Biology subjects

Zhang, D.

Publications and source records attributed to Zhang, D..

At least 379 records · Page 21Linked to original sources

bioRxiv2021

Spatial Epigenome Sequencing at Tissue Scale and Cellular Level

Spatial biology is emerging as a new frontier of biomedical research in development and disease, but currently limited to transcriptome and a panel of proteins. Here we present spatial epigenome profiling for three histone modifications (H3K27me3, H3K4me3, H3K27ac) via next-generation sequencing by combining in-tissue CUT&Tag chemistry and microfluidic deterministic barcoding. Spatial chromatin states in mouse embryos or olfactory bulbs revealed tissue type-specific epigenetic regulations, in concordance with ENCODE reference data, but providing spatially resolved genome-wide profiles at tissue scale. Using fluorescence imaging to identify the tissue pixels (20m) each containing one nucleus allowed us to extract single-cell epigenomes in situ. Spatial chromatin state profiling in tissue may enable unprecedented opportunities to study epigenetic regulation, cell function and fate decision in normal physiology and pathogenesis.

genomics↗

bioRxiv2021

Leveraging omic features with F3UTER enables identification of unannotated 3'UTRs for synaptic genes

There is growing evidence for the importance of 3 untranslated region (3UTR) dependent regulatory processes. However, our current human 3UTR catalogue is incomplete. Here, we developed a machine learning-based framework, leveraging both genomic and tissue-specific transcriptomic features to predict previously unannotated 3UTRs. We identify unannotated 3UTRs associated with 1,513 genes across 39 human tissues, with the greatest abundance found in brain. These unannotated 3UTRs were significantly enriched for RNA binding protein (RBP) motifs and exhibited high human lineage-specificity. We found that brain-specific unannotated 3UTRs were enriched for the binding motifs of important neuronal RBPs such as TARDBP and RBFOX1, and their associated genes were involved in synaptic function and brain- related disorders. Our data is shared through an online resource F3UTER (https://astx.shinyapps.io/F3UTER/). Overall, our data improves 3UTR annotation and provides novel insights into the mRNA-RBP interactome in the human brain, with implications for our understanding of neurological and neurodevelopmental diseases.

bioinformatics↗

bioRxiv2021

Instructor-learner neural synchronization during elaborated feedback predicts learning transfer

The provision of feedback with complex information beyond the correct answer, i.e., elaborated feedback, can powerfully shape learning outcomes such as transfer, i.e., the ability to extend what has been learned in one context to new contexts. However, an understanding of neurocognitive processes of elaborated feedback during instructor-learner interactions remains elusive. Here, a two-person interactive design is used during simultaneous recording of functional near-infrared spectroscopy (fNIRS) signals from adult instructor-learner dyads. Instructors either provided elaborated feedback (i.e., correct answer and an example) or simple feedback (i.e., correct answer only) to learners during a concept learning task. Our results showed that elaborated feedback produced comparable levels of retention to simple feedback, however, transfer was significantly enhanced by elaboration. We also noted significant instructor-learner neural synchronization in frontoparietal regions during the provision of elaborated feedback, especially when examples were provided. Further, interpersonal neural synchronization in the parietal cortex successfully predicted transfer of knowledge to novel contexts. This prediction was retained for both learner-delayed and learner-preceding neural synchronization. These findings point toward transfer effects of elaborated feedback provided in a social context can be predictable through interpersonal neural synchronization, which may hold important implications for real-world learning and pedagogical efficacy. Educational Impact and Implications StatementFeedback provides learners with crucial information regarding the gap between what has currently been achieved and what remains to be achieved, and thus plays a critical role in any learning process. In real-world settings, feedback is typically provided and received through social interaction, and high-quality "elaborated feedback" contains complex information that goes beyond the correct answer. This study aims to elucidate the neurocognitive processes underpinning elaborated feedback during instructor-learner interactions. We detected significant instructor-learner neural synchronization in mutual frontoparietal brain regions during elaborated feedback, particularly during the provision of specific elaborated information (i.e., concrete examples). Moreover, this synchronization (including learner-delayed and learner-preceded synchronization) in the parietal region predicted whether the learners transferred learning to novel examples of learned psychology concepts. This study advances current understanding on the neural mechanisms for elaborated feedback and the role of social interaction in feedback effects. These results may have important implications for successful real-world learning and communication, and related pedagogical applications in educational settings.

scientific communication and education↗

bioRxiv2021

miRNA-dependent poly(A) length control in uncoupling transcription and translation of haploid male germ cells

As one of the post-transcriptional regulatory mechanisms, transcription and translations uncoupling plays an essential role in development and adulthood physiology. However, it remains elusive how thousands of mRNAs get translationally silenced while stability is maintained for up to hours or even days before translation. In addition to oocytes and neurons, developing spermatids have significant uncoupling of transcription and translation for delayed translation. Therefore, spermiogenesis represents an excellent in vivo model for investigating the mechanism underlying uncoupled transcription and translation. Through full-length poly(A) deep sequencing, we discovered dynamic changes in poly(A) length through deadenylation and re-polyadenylation. Deadenylation appeared to be mediated by microRNAs (miRNAs), and transcripts with shorter poly(A) tails tend to be sequestered into ribonucleoproteins (RNPs) for translational repression and stabilization. In contrast, re-polyadenylation allows for translocation of the translationally repressed transcripts from RNPs to polysomes for translation. Overall, our data suggest that miRNA-dependent poly(A) length control represents a novel mechanism underlying uncoupled translation and transcription in haploid male germ cells.

molecular biology↗

bioRxiv2021

Familial prion disease-related mutation E196K displays a novel amyloid fibril structure revealed by cryo-EM

Prion diseases are caused by the conformational conversion of prion protein (PrP) from its cellular form (PrPC) into a protease-resistant, aggregated form (PrPSc). 42 different familial mutations were identified in human PrP, which lead to genetic prion diseases with distinct clinical syndromes. Here we report cryo-EM structure of an amyloid fibril formed by full-length human PrP with E196K mutation, a familial Creutzfeldt-Jakob disease-related mutation. This mutation disrupts key interactions in wild-type PrP fibril and results in a rearrangement of the overall structure, forming an amyloid fibril with a conformation distinct from wild-type PrP fibril. The E196K fibril consists of two protofibrils intertwined into a left-handed helix. Each subunit forms five {beta}-strands stabilized by a disulfide bond and an unusual hydrophilic cavity. Two pairs of amino acids (Lys194 and Glu207; Lys196 and Glu200) from opposing subunits form four salt bridges to stabilize the zigzag interface of the two protofibrils. Furthermore, the E196K fibril exhibits a significantly lower conformational stability and protease resistance activity than the wild-type fibril. Our results provide direct structural evidences of the diverse mammalian prion strains and fibril polymorphism of PrP, and highlight the importance of familial mutations in determining the different prion strains.

biochemistry↗

bioRxiv2021

Aggregation-induced emission dots assisted non-invasive fluorescence hysterography in near-infrared IIb window

Uterine diseases seriously threaten the physical and mental health of women. The main principle, when clinicians adopt examinations, is to achieve efficient diagnosis without negative effect on the physical function including fertility. Hysterography in near-infrared (NIR) IIb window (1500-1700 nm) presents perceptibly enhanced signal to background ratio (SBR) and higher penetration capability compared with those beyond 1000 nm and 1300 nm, but lays down high requirements for the biosafety of fluorophores at the same time. Assisted by the biologically excretable aggregation-induced emission (AIE) dots, non-invasive NIR-IIb fluorescence hysterography visualized typical Y-shaped uteruses, real-time uterine peristalsis or the uterine lesions (mimetic disease statuses in clinic) in mouse models. Significantly, after intrauterine perfusion, the reproductive capacity was unimpaired via fertility assessment and histological analysis. This work could inspire some new ideas for non-invasive clinical diagnosis of uterine diseases and effectively promote the clinical translation of AIE dots.

bioengineering↗

bioRxiv2021

A potent bispecific nanobody protects hACE2 mice against SARS-CoV-2 infection via intranasal administration

The dramatically expanding COVID-19 needs multiple effective countermeasures. Neutralizing antibodies are a potential therapeutic strategy for treating COVID-19. A number of neutralizing nanobodies (Nbs) were reported for their in vitro activities. However, in vivo protection of these nanobodies was not reported in animal models. In the current report, we characterized several RBD-specific Nbs isolated from a screen of an Nb library derived from an alpaca immunized with SARS-CoV-2 spike glycoprotein (S); among them, three Nbs exhibited picomolar potency against SARS-CoV-2 live virus, pseudotyped viruses, and 15 circulating SARS-CoV-2 variants. To improve the efficacy, various configurations of Nbs were engineered. Nb15-NbH-Nb15, a novel trimer constituted of three Nbs, was constructed to be bispecific for human serum albumin (HSA) and RBD of SARS-CoV-2. Nb15-NbH-Nb15 exhibited sub-ng/ml neutralization potency against the wild-type and currently circulating variants of SARS-CoV-2 with a long half-life in vivo. In addition, we showed that intranasal administration of Nb15-NbH-Nb15 provided 100% protection for both prophylactic and therapeutic purposes against SARS-CoV-2 infection in transgenic hACE2 mice. Nb15-NbH-Nb15 is a potential candidate for both prevention and treatment of SARS-CoV-2 through respiratory administration. One sentence summaryNb15-NbH-Nb15, with a novel heterotrimeric bispecific configuration, exhibited potent and broad neutralization potency against SARS-CoV-2 in vitro and provided in vivo protection against SARS-CoV-2 infection in hACE2 transgenic mice via intranasal delivery. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=156 SRC="FIGDIR/small/429275v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@a30bc8org.highwire.dtl.DTLVardef@5a872eorg.highwire.dtl.DTLVardef@1610f74org.highwire.dtl.DTLVardef@13d9bd8_HPS_FORMAT_FIGEXP M_FIG Graphical abstract: C_FIG HighlightsO_LIWe described a novel heterotrimeric configuration of Nb-NbH-Nb (Nb15-NbH-Nb15) that exhibited improved viral inhibition and stability. C_LIO_LINb15-NbH-Nb15 provides ultrahigh neutralization potency against SARS-CoV-2 wild type and 18 mutant variants, including the current circulating variants of D614G and N501Y predominantly in the UK and South Africa. C_LIO_LIIt is the first to demonstrate the Nbs efficacy in preventing and treating SARS-CoV-2 infection in hACE2 transgenic mice via intranasal delivery. C_LI

microbiology↗

bioRxiv2021

Human brain mitochondrial-nuclear cross-talk is cell-type specific and is perturbed by neurodegeneration

Mitochondrial dysfunction contributes to the pathogenesis of many neurodegenerative diseases as mitochondria are essential to neuronal function. The mitochondrial genome encodes a small number of core respiratory chain proteins, whereas the vast majority of mitochondrial proteins are encoded by the nuclear genome. Here we focus on establishing a profile of nuclear-mitochondrial transcriptional relationships in healthy human central nervous system tissue data, before examining perturbations of these processes in Alzheimer&#8217s disease using transcriptomic data originating from affected human brain tissue. Through cross-central nervous system analysis of mitochondrial-nuclear gene pair relationships, we find that the cell type composition underlies regional variation, and variation is driven at the subcellular level by heterogeneity of nuclear-mitochondrial coordination in post-synaptic regions. We show that nuclear genes causally implicated in sporadic Parkinson&#8217s disease and Alzheimer&#8217s disease show much stronger relationships with the mitochondrial genome than expected by chance, and that nuclear-mitochondrial relationships are significantly perturbed in Alzheimer&#8217s disease cases, particularly amongst genes involved in synaptic and lysosomal pathways. Finally, we present MitoNuclearCOEXPlorer, a web tool designed to allow users to interrogate and visualise key mitochondrial-nuclear relationships in multi-dimensional brain data. We conclude that mitochondrial-nuclear relationships differ significantly across regions of the healthy brain, which appears to be driven by the functional specialisation of different cell types. We also find that mitochondrial-nuclear co-expression in critical pathways is disrupted in Alzheimer&#8217s disease, potentially implicating the regulation of energy balance and removal of dysfunctional mitochondria in the etiology or progression of the disease and making the case for the relevance of bi-genomic co-ordination in the pathogenesis of neurodegenerative diseases.

genomics↗

bioRxiv2021

Compensating complete loss of signal recognition particle during co-translational protein targeting by the translation speed and accuracy

Signal recognition particle (SRP) is critical for delivering co-translational proteins to the bacterial inner membrane. Previously, we identified SRP suppressors in Escherichia coli that inhibit translation initiation and elongation, which provided insights into the mechanism of bypassing the requirement of SRP. Suppressor mutations tended to be located in regions that govern protein translation under evolutionary pressure. To test this hypothesis, we re-executed the suppressor screening of SRP. Here we isolated a novel SRP suppressor mutation located in the Shine-Dalgarno sequence of the S10 operon, which partially offset the targeting defects of SRP-dependent proteins. We found that the suppressor mutation decreased the protein translation rate, which extended the time window of protein targeting. This increased the possibility of the correct localization of inner membrane proteins. Furthermore, the fidelity of translation was decreased in suppressor cells, suggesting that the quality control of translation was inactivated to provide an advantage in tolerating toxicity caused by the loss of SRP. Our results demonstrated that the inefficient protein targeting due to SRP deletion can be rescued through modulating translational speed and accuracy.

microbiology↗

bioRxiv2020

An Ultrasensitive Biosensor for Quantifying the Interaction of SARS-CoV-2 and Its Receptor ACE2 in Cells and in vitro

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is currently spreading and mutating with increasing speed worldwide. Therefore, there is an urgent need for a simple, sensitive, and high-throughput (HTP) assay to quantify virus-host interaction in order to quickly evaluate infectious ability of mutant virus and develop or validate virus-inhibiting drugs. Here we have developed an ultrasensitive bioluminescent biosensor to evaluate virus-cell interaction by quantifying the interaction between SARS-CoV-2 receptor binding domain (RBD) and its cellular receptor angiotensin-converting enzyme 2 (ACE2) both in living cells and in vitro. We have successfully used this novel biosensor to analyze SARS-CoV-2 RBD mutants, and evaluated candidate small molecules (SMs), antibodies, and peptides that may block RBD:ACE2 interaction. This simple, rapid and HTP biosensor tool will significantly expedite detection of viral mutants and anti-COVID-19 drug discovery processes.

biochemistry↗

bioRxiv2020

Megadepth: efficient coverage quantification for BigWigs and BAMs

MotivationA common way to summarize sequencing datasets is to quantify data lying within genes or other genomic intervals. This can be slow and can require different tools for different input file types. ResultsMegadepth is a fast tool for quantifying alignments and coverage for BigWig and BAM/CRAM input files, using substantially less memory than the next-fastest competitor. Megadepth can summarize coverage within all disjoint intervals of the Gencode V35 gene annotation for more than 19,000 GTExV8 BigWig files in approximately one hour using 32 threads. Megadepth is available both as a command-line tool and as an R/Bioconductor package providing much faster quantification compared to the rtracklayer package. Availabilityhttps://github.com/ChristopherWilks/megadepth, https://bioconductor.org/packages/megadepth. Contactchris.wilks@jhu.edu

bioinformatics↗

bioRxiv2020

A phenotypic screen identifies a compound series that induces differentiation of acute myeloid leukemia cells in vitro and shows anti-tumour effects in vivo

Induction of differentiation is a promising therapeutic strategy against acute myeloid leukemia. However, current differentiation therapies are effective only to specific patient populations. To identify novel differentiation agents with wider efficacy, we developed a phenotypic high-throughput screen with a range of genetically diverse cell lines. From the resulting hits, one chemical scaffold was optimised in terms of activity and physicochemical properties to yield OXS007417, which was also able to decrease tumour volume in a murine in vivo xenograft model.

cancer biology↗

bioRxiv2020

Informing NMR experiments with molecular dynamics simulations to characterize the dominant activated state of the KcsA ion channel

As the first potassium channel with a X-ray structure determined, and given its homology to eukaryotic channels, the pH-gated prokaryotic channel KcsA has been extensively studied. Nevertheless, questions related in particular to the allosteric coupling between its gates remain open. The many currently available X-ray crystallography structures appear to correspond to various stages of activation and inactivation, offering insights into the molecular basis of these mechanisms. Since these studies have required mutations, complexation with antibodies, and substitution of detergents in place of lipids, examining the channel under more native conditions is desirable. Solid-state NMR (SSNMR) can be used to study the wild-type protein under activating conditions (low pH), at room temperature, and in bacteriomimetic liposomes. In this work, we sought to structurally assign the activated state present in SSNMR experiments. We used a combination of molecular dynamics (MD) simulations, chemical shift prediction algorithms, and Bayesian inference techniques to determine which of the most plausible X-ray structures resolved to date best represents the activated state captured in SSNMR. We first identified specific nuclei with simulated NMR chemical shifts that differed significantly when comparing partially open vs. fully open ensembles from MD simulations. The simulated NMR chemical shifts for those specific nuclei were then compared to experimental ones, revealing that the simulation of the partially open state was in good agreement with the SSNMR data. Nuclei that discriminate effectively between partially and fully open states belong to residues spread over the sequence and provide a molecular level description of the conformational change.

biophysics↗

bioRxiv2020

Effects of Ca2+ on encystment and growth in Scrippsiella trochoidea

Cysts serve as a seed source for the initiation and recurrence of a harmful algal bloom (HAB) caused by dinoflagellates. And the influence of calcium on cyst formation has been relatively understudied. In the present study, we investigated the effects of calcium (Ca2+) on the growth and encystment of Scrippsiella trochoidea. We incubated S. trochoidea in modified f/2 media in flasks which were divided into five groups and treated with different Ca2+ concentrations (0, 0.2, 0.4, 0.6, and 0.8 g{middle dot}L-1). We revealed that cell density increased with increasing Ca2+ concentrations; however, cell density was reduced when Ca2+ concentrations exceeded 0.2 g{middle dot}mL-1. Additionally, the number of cysts and the cyst formation rate similarly increased as Ca2+ concentrations increased, but these were reduced when Ca2+ concentrations exceeded 0.4 g{middle dot}mL-1. Lastly, S. trochoidea absorbed Ca2+ from the water when cysts were formed and under high Ca2+ concentrations, more calcareous thorn cysts formed.

ecology↗

bioRxiv2020

Gating transitions and modulation of a hetero-octameric AMPA glutamate receptor

AMPA glutamate receptors (AMPARs) mediate the majority of excitatory transmission in the brain, and enable synaptic plasticity that underlies learning 1. A diverse array of AMPAR signaling complexes are established by receptor auxiliary subunits, associating in various combinations to modulate trafficking, gating and synaptic strength 2. However, their mechanisms of action are poorly understood. Here, we determine cryo-electron microscopy structures of the heteromeric GluA1/2 receptor assembled with both TARP-{gamma}8 and CNIH2, the predominant AMPAR complex in the forebrain, in both resting and active states (at 3.2 and 3.7 [A], respectively). Consequential for gating regulation, two {gamma}8 and two CNIH2 subunits lodge at distinct sites beneath the ligand-binding domains of the receptor tetramer, with site-specific lipids shaping each interaction. Activation leads to a stark asymmetry between GluA1 and GluA2 along the ion conduction path, and an outward expansion of the channel triggers counter-rotations of both auxiliary subunit pairs, that promotes the active-state conformation. In addition, both {gamma}8 and CNIH2 pivot towards the pore exit on activation, extending their reach for cytoplasmic receptor elements. CNIH2 achieves this through its uniquely extended M2 helix, which has transformed this ER-export factor into a powerful positive AMPAR modulator, capable of providing hippocampal pyramidal neurons with their integrative synaptic properties.

neuroscience↗

bioRxiv2020

The pan-cancer lncRNA PLANE regulates an alternative splicing program to promote cancer pathogenesis

Genomic amplification of the distal portion of chromosome 3q, which encodes a number of oncogenic proteins, is one of the most frequent chromosomal abnormalities in malignancy. Here we functionally characterise a non-protein product of the 3q region, the long noncoding RNA (lncRNA) PLANE, which is upregulated in diverse cancer types through copy number gain as well as E2F1-mediated transcriptional activation. PLANE forms an RNA-RNA duplex with the nuclear receptor co-repressor 2 (NCOR2) pre-mRNA at intron 45, binds to heterogeneous ribonucleoprotein M (hnRNPM) and facilitates the association of hnRNPM with the intron, thus leading to repression of the alternative splicing (AS) event generating NCOR2-202, a major protein-coding NCOR2 AS variant. In consequence, PLANE promotes cancer cell proliferation and tumorigenicity and its upregulation is associated with poor patient outcomes. These results uncover the function and regulation of PLANE and suggest that PLANE may constitute a therapeutic target in the pan-cancer context.

cancer biology↗

bioRxiv2020

Light-activated tetrazines enable live-cell spatiotemporal control of bioorthogonal reactions

Bioorthogonal ligations encompass coupling chemistries that have considerable utility in living systems.1-3 Among the numerous bioorthogonal chemistries described to date, cycloaddition reactions between tetrazines and strained dienophiles are widely used in proteome, lipid, and glycan labeling due to their extremely rapid kinetics.4,5 In addition, a variety of functional groups can be released after the cycloaddition reaction,6,7 and drug delivery triggered by in vivo tetrazine ligation8 is in human phase I clinical trials.9 While applications of tetrazine ligations are growing in academia and industry, it has so far not been possible to control this chemistry to achieve the high degrees of spatial and temporal precision necessary for modifying mammalian cells with single-cell resolution. Here we demonstrate visible light-activated formation of tetrazines from photocaged dihydrotetrazines, which enables live-cell spatiotemporal control of rapid biorthogonal cycloaddition reactions between tetrazines and dienophiles such as trans-cyclooctenes (TCOs). Photocaged dihydrotetrazines are stable in conditions that normally degrade tetrazines, enabling efficient early-stage incorporation of bioorthogonal handles into biomolecules such as peptides. Photocaged dihydrotetrazines allow the use of non-toxic visible light to trigger tetrazine ligations on live mammalian cells. By tagging reactive phospholipids with fluorophores, we demonstrate modification of HeLa cell membranes with single-cell spatial resolution. Finally, we show that photo-triggered therapy is possible by coupling tetrazine photoactivation with strategies that uncage prodrugs in response to tetrazine ligation, opening up new methods for photopharmacology and precision drug delivery using bioorthogonal chemistry.

biochemistry↗

bioRxiv2020

Retinal glial cells under hypoxia possibly function in mammalian myopia by responding to mechanical stimuli, affecting hormone metabolism and peptide secretion

PurposeChanges in the retina and the choroid blood vessels are regularly observed in myopia. The aim of this study is to test if the retinal glial cells, which directly contact blood vessels, play a role in mammalian myopia. MethodWe adapted the common form-deprivation myopia mouse model and used the retina slice and whole-mount immunofluorescence technique to evaluate changes in the morphology, and distribution of retinal glial cells. We then searched the Gene Expression Omnibus database for series on myopia and retinal glial cells (astrocytes and Muller cells). Using review articles and the National Center for Biotechnology Information gene database, we obtained clusters of myopia-related gene lists. By searching SwissTargetPrediction, we collected information on atropine target proteins. We then used online tools to find the Gene Ontology analysis enriched clusters, pathways, and proteins that provide correlative evidence. ResultGlial fibrillary acidic protein fluorescence was observed in mice eyes that were covered and deprived of light for five days compared to uncovered eyes, and the cell morphology became more star-like. From in silico experiments, we identified several pathways and proteins that were common to both myopia and retinal glial cells in hypoxic conditions. The common pathways in human astrocytes represented response to mechanical stimuli, peptide secretion, skeletal system development, and monosaccharide binding. In mouse Muller cells, the pathways represented membrane raft and extracellular structure organisation. The proteins common to myopia and hypoxic glial cells were highly relevant to atropine target proteins. ConclusionRetinal astrocytes and Muller cells under hypoxic conditions contribute to the development of myopia, and may be a valid target for atropine.

bioinformatics↗