Biology subjects

Zhao, W.

Publications and source records attributed to Zhao, W..

At least 271 records · Page 15Linked to original sources

bioRxiv2021

Regulation of a bacterial histidine kinase by a phase separating scaffolding protein

Scaffolding proteins customize the response of signaling networks to support cell development and behaviors. We investigated how the bacterial scaffolding protein PodJ regulates the histidine kinase PleC involved in the asymmetric cell division of Caulobacter crescentus. We reconstituted the PleC-PodJ signaling complex through both heterologous expression in E. coli and in vitro studies. In vitro PodJ phase separates as a biomolecular condensate that recruits and inhibits PleC kinase activity. By constructing an in vivo PleC-CcaS chimeric histidine kinase reporter assay, we have demonstrated how PodJ leverages its intrinsically disordered region (IDR) to bind and regulate PleC-CcaS signaling. Moreover, we observed that full-length PodJL regulates PleC-CcaS signaling, while a truncated PodJs could not regulate signaling activity. These results support a model where PodJ biomolecular condensate formation regulates the localization and activity of the cell fate determining kinase PleC.

synthetic biology↗

bioRxiv2021

Bioimage postprocessing based on discrete wavelet transform and Lucy-Richardson deconvolution methods

Accompanied with the increasing requirements of probing micro/nanoscopic structures of biological samples, a variety of image processing algorithms have been developed for visualization or to facilitate data analysis. However, it remains challenging to enhance both the signal-to-noise ratio and image resolution using a single algorithm. In this investigation, we propose an approach utilizing discrete wavelet transform (DWT) in combination with Lucy-Richardson (LR) deconvolution (DWDC). Our results demonstrate that the signal-to-noise ratio and resolution of live cells microtubule network are considerably improved, allowing recognition of features as small as 120 nm. Notably, the approach is independent of imaging system and shows robustness in processing fibrous structures, e.g. the cytoskeleton networks.

biophysics↗

bioRxiv2021

The Genome Sequence Archive Family: Towards Explosive Data Growth and Diverse Data Types

The Genome Sequence Archive (GSA) is a data repository for archiving raw sequence data, which provides data storing and sharing services for worldwide scientific communities. Considering explosive data growth with diverse data types, here we present the GSA family by expanding into a set of resources for raw data archive with different purposes, namely, GSA (https://ngdc.cncb.ac.cn/gsa/), GSA for Human (GSA-Human, https://ngdc.cncb.ac.cn/gsa-human/), and Open Archive for Miscellaneous Data (OMIX, https://ngdc.cncb.ac.cn/omix/). Compared with the 2017 version, GSA has been significantly updated in data model, online functionalities, and web interfaces. GSA-Human, as a new partner of GSA, is a data repository specialized in human genetics-related data with controlled access and security. OMIX, as a critical complement to the two resources mentioned above, is an open archive for miscellaneous data. Together, all these resources form a family of resources dedicated to archiving explosive data with diverse types, accept data submissions from all over the world and provide free open access to all publicly available data in support of worldwide research activities.

bioinformatics↗

bioRxiv2021

Hexapeptide induces M2 macrophage polarization via the JAK1/STAT6 pathway to promote angiogenesis in bone repair

Angiogenesis is essential for successful bone defect repair. In normal tissue repair, the physiological inflammatory response is the main regulator of angiogenesis through the activity of macrophages and the cytokines secreted by them. In particular, M2 macrophages which secrete high levels of PDGF-BB are typically considered to promote angiogenesis. A hexapeptide [WKYMVm, (Trp-Lys-Tyr-Met-Val-D-Met-NH2)] has been reported to modulate inflammatory activities. However, the underlying mechanisms by which WKYMVm regulates macrophages remain unclear. In this study, the possible involvement by which WKYMVm induces the polarization of macrophages and affects their behaviors was evaluated. In vitro results showed that macrophages were induced to an M2 rather than M1 phenotype and the M2 phenotype was enhanced by WKYMVm through activation of the JAK1/STAT6 signaling pathway. It was also found that WKYMVm played an important role in the PDGF-BB production increase and proangiogenic abilities in M2 macrophages. Consistent with the results in vitro, the elevated M2/M0 ratio induced by WKYMVm enhanced the formation of new blood vessels in a femoral defect mouse model. In summary, these findings suggest that WKYMVm could be a promising alternative strategy for angiogenesis in bone repair by inducing M2 macrophage polarization.

biochemistry↗

bioRxiv2021

Spatial transcriptomics using combinatorial fluorescence spectral and lifetime encoding, imaging and analysis

Multiplexed mRNA profiling in the spatial context provides important new information enabling basic research and clinical applications. Unfortunately, most existing spatial transcriptomics methods are limited due to either low multiplexing or assay complexity. Here, we introduce a new spatialomics technology, termed Multi Omic Single-scan Assay with Integrated Combinatorial Analysis (MOSAICA), that integrates in situ labeling of mRNA and protein markers in cells or tissues with combinatorial fluorescence spectral and lifetime encoded probes, spectral and time-resolved fluorescence imaging, and machine learning-based target decoding. This technology is the first application combining the biophotonic techniques, Spectral and Fluorescence Lifetime Imaging and Microscopy (FLIM), to the field of spatial transcriptomics. By integrating the time dimension with conventional spectrum-based measurements, MOSAICA enables direct and highly-multiplexed in situ spatial biomarker profiling in a single round of staining and imaging while providing error correction removal of background autofluorescence. We demonstrate mRNA encoding using combinatorial spectral and lifetime labeling and target decoding and quantification using a phasor-based image segmentation and machine learning clustering technique. We then showcase MOSAICAs multiplexing scalability in detecting 10-plex targets in fixed colorectal cancer cells using combinatorial labeling of only five fluorophores with facile error-correction and removal of autofluorescent moieties. MOSAICAs analysis is strongly correlated with sequencing data (Pearsons r = 0.9) and was further benchmarked using RNAscopeand LGC Stellaris. We further apply MOSAICA for multiplexed analysis of clinical melanoma Formalin-Fixed Paraffin-Embedded (FFPE) tissues that have a high degree of tissue scattering and intrinsic autofluorescence, demonstrating the robustness of the approach. We then demonstrate simultaneous co-detection of protein and mRNA in colorectal cancer cells. MOSAICA represents a simple, versatile, and scalable tool for targeted spatial transcriptomics analysis that can find broad utility in constructing human cell atlases, elucidating biological and disease processes in the spatial context, and serving as companion diagnostics for stratified patient care.

bioengineering↗

bioRxiv2021

The Prediction of Active Ingredients and Polypharmacological Mechanisms of Orthosiphon stamineus Benth. (Lamiaceae) of Dai medicine based on Network Pharmacology

BackgroundOrthosiphon stamineus Benth. was a dietary supplement or a traditional folk herb with widespread clinical application, but it was lack of comprehensive understanding of its active ingredients and polypharmacological mechanisms. The aim of this work was to systematically study its natural compounds and the molecular mechanisms of the O. stamineus via network pharmacology. MethodsCompounds from O. stamineus were collected by literature retrieval and evaluated by SwissADME with the physicochemical property (ADMET model) and the likelihood for a natural medicine. The connection of active ingredients and target genes was built and confirmed by Cytoscape and AutoDock vina. Then, Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were separately performed to obtain a more in-depth understanding of O. stamineus. Finally, the relationship among active ingredients, targets, and diseases was built to clarify the polypharmacological mechanisms and found relevant active substances for further drug discovery. ResultsA total of 159 compounds of O. stamineus were collected, and 22 potentially active ingredients were screened out. The Ingredient-Target Interaction Network was built with 12 flavonoids, 3 diterpenes, 3 phenols, and 4 volatile oils, and 65 targets. The Docking analysis indicated that the ingredient-target interaction network was reliable; most ligand-receptor had a strong binding affinity (lowest binding energy: -6.9 kcal/mol). After pathway analysis, 185 significant biological processes and 36 signal pathways were found, and the ingredient-target-disease network of O. stamineus was constructed for polypharmacological mechanisms. ConclusionOur study clarified the polypharmacological mechanisms via the relationship among active ingredients, targets, and diseases and provided better guidance for subsequent experiments and potential active ingredients for drug discovery or health promotion.

systems biology↗

bioRxiv2021

CCPE: Cell Cycle Pseudotime Estimation for Single Cell RNA-seq Data

The rapid development of single-cell RNA-sequencing (scRNA-seq) technologies makes it possible to characterize cellular heterogeneity by detecting and quantifying transcriptional changes at the single-cell level. Pseudotime analysis enables to characterize the continuous progression of various biological processes, such as cell cycle. Cell cycle plays an important regulatory role in cell fate decisions and differentiation and is also often regarded as a confounder in scRNA-seq data analysis when analyzing the role of other factors on transcriptional regulation. Therefore, accurate prediction of cell cycle pseudotime and identify cell stages are important steps for characterizing the development-related biological processes, identifying important regulatory molecules and promoting the analysis of transcriptional heterogeneity. Here, we develop CCPE, a novel cell cycle pseudotime estimation method to characterize cell cycle timing and determine cell cycle phases from single-cell RNA-seq data. CCPE uses a discriminative helix to characterize the circular process and estimates pseudotime in the cell cycle. We evaluated the model performance based on a variety of simulated and real scRNA-seq datasets. Our results indicate that CCPE is an effective method for cell cycle estimation and competitive in various downstream analyses compared with other existing methods. CCPE successfully identified cell cycle marker genes and is robust to dropout events in scRNA-seq data. CCPE also has excellent performance on small datasets with fewer genes or cells. Accurate prediction of the cell cycle in CCPE effectively contributes to cell cycle effect removal across cell types or conditions.

bioinformatics↗

bioRxiv2021

PHARP: A pig haplotype reference panel for genotype imputation

Pigs not only function as a major meat source worldwide but also are commonly used as an animal model for studying human complex traits. A large haplotype reference panel has been used to facilitate efficient phasing and imputation of relatively sparse genome-wide microarray chips and low-coverage sequencing data. Using the imputed genotypes in the downstream analysis, such as GWASs, TWASs, eQTL mapping and genomic prediction (GS), is beneficial for obtaining novel findings. However, currently, there is still a lack of publicly available and high-quality pig reference panels with large sample sizes and high diversity, which greatly limits the application of genotype imputation in pigs. In response, we built the pig Haplotype Reference Panel (PHARP) database. PHARP provides a reference panel of 2,012 pig haplotypes at 34 million SNPs constructed using whole-genome sequence data from more than 49 studies of 71 pig breeds. It also provides Web-based analytical tools that allow researchers to carry out phasing and imputation consistently and efficiently. PHARP is freely accessible at http://alphaindex.zju.edu.cn/PHARP/index.php. We demonstrate its applicability for pig commercial 50K SNP arrays, by accurately imputing 2.6 billion genotypes at a concordance rate value of 0.971 in 81 Large White pigs (~ 17x sequencing coverage). We also applied our reference panel to impute the low-density SNP chip into the high-density data for three GWASs and found novel significantly associated SNPs that might be casual variants.

genomics↗

bioRxiv2021

Transient nuclear deformation primes epigenetic state and promotes cell reprogramming

Cell reprogramming has wide applications in tissue regeneration, disease modeling and personalized medicine, but low reprogramming efficiency remains a challenge. In addition to biochemical cues, biophysical factors can modulate the epigenetic state and a variety of cell functions. However, how biophysical factors help overcome the epigenetic barrier for cell reprogramming are not well understood. Here we utilized microfluidic channels to induce a transient deformation of the cell nucleus, which caused the disassembly of the nuclear lamina and a downregulation of DNA methylation and histone (H3K9) for 12-24 hours. These global decreases of heterochromatin marks at the early stage of cell reprogramming strikingly enhanced the conversion of fibroblasts into neurons and induced pluripotent stem cells. Consistently, inhibition of DNA methylation and H3K9 methylation partially mimicked the effects of mechanical squeezing on iN reprogramming efficiency. Knocking down lamin A had similar effects to squeezing on enhancing the reprogramming efficiency. Based on these findings, we developed a scalable microfluidic system that enabled a continuous cell processing to effectively prime the epigenetic state for cell reprogramming, demonstrating the potential of mechano-biotechnology for cell engineering.

biophysics↗

bioRxiv2021

The mirror neuron system compensates for amygdala dysfunction-associated social deficits in individuals with higher autistic traits

The amygdala is a core node in the social brain which exhibits structural and functional abnormalities in Autism spectrum disorder and there is evidence that the mirror neuron system (MNS) can functionally compensate for impaired emotion processing following amygdala lesions. In the current study, we employed an fMRI paradigm in 241 subjects investigating MNS and amygdala responses to observation, imagination and imitation of dynamic facial expressions and whether these differed in individuals with higher as opposed to lower autistic traits. Results indicated that individuals with higher compared to lower autistic traits showed reduced left amygdala responses to imitation and enhanced responses in the left superior temporal sulcus (STS) of the MNS to observation, imagination and imitation. Additionally, functional connectivity between the left amygdala and the left STS as well as some other MNS regions was increased in individuals with higher autistic traits, especially during imitation of fearful expressions. The amygdala-MNS connectivity significantly moderated autistic group differences on recognition memory for fearful faces and real-life social network indices, indicating that increased amygdala-MNS connectivity could diminish the social behavioral differences between higher and lower autistic trait groups. Overall, findings demonstrate decreased imitation-related amygdala activity in individuals with higher autistic traits in the context of increased cortical MNS activity and amygdala-MNS connectivity which may functionally compensate for amygdala dysfunction and social deficits. Training targeting the MNS may capitalize on this compensatory mechanism for therapeutic benefits in Autism spectrum disorder.

neuroscience↗

bioRxiv2021

Generalization of Cortical Multivariate Genome-Wide Associations Within and Across Samples

Genome-Wide Association studies have typically been limited to single phenotypes, given that high dimensional phenotypes incur a large multiple comparisons burden: ~1 million tests across the genome times the number of phenotypes. Recent work demonstrates that a Multivariate Omnibus Statistic Test (MOSTest) is well powered to discover genomic effects distributed across multiple phenotypes. Applied to cortical brain MRI morphology measures, MOSTest has resulted in a drastic improvement in power to discover loci when compared to established approaches (min-P). One question that arises is how well these discovered loci replicate in independent data. Here we perform 10 times cross validation within 35,644 individuals from UK Biobank for imaging measures of cortical area, thickness and sulcal depth (>1,000 dimensionality for each). By deploying a replication method that aggregates discovered effects distributed across multiple phenotypes, termed PolyVertex Score (PVS), we demonstrate a higher replication yield and comparable replication rate of discovered loci for MOSTest (# replicated loci: 348-845, replication rate: 94-95%) in independent data when compared with the established min-P approach (# replicated loci: 31-68, replication rate: 65-80%). An out-of-sample replication of discovered loci was conducted with a sample of 8,336 individuals from the Adolescent Brain Cognitive Development(R) (ABCD) study, who are on average 50 years younger than UK Biobank individuals. We observe a higher replication yield and comparable replication rate of MOSTest compared to min-P. This finding underscores the importance of using well-powered multivariate techniques for both discovery and replication of high dimensional phenotypes in Genome-Wide Association studies.

genomics↗

bioRxiv2021

Anxiolytic effects of chronic intranasal oxytocin on neural responses to threat are dose-frequency dependent

Anxiety disorders are prevalent psychiatric conditions characterized by exaggerated anxious arousal and threat reactivity. Animal and human studies suggest an anxiolytic potential of the neuropeptide oxytocin (OT), yet, while a clinical application will require chronic administration protocols previous studies in humans have exclusively focused on single-dose (acute) intranasal OT effects. We aimed at determining whether the anxiolytic effects of OT are maintained with repeated (chronic) administration or are influenced by dose frequency and trait anxiety. A double-blind randomized, placebo-controlled pharmaco-fMRI trial (n=147) determined acute (single-dose) as well as chronic effects of two different dose frequencies of OT (OT administered daily for 5 days or every other day) on emotional reactivity in healthy subjects with high versus low trait anxiety. OT produced valence, dose frequency and trait anxiety specific effects, such that the low-frequency (intermittand) chronic dosage specifically attenuated neural reactivity in amygdala-insula-prefrontal regions in high anxious subjects in response to threatening but not positive stimuli. The present trial provides evidence that low dose frequency chronic oxytocin nasal spray has the potential to alleviate exaggerated neural threat reactivity in subjects with elevated anxiety levels underscoring a treatment potential for anxiety disorders.

neuroscience↗

bioRxiv2021

Extracellular vesicles Transfer Polarized Mitochondria and Increase Cellular Energetics in Ischemic Endothelial Cells

We have demonstrated, for the first time that microvesicles, a sub-type of extracellular vesicles (EVs) derived from hCMEC/D3: a human brain endothelial cell (BEC) line transfer polarized mitochondria to recipient BECs in culture and to neurons in mice acute brain cortical and hippocampal slices. This mitochondrial transfer increased ATP levels by 100 to 200-fold (relative to untreated cells) in the recipient BECs exposed to oxygen-glucose deprivation, an in vitro model of cerebral ischemia. We have also demonstrated that transfer of microvesicles, the larger EV fraction, but not exosomes resulted in increased mitochondrial function in hypoxic endothelial cultures. Gene ontology and pathway enrichment analysis of EVs revealed a very high association to glycolysis-related processes. In comparison to heterotypic macrophage- derived EVs, BEC-derived EVs demonstrated a greater selectivity to transfer mitochondria and increase endothelial cell survival under ischemic conditions. HighlightsO_LIMicrovesicles transfer mitochondria to endothelial cells and brain slice neurons C_LIO_LIMitochondrial transfer increased ATP in ischemic brain endothelial cells (BECs) C_LIO_LITransfer of microvesicles increased mitochondrial function in hypoxic BECs C_LIO_LITransfer of exosomes did not affect mitochondrial function in hypoxic BECs C_LIO_LIHomotypic BEC-derived EVs result in greater ATP levels in the recipient BECs C_LI

bioengineering↗

bioRxiv2021

Genome-wide survey of the F-box/Kelch (FBK) members and molecular identification of a novel FBK gene TaAFR in wheat

F-box proteins play critical roles in plant responses to biotic/abiotic stresses. In the present study, a total of 68 wheat F-box/Kelch (TaFBK) gene sequences encoding for 74 proteins were obtained in a genome-wide survey against EnsemblPlants. The 74 TaFBK proteins were divided into 5 categories based on their domain structures. The FBK proteins from wheat, Arabidopsis, and three other cereal species were grouped into 7 clades, and the number of Kelch domains present was their key clustering criterion. Sixty-eight TaFBK genes were unevenly distributed on 21 chromosomes. Most of TaFBKs were predicted to localize in the nucleus and cytoplasm. In silico analysis of a digital PCR revealed that TaFBKs were expressed at multiple developmental stages and tissues, and in response to drought and/or heat stresses. The TaFBK19 gene, a homologous to the Attenuated Far-Red Response (AFR) genes in other plant species, and hence named TaAFR, was selected for further analysis. The gene was isolated from the wheat line TcLr15 and its expression evaluated by quantitative real-time PCR. TaAFR transcripts were primarily detected in wheat leaves, and its expression was found to be regulated by various abiotic and biotic stresses as well as plant signaling hormones. Of particular interest, TaAFR expression was differentially regulated in the compatible vs incompatible wheat leaf rust reaction. Subcellular localization studies showed that TaAFR accumulates in the nucleus and cytoplasm. Three TaAFR-interacting proteins were identified experimentally: Skp1/ASK1-like protein (Skp1), ADP-ribosylation factor 2-like isoform X1 (ARL2) and phenylalanine ammonia-lyase (PAL). Further analysis revealed that the Skp1 protein interacted specifically with the F-box domain of TaAFR, while ARL2 and PAL were recognized by the Kelch domain. The data presented herein provides a solid foundation from which the function and metabolic network of TaAFR and other wheat FBKs can be further explored.

molecular biology↗

bioRxiv2021

A Chemical-Enhanced System for CRISPR-Based Nucleic Acid Detection

The CRISPR-based nucleic acid detection systems such as SHERLOCK, DETECTR and HOLMES have shown great potential for point-of-care testing of viral pathogens, especially in the context of COVID-19 pandemic. Here we optimize several key parameters of reaction chemistry and develop a Chemical Enhanced CRISPR Detection system for nucleic acid (termed CECRID). For the Cas12a/Cas13a-based signal detection phase, we determine buffer conditions and substrate range for optimal detection performance. By comparing several chemical additives, we find that addition of L-proline can secure or enhance Cas12a/Cas13a detection capability. For isothermal amplification phase with typical LAMP and RPA methods, inclusion of L-proline can also enhance specific target amplification as determined by CRISPR detection. Using SARS-CoV-2 pseudovirus, we demonstrate CECRID has enhanced detection sensitivity over chemical additive-null method with either fluorescence or lateral flow strip readout. Thus, CECRID provides an improved detection power and system robustness towards practical application of CRISPR-based diagnostics.

molecular biology↗

bioRxiv2021

Defining proximity proteomics of post-translationally modified proteins by antibody-mediated protein A-APEX2 labeling

Proximity labeling catalyzed by promiscuous enzymes, such as APEX2, has emerged as a powerful approach to characterize multiprotein complexes and protein-protein interactions. However, current methods depend on the expression of exogenous fusion proteins and cannot be applied to post-translational modifications. To address this limitation, we developed a new method to label proximal proteins of interest by antibody-mediated protein A-APEX2 labeling (AMAPEX). In this method, a modified protein is bound in situ by a specific antibody, which then tethers a protein A-APEX2 (pA-APEX2) fusion protein. Activation of APEX2 labels the nearby proteins with biotin; these proteins are then purified using streptavidin beads and are identified by mass spectrometry. We demonstrate the utility of this approach by profiling the binding proteins of histone modifications including H3K27me3, H3K9me3, H3K4me3, H4K5ac and H4K12ac, and we verified the genome-wide colocalization of these identified proteins with bait proteins by published ChIP-seq analysis. Overall, AMAPEX is an efficient tool to identify proteins that interact with modified proteins.

molecular biology↗

bioRxiv2021

AIM-CICs: automatic identification method for Cell-in-cell structures based on convolutional neural network

Whereas biochemical markers are available for most types of cell death, current studies on non-autonomous cell death by entosis relays strictly on the identification of cell-in-cell structure (CICs), a unique morphological readout that can only be quantified manually at present. Moreover, the manual CICs quantification is generally over-simplified as CICs counts, which represents a major hurdle against profound mechanistic investigations. In this study, we take advantage of artificial intelligence (AI) technology to develop an automatic identification method for CICs (AIM-CICs), which performs comprehensive CICs analysis in an automated and efficient way. The AIM-CICs, developed on the algorithm of convolutional neural network (CNN), can not only differentiate between CICs and non-CICs (AUC > 0.99), but also accurately categorize CICs into five subclasses based on CICs stages and cell number involved (AUC > 0.97 for all subclasses). The application of AIM-CICs would systemically fuel researches on CICs-mediated cell death such as high-throughput screening.

cell biology↗

bioRxiv2021

Modulating the voltage sensor of a cardiac potassium channel shows antiarrhythmic effects

Cardiac arrhythmias are the most common cause of sudden cardiac death worldwide. Lengthening the ventricular action potential duration (APD) either congenitally or via pathologic or pharmacologic means, predisposes to a life-threatening ventricular arrhythmia, Torsade de Pointes. IKs, a slowly activating K+ current plays a role in action potential repolarization. In this study, we screened a chemical library in silico by docking compounds to the voltage sensing domain (VSD) of the IKs channel. Here we show that C28 specifically shifted IKs VSD activation in ventricle to more negative voltages and reversed drug-induced lengthening of APD. At the same dosage, C28 did not cause significant changes of the normal APD in either ventricle or atrium. This study provides evidence in support of a computational prediction of IKs VSD activation as a potential therapeutic approach for all forms of APD prolongation. This outcome could expand the therapeutic efficacy of a myriad of currently approved drugs that may trigger arrhythmias. Significance statementC28, identified by in silico screening, specifically facilitated voltage dependent activation of a cardiac potassium ion channel, IKs. C28 reversed drug-induced prolongation of action potentials, but minimally affected the normal action potential at the same dosage. This outcome supports a computational prediction of modulating IKs activation as a potential therapy for all forms of action potential prolongation, and could expand therapeutic efficacy of many currently approved drugs that may trigger arrhythmias.

biophysics↗