Biology subjects

Meng, X.

Publications and source records attributed to Meng, X..

At least 127 records · Page 7Linked to original sources

bioRxiv2021

HyMM: Hybrid method for disease-gene prediction by integrating multiscale module structures

MotivationIdentifying disease-related genes is important for the study of human complex diseases. Module structures or community structures are ubiquitous in biological networks. Although the modular nature of human diseases can provide useful insights, the mining of information hidden in multiscale module structures has received less attention in disease-gene prediction. ResultsWe propose a hybrid method, HyMM, to predict disease-related genes more effectively by integrating the information from multiscale module structures. HyMM consists of three key steps: extraction of multiscale modules, gene rankings based on multiscale modules and integration of multiple gene rankings. The statistical analysis of multiscale modules extracted by three multiscale-module-decomposition algorithms (MO, AS and HC) shows that the functional consistency of the modules gradually improves as the resolution increases. This suggests the existence of different levels of functional relationships in the multiscale modules, which may help reveal disease-gene associations. We display the effectiveness of multiscale module information in the disease-gene prediction and confirm the excellent performance of HyMM by 5-fold cross-validation and independent test. Specifically, HyMM with MO can more effectively enhance the ability of disease-gene prediction; HyMM (MO, RWR) and HyMM (MO, RWRH) are especially preferred due to their excellent comprehensive performance, and HyMM (AS, RWRH) is also good choice due to its local performance. We anticipate that this work could provide useful insights for disease-module analysis and disease-gene prediction based on multi-scale module structures. Availabilityhttps://github.com/xiangiu0208/HvMM Contactlimin@mail.csu.edu.cn Supplementary informationSupplementary data are available at Bioinformatics online.

bioinformatics↗

bioRxiv2021

Discovery of SARS-CoV-2 papain-like protease inhibitors through a combination of high-throughput screening and FlipGFP-based reporter assay

The papain-like protease (PLpro) of SARS-CoV-2 is a validated antiviral drug target. PLpro is involved in the cleavage of viral polyproteins and antagonizing host innate immune response through its deubiquitinating and deISG15ylating activities, rendering it a high profile antiviral drug target. Through a FRET-based high-throughput screening, several hits were identified as PLpro inhibitors with IC50 values at the single-digit micromolar range. Subsequent lead optimization led to potent inhibitors with IC50 values ranging from 0.56 to 0.90 {micro}M. To help prioritize lead compounds for the cellular antiviral assay against SARS-CoV-2, we developed the cell-based FlipGFP assay that is suitable for quantifying the intracellular enzymatic inhibition potency of PLpro inhibitors in the BSL-2 setting. Two compounds selected from the FlipGFP-PLpro assay, Jun9-53-2 and Jun9-72-2, inhibited SARS-CoV-2 replication in Caco-2 hACE2 cells with EC50 values of 8.89 and 8.32 {micro}M, respectively, which were 3-fold more potent than GRL0617 (EC50 = 25.1 {micro}M). The X-ray crystal structures of PLpro in complex with GRL0617 showed that binding of GRL0617 to SARS-CoV-2 induced a conformational change in the BL2 loop to the more closed conformation. Overall, the PLpro inhibitors identified in this study represent promising starting points for further development as SARS-CoV-2 antivirals, and FlipGFP-PLpro assay might be a suitable surrogate for screening PLpro inhibitors in the BSL-2 setting.

pharmacology and toxicology↗

bioRxiv2021

Novel inhibition mechanism of SARS-CoV-2 main protease by ebselen and its derivatives

The global emergence of SARS-CoV-2 has triggered numerous efforts to develop therapeutic options for COVID-19 pandemic. The main protease of SARS-CoV-2 (Mpro), which is a critical enzyme for transcription and replication of SARS-CoV-2, is a key target for therapeutic development against COVID-19. An organoselenium drug called ebselen has recently been demonstrated to have strong inhibition against Mpro and antiviral activity but its molecular mode of action is unknown preventing further development. We have examined the binding modes of ebselen and its derivative in Mpro via high resolution co-crystallography and investigated their chemical reactivity via mass spectrometry. Stronger Mpro inhibition than ebselen and potent ability to rescue infected cells were observed for a number of ebselen derivatives. A free selenium atom bound with cysteine 145 of Mpro catalytic dyad has been revealed by crystallographic studies of Mpro with ebselen and MR6-31-2 suggesting hydrolysis of the enzyme bound organoselenium covalent adduct, formation of a phenolic by-product is confirmed by mass spectrometry. The target engagement of these compounds with an unprecedented mechanism of SARS-CoV-2 Mpro inhibition suggests wider therapeutic applications of organo-selenium compounds in SARS-CoV-2 and other zoonotic beta-corona viruses.

biophysics↗

bioRxiv2021

Potent in vitro Neutralization of SARS-CoV-2 by Hetero-bivalent Alpaca Nanobodies Targeting the Spike Receptor-Binding Domain

Cell entry by SARS-CoV-2 requires the binding between the receptor-binding domain (RBD) of the viral Spike protein and the cellular angiotensin-converting enzyme 2 (ACE2). As such, RBD has become the major target for vaccine development, while RBD-specific antibodies are pursued as therapeutics. Here, we report the development and characterization of SARS-CoV-2 RBD-specific VHH/nanobody (Nb) from immunized alpacas. Seven RBD-specific Nbs with high stability were identified using phage display. They bind to SARS-CoV-2 RBD with affinity KD ranging from 2.6 to 113 nM, and six of them can block RBD-ACE2 interaction. The fusion of the Nbs with IgG1 Fc resulted in homodimers with greatly improved RBD-binding affinities (KD ranging from 72.7 pM to 4.5 nM) and nanomolar RBD-ACE2 blocking abilities. Furthermore, fusion of two Nbs with non-overlapping epitopes resulted in hetero-bivalent Nbs, namely aRBD-2-5 and aRBD-2-7, with significantly higher RBD binding affinities (KD of 59.2 pM and 0.25 nM) and greatly enhanced SARS-CoV-2 neutralizing potency. The 50% neutralization dose (ND50) of aRBD-2-5 and aRBD-2-7 was 1.22 ng/mL ([~]0.043 nM) and 3.18 ng/mL ([~]0.111 nM), respectively. These high-affinity SARS-CoV-2 blocking Nbs could be further developed into therapeutics as well as diagnosis reagents for COVID-19. ImportanceTo date, SARS-CoV-2 has caused tremendous loss of human life and economic output worldwide. Although a few COVID-19 vaccines have been approved in several countries, the development of effective therapeutics including SARS-CoV-2 targeting antibodies remains critical. Due to their small size (13-15 kDa), highly solubility and stability, Nbs are particularly well suited for pulmonary delivery and more amenable to engineer into multi-valent formats, compared to the conventional antibody. Here, we report a serial of new anti-SARS-CoV-2 Nbs isolated from immunized alpaca and two engineered hetero-bivalent Nbs. These potent neutralizing Nbs showed promise as potential therapeutics against COVID-19.

immunology↗

bioRxiv2021

Mitochondrial function is essential for humoral immunity by controlling flux of the TCA cycle, phosphatidic acid and mTOR activity in B cells

The function of mitochondrial respiration during B cell fate decisions and differentiation remains equivocal. This study reveals that selection for mitochondrial fitness occurs during B cell activation and is essential for subsequent plasma cell differentiation. By expressing a mutated mitochondrial helicase in transitional B cells, we depleted mitochondrial DNA during B cell maturation, resulting in reduced oxidative phosphorylation. Although no changes in follicular B cell development were evident, germinal centers, class switch recombination to IgG, plasma cell generation and humoral immunity were diminished. Defective oxidative phosphorylation led to aberrant flux of the tricarboxylic acid cycle and lowered the amount of saturated phosphatidic acid. Consequently, MTOR activity and BLIMP-1 induction were curtailed whereas HIF1, glycolysis and AMPK activity were amplified. Exogenous phosphatidic acid increased mTOR activity in activated B cells. Hence, mitochondrial function is required and selected for in activated B cells for the successful generation of functional plasma cells.

immunology↗

bioRxiv2021

Cas9-cleavage sequences in minimal plasmids enhance non-viral genome targeting of CARs in primary human T cells

T cell genome editing holds great promise to advance a range of immunotherapies but is encumbered by the dependence on difficult-to-produce and expensive viral vectors. Here we have designed small double-stranded plasmid DNA modified to mediate high-efficiency homologous recombination. The resulting chimeric antigen receptor (CAR)-T cells display a similar phenotype, transcriptional profile and in vivo potency as CAR-T cells generated using adeno-associated viral (AAV) vector. This method should simplify and accelerate the use of precision engineering to produce edited T cells for research and clinical purposes.

bioengineering↗

bioRxiv2020

An expedited approach towards the rationale design of non-covalent SARS-CoV-2 main protease inhibitors with in vitro antiviral activity

The main protease (Mpro) of SARS-CoV-2 is a validated antiviral drug target. Several Mpro inhibitors have been reported with potent enzymatic inhibition and cellular antiviral activity, including GC376, boceprevir, calpain inhibitors II and XII, each containing a reactive warhead that covalently modifies the catalytic Cys145. In this study, we report an expedited drug discovery approach by coupling structure-based design and Ugi four-component (Ugi-4CR) reaction methodology to the design of non-covalent Mpro inhibitors. The most potent compound 23R had cellular antiviral activity similar to covalent inhibitors such as GC376. Our designs were guided by overlaying the structure of SARS-CoV Mpro + ML188 (R), a non-covalent inhibitor derived from Ug-4CR, with the X-ray crystal structures of SARS-CoV-2 Mpro + calpain inhibitor XII/GC376/UAWJ247. Binding site analysis suggests a strategy of extending the P2 and P3 substitutions in ML188 (R) to achieve optimal shape complementary with SARS-CoV-2 Mpro. Lead optimization led to the discovery of 23R, which inhibits SARS-CoV-2 Mpro and SARS-CoV-2 viral replication with an IC50 of 0.31 M and EC50 of 1.27 M, respectively. The binding and specificity of 23R to SARS-CoV-2 Mpro were confirmed in a thermal shift assay and native mass spectrometry assay. The co-crystal structure of SARS-CoV-2 Mpro with 23R revealed the P2 biphenyl fits snuggly into the S2 pocket and the benzyl group in the -methylbenzyl faces towards the core of the enzyme, occupying a previously unexplored binding site located in between the S2 and S4 pockets. Overall, this study revealed the most potent non-covalent SARS-CoV-2 Mpro inhibitors reported to date and a novel binding pocket that can be explored for Mpro inhibitor design.

pharmacology and toxicology↗

bioRxiv2020

Fast-Bonito: A Faster Basecaller for Nanopore Sequencing

Oxford Nanopore Technologies (ONT) is a promising sequencing technology that could generate relatively longer sequencing reads compared to the next generation sequencing (NGS) technology. The base calling process is very important for TGS. It translates the original electrical signals from the sequencer to the nucleotide sequence. By doing that, the base calling could significantly influence the accuracy of downstream analysis. Bonito is a recently developed basecaller based on deep neuron network, the neuron network architecture of which is composed of a single convolutional layer followed by three stacked bidirectional GRU layers. Although Bonito achieved the state-of-the-art accuracy, its speed is so slow that it is not likely to be used in production. We therefore implement Fast-Bonito, which introduces systematic optimization to speed up Bonito. Fast-Bonito archives 53.8% faster than the original version on NVIDIA V100 and could be further speed up by HUAWEI Ascend 910 NPU, achieving 565% faster than the original version. The accuracy of Fast-Bonito is also slightly higher than the original Bonito.

bioinformatics↗

bioRxiv2020

Rational Tuning of CAR Tonic Signaling Yields Superior T-Cell Therapy for Cancer

Chimeric antigen receptors (CARs) are modular proteins capable of redirecting immune cells toward a wide variety of disease-associated antigens. Here, we explore the effects of CAR protein sequence and structure on CAR-T cell function. Based on the empirical observation that CD20 CARs with similar sequences exhibit divergent tonic-signaling and anti-tumor activities, we devised engineering strategies that aimed to improve CAR-T cell function by tuning the intensity of tonic signaling. We found that CARs designed to exhibit low but non-zero levels of tonic signaling show robust effector function upon antigen stimulation while avoiding premature functional exhaustion by CAR-T cells. Through alterations of the CARs ligand-binding domain and overall protein conformation, we generated CD20 CAR variants that outperform the CD19 CAR in mouse models of human lymphoma. We further demonstrate that rational modification of protein confirmation can be generalized to improve GD2 CAR-T cell efficacy against neuroblastoma. These findings point to tonic signaling and basal T-cell activation as informative parameters to guide the rational design of next-generation CARs for cancer therapy.

bioengineering↗

bioRxiv2020

Predicting transcriptional responses to cold stress across plant species

Although genome sequence assemblies are available for a growing number of plant species, gene expression responses to stimuli have been catalogued for only a subset of these species. Many genes show altered transcription patterns in response to abiotic stresses. However, orthologous genes in related species often exhibit different responses to a given stress. Accordingly, data on the regulation of gene expression in one species are not reliable predictors of orthologous gene responses in a related species. Here, we trained a supervised classification algorithm to identify genes that transcriptionally respond to cold stress. A model trained with only features calculated directly from genome assemblies exhibited only modest decreases in performance relative to models trained using genomic, chromatin, and evolution/diversity features. Models trained with data from one species successfully predicted which genes would respond to cold stress in other related species. Cross-species predictions remained accurate when training was performed in cold-sensitive species and predictions were performed in cold-tolerant species and vice versa. Models trained with data on gene expression in multiple species outperformed models trained with data from any single species. These results suggest that classifiers trained on stress data from well-studied species may suffice for predicting gene expression patterns in related, less-studied species with sequenced genomes.

plant biology↗

bioRxiv2020

Converting Escherichia coli MG1655 into a chemical overproducer through inactivating defense system against exogenous DNA

Escherichia coli strain K-12 MG1655 has been proposed as an appropriate host strain for industrial production. However, the direct application of this strain suffers from the transformation inefficiency and plasmid instability. Herein, we conducted genetic modifications at a serial of loci of MG1655 genome, generating a robust and universal host strain JW128 with higher transformation efficiency and plasmid stability that can be used to efficiently produce desired chemicals after introducing the corresponding synthetic pathways. Using JW128 as the host, the titer of isobutanol reached 5.76 g/L in shake-flask fermentation, and the titer of lycopene reached 1.91 g/L in test-tube fermentation, 40-fold and 5-fold higher than that of original MG1655, respectively. These results demonstrated JW128 is a promising chassis for high-level production of value-added chemicals.

genetics↗

bioRxiv2020

Structure and inhibition of the SARS-CoV-2 main protease reveals strategy for developing dual inhibitors against Mpro and cathepsin L

The main protease (Mpro) of SARS-CoV-2, the pathogen responsible for the COVID-19 pandemic, is a key antiviral drug target. While most SARS-CoV-2 Mpro inhibitors have a {gamma}-lactam glutamine surrogate at the P1 position, we recently discovered several Mpro inhibitors have hydrophobic moieties at the P1 site, including calpain inhibitors II/XII, which are also active against human cathepsin L, a host-protease that is important for viral entry. To determine the binding mode of these calpain inhibitors and establish a structure-activity relationship, we solved X-ray crystal structures of Mpro in complex with calpain inhibitors II and XII, and three analogues of GC-376, one of the most potent Mpro inhibitors in vitro. The structure of Mpro with calpain inhibitor II confirmed the S1 pocket of Mpro can accommodate a hydrophobic methionine side chain, challenging the idea that a hydrophilic residue is necessary at this position. Interestingly, the structure of calpain inhibitor XII revealed an unexpected, inverted binding pose where the P1 pyridine inserts in the S1 pocket and the P1 norvaline is positioned in the S1 pocket. The overall conformation is semi-helical, wrapping around the catalytic core, in contrast to the extended conformation of other peptidomimetic inhibitors. Additionally, the structures of three GC-376 analogues UAWJ246, UAWJ247, and UAWJ248 provide insight to the sidechain preference of the S1, S2, S3 and S4 pockets, and the superior cell-based activity of the aldehyde warhead compared with the -ketoamide. Taken together, the biochemical, computational, structural, and cellular data presented herein provide new directions for the development of Mpro inhibitors as SARS-CoV-2 antivirals.

biophysics↗

bioRxiv2020

Trk Agonist Drugs Rescue Noise-Induced Hidden Hearing Loss

TrkB agonist drugs are shown here to have a significant effect on the regeneration of afferent cochlear synapses after noise-induced synaptopathy. The effects were consistent with regeneration of cochlear synapses that we observed in vitro after synaptic loss due to kainic acid-induced glutamate toxicity and were elicited by administration of TrkB agonists, amitriptyline and 7,8-dihydroxyflavone, directly into the cochlea via the posterior semicircular canal 48 h after exposure to noise. Synaptic counts at the inner hair cell and wave 1 amplitudes in the ABR were partially restored 2 weeks after drug treatment. Effects of amitriptyline on wave 1 amplitude and afferent auditory synapse numbers in noise-exposed ears after systemic (as opposed to local) delivery were profound and long-lasting; synapses in the treated animals remained intact one year after the treatment. However, the effect of systemically delivered amitriptyline on synaptic rescue was dependent on dose and the time window of administration: it was only effective when given before noise exposure at the highest injected dose. The long-lasting effect and the efficacy of post-exposure treatment indicate a potential broad application for the treatment of synaptopathy, which often goes undetected until well after the original damaging exposure(s).

neuroscience↗

bioRxiv2020

BATF3-dependent induction of IL-27 by B cells bridges the innate and adaptive stages of the antibody response

B cells are exposed to innate and T cell stimuli during the antibody response, although whether and how they functionally integrate such signals are unclear. Here we have identified IL-27 as the cytokine specifically produced by murine B cells upon sequential stimulation by TLR ligands and then CD154 and IL-21, the hallmark factors of T follicular helper cells, and during the T-dependent antibody response to a conjugated hapten or virus infection. B-cell Il27p28 transcription is concomitant with increased locus accessibility and depends on newly induced BATF3 transcription factor. IL-27-producing B cells are inefficient in antibody secretion, but cooperate with IFN{gamma} to promote proliferation, survival, class-switching and plasma cell differentiation of CD40-activated B cells, leading to optimal IgG2a and IgG1 responses. Overall, IL-27-producing B cells function as "helper" B cells that integrate the innate and adaptive stages of the antibody response. One-sentence summaryB cells integrate innate TLR and adaptive CD40 signals to induce BATF3 transcription factor for production of IL-27, which together with INFg optimizes antibody responses.

immunology↗

bioRxiv2020

Hypomyelination leads to alterations in inhibitory function and parvalbumin-positive neuron density in the auditory cortex

For a long time, myelin was thought to be restricted to excitatory neurons, and studies on dysmyelination focused primarily on excitatory cells. Recent evidence showed that axons of inhibitory neurons in the neocortex are also myelinated, but the role of myelin on inhibitory circuits remains unknown. Here we studied the impact of mild hypomyelination on both excitatory and inhibitory connectivity in the primary auditory cortex (A1) with well-characterized mouse models of hypomyelination due to loss of oligodendrocyte ErbB receptor signaling. Using laser-scanning photostimulation, we found that mice with mild hypomyelination have reduced functional inhibitory connections to A1 L2/3 neurons without changes in excitatory connections, resulting in altered excitatory/inhibitory balance. These effects are not associated with altered expression of GABAergic and glutamatergic synaptic components, but with reduced density of parvalbumin-positive (PV+) neurons, which reflects reduced PV expression by interneurons rather than PV+ neuronal loss. While immunostaining shows that hypomyelination occurs in both PV+ and PV- axons, there is a strong correlation between MBP and PV expression suggesting that myelination influences PV expression. Together, the results demonstrate that mild hypomyelination impacts A1 neuronal networks, reducing inhibitory activity, and shifting networks towards excitation.

neuroscience↗

bioRxiv2020

Developmental Normalization of Phenomics Data Generated by High Throughput Plant Phenotyping Systems

BackgroundSowing time is commonly used as the temporal reference for Arabidopsis thaliana (Arabidopsis) experiments in high throughput plant phenotyping (HTPP) systems. This relies on the assumption that germination and seedling establishment are uniform across the population. However, individual seeds have different development trajectories even under uniform environmental conditions. This leads to increased variance in quantitative phenotyping approaches. We developed the Digital Adjustment of Plant Development (DAPD) normalization method. It normalizes time-series HTPP measurements by reference to an early developmental stage and in an automated manner. The timeline of each measurement series is shifted to a reference time. The normalization is determined by cross-correlation at multiple time points of the time-series measurements, which may include rosette area, leaf size, and number. ResultsThe DAPD method improved the accuracy of phenotyping measurements by decreasing the statistical dispersion of quantitative traits across a time-series. We applied DAPD to evaluate the relative growth rate in A. thaliana plants and demonstrated that it improves uniformity in measurements, permitting a more informative comparison between individuals. Application of DAPD decreased variance of phenotyping measurements by up to 2.5 times compared to sowing-time normalization. The DAPD method also identified more outliers than any other central tendency technique applied to the non-normalized dataset.

plant biology↗

bioRxiv2020

Computational Study of Ions and Water Permeation and Transportation Mechanisms of the SARS-CoV-2 Pentameric E Protein Channel

Coronavirus disease 2019 (COVID-19) is caused by a novel coronavirus (SARS-CoV-2) and represents the causative agent of a potentially fatal disease that is of public health emergency of international concern. Coronaviruses, including SARS-CoV-2, encode an envelope (E) protein, which is a small, hydrophobic membrane protein; the E protein of SARS-CoV-2 has high homology with that of severe acute respiratory syndrome coronavirus. (SARS-CoV) In this study, we provide insights into the function of the SARS-CoV-2 E protein channel and the ion and water permeation mechanisms on the basis of combined in silico methods. Our results suggest that the pentameric E protein promotes the penetration of monovalent ions through the channel. Analysis of the potential mean force (PMF), pore radius and diffusion coefficient reveals that Leu10 and Phe19 are the hydrophobic gates of the channel. In addition, the pore demonstrated a clear wetting/dewetting transition with monovalent cation selectivity under transmembrane voltage, which indicates that it is a hydrophobic voltage-dependent channel. Overall, these results provide structural-basis insights and molecular-dynamic information that are needed to understand the regulatory mechanisms of ion permeability in the pentameric SARS-CoV-2 E protein channel.

biophysics↗

bioRxiv2020

Transient coupling between subplate and subgranular layers to L1 neurons before and during the critical period

Cortical layer 1 (L1) contains a diverse population of interneurons which can modulate processing in superficial cortical layers but the intracortical sources of synaptic input to these neurons and how these inputs change over development is unknown. We here investigated the changing intracortical connectivity to L1 in primary auditory cortex (A1) in slices of mouse A1 across development using laser-scanning photostimulation. Before P10 L1 cells receive most excitatory input from within L1, L2/3, L4 and L5/6 as well as the subplate. Excitatory inputs from all layers increase and peak during P10-P16, the peak of the critical period. Inhibitory inputs followed a similar pattern. Functional circuit diversity in L1 emerges after P16. In adult, L1 neurons receive ascending inputs from superficial L2/3 and subgranular L5/6, but only few inputs from L4. A subtype of L1 neurons, NDNF+ neurons, follow a similar pattern, suggesting that transient hyperconnectivity is a universal feature of developing cortical circuits. Our results demonstrate that deep excitatory and superficial inhibitory circuits are tightly linked in early development and might provide a functional scaffold for the layers in between. These results suggest that early thalamic driven spontaneous and sensory activity in subplate can be relayed to L1 from the earliest ages on, that the critical period is characterized by high transient columnar hyperconnectivity, and that in particular circuits originating in L5/6 and subplate might play a key role.

neuroscience↗