococcal challenge model to explore whether pneumococcal colonisation (or co-colonisation with a respiratory virus) is a symptomatic event in healthy adults.\n\nMethodsHealthy volunteers aged 18-50 were recruited and inoculated intra-nasally with either Streptococcus pneumoniae (serotypes 6B, 23F) or saline as a control. Respiratory viral swabs were obtained prior to inoculation. Nasal and non-nasal symptoms were then assessed using a modified Likert score between 1 (no symptoms) to 7 (cannot function). The rate of symptoms reported between groups was compared and a correlation analysis performed.\n\nResultsData from 54 participants were analysed. 46 were inoculated with S. pneumoniae (29 with 6B, 17 with 23F) and 8 received saline. In total, 14 became experimentally colonised (30.4%), all of which were inoculated with 6B serotype. There was no statistically significant difference in nasal (p= 0.45) or non-nasal symptoms (p=0.28) between the pneumococcal inoculation group and the saline group. There was no direct correlation between colonisation density and symptom severity in those who were colonised. In the 22% (12/52) who were co-colonised with pneumococcus and respiratory viruses there was no statistical difference in either nasal or non-nasal symptoms (virus positive p=0.74 and virus negative p=1.0).\n\nConclusionPneumococcal colonisation is asymptomatic in healthy adults, regardless of bacterial density or viral co-colonisation.

microbiology↗

bioRxiv2019

Age-dependent sexual dimorphism in the adult human gut microbiota

A decade of studies has established the importance of the gut microbiome in human health. In spite of sex differences in the physiology, lifespan, and prevalence of many age-associated diseases, sex and age disparities in the gut microbiota have been little studied. Here we show age-related sex differences in the adult gut microbial composition and functionality in two community-based cohorts from Northern China and the Netherlands. Consistently, women harbour a more diverse and stable microbial community across broad age ranges, whereas men exhibit a more variable gut microbiota strongly correlated with age. Reflecting the sex-biased age-gut microbiota interaction patterns, sex differences observed in younger adults are considerably reduced in the elderly population. Our findings highlight the age- and sex-biased differences in the adult gut microbiota across two ethnic population and emphasize the need for considering age and sex in studies of the human gut microbiota.

genomics↗

bioRxiv2019

Enrichment in conservative amino acid changes among fixed and standing missense variations in slow evolving proteins

Proteins were first used in the early 1960s to discover the molecular clock dating method and remain in common usage today in phylogenetic inferences based on neutral variations. To avoid substitution saturation, it is necessary to use slow evolving genes. However, it remains unclear whether fixed and standing missense changes in such genes may qualify as neutral. Here, based on the evolutionary rates as inferred from identity scores between orthologs in human and Macaca monkey, we found that the fraction of conservative amino acid mismatches between species was significantly higher in slow evolving proteins. We also examined the single nucleotide polymorphisms (SNPs) by using the 1000 genomes project data and found that missense SNPs in slow evolving proteins also had higher fraction of conservative changes, especially for common SNPs, consistent with more natural selection for SNPs, particularly rare ones, in fast evolving proteins. These results suggest that fixed and standing missense variations in slow evolving proteins are more likely to be neutral and hence better qualified for use in phylogenetic inferences.

evolutionary biology↗

bioRxiv2019

CDK1 phosphorylates ULK1-ATG13 complex to regulate mitotic autophagy and Taxol chemosensitivity

ULK1-ATG13 is the most upstream autophagy initiation complex that is phosphorylated by mTORC1 and AMPK to induce autophagy in asynchronous conditions. However, the phospho-regulation and function of ULK1-ATG13 in mitosis and cell cycle remains unknown. Here we show that ULK1-ATG13 complex is differentially regulated throughout the cell cycle. Notably, in mitosis, both ULK1 and ATG13 are highly phosphorylated by CDK1/cyclin B, the key cell cycle machinery. Combining mass spectrometry and site-directed mutagenesis, we found that CDK1-induced ULK1-ATG13 phosphorylation positively regulates mitotic autophagy and Taxol chemosensitivity, and some phosphorylation sites occur in cancer patients. Moreover, double knockout of ULK1 and ATG13 could block cell cycle progression and significantly decrease cancer cell proliferation in cell line and mouse models. Our results not only bridge the mutual regulation between the core machineries of autophagy and mitosis, illustrate the mitotic autophagy regulation mechanism, but also provide ULK1-ATG13 as potential targets for cancer therapy.

cell biology↗

bioRxiv2019

Sing cell RNA-seq reveals cellular diversity and developmental characteristics of human infant retina

Retina, located in the innermost layer of the eye of human, holds the decisive role in visual perception. Dissecting the heterogeneity of retina is essential for understanding the mechanism of vision formation and even the development of central nervous system (CNS). Here, we performed single cell RNA-seq, analyzed 57,832 cells from human infant donors, resulting in 20 distinct clusters representing major cell types in retina: rod photoreceptors, cone photoreceptors, bipolar cells, horizontal cells, amacrine cells, Muller glia cells and microglia. We next constructed extensive networks of intercellular communication and identified ligand-receptor interactions playing crucial roles in regulating neural cell development and immune homeostasis in retina. Though re-clustering, we identified known subtypes in cone PRs and additional unreported subpopulations and corresponding markers in rod PRs as well as bipolar cells. Additionally, we linked inherited retinal disease to certain cell subtypes or subpopulations through enrichment analysis. Intriguingly, we found that status and functions of photoreceptors changed drastically between early and late retina. Overall, our study offers the first retinal cell atlas in human infants, dissecting the heterogeneity of retina and identifying the key molecules in the developmental process, which provides an important resource that will pave the way for retina development mechanism research and regenerative medicine concerning retinal biology.

cell biology↗