Biology subjects

Zhao, B.

Publications and source records attributed to Zhao, B..

At least 145 records · Page 8Linked to original sources

bioRxiv2022

Development of molecular markers of the KTI mutant alleles generated by CRISPR/Cas9-mediated mutagenesis in soybean

The digestibility of soybean meal can be severely impacted by trypsin inhibitor (TI), one of the most abundant anti-nutritional factors present in soybean seeds. TI can restrain the function of trypsin, a critical enzyme that breaks down proteins in the digestive tract. Soybean accessions with low TI content have been identified. However, it is challenging to breed the low TI trait into elite cultivars due to a lack of molecular markers associated with low TI traits. We identified Kunitz trypsin inhibitor 1 (KTI1, Glyma01g095000) and KTI3 (Glyma08g341500) as two seed-specific TI genes. Mutant kti1 and kti3 alleles carrying small deletions or insertions within the gene open reading frames were created in the soybean cultivar Glycine max cv. Williams 82 (WM82) using the CRISPR/Cas9-mediated genome editing approach. The KTI content and TI activity both remarkably reduced in kti1/3 mutants compared to the WM82 seeds. There was no significant difference in terms of plant growth or maturity days of kti1/3 transgenic and WM82 plants in greenhouse condition. We further identified a T1 line, #5-26, that carried double homozygous kti1/3 mutant alleles, but not the Cas9 transgene. Based on the sequences of kti1/3 mutant alleles in #5-26, we developed markers to co-select for these mutant alleles by using a gel-electrophoresis-free method. The kti1/3 mutant soybean line and associated selection markers will assist in accelerating the introduction of low TI trait into elite soybean cultivars in the future.

plant biology↗

bioRxiv2022

DUnet: A deep learning guided protein-ligand binding pocket prediction

Investigating protein-ligand binding sites is the key step in engineering protein/enzyme activity and selectivity. In this study, we developed a 3D convolutional neural network DUnet that derived from DenseNet and UNet for predicting the protein-ligand binding sites. To train DUnet, the features of protein 3D structure were extracted by describing the atomic physical characters, and the ligand binding sites were used as training labels. DUnet was trained using three dataset, the scPDB dataset (collecting of protein-ligand complexes from Protein Data Bank), scPDB and SC6K (collecting of protein-ligand complexes deposited after January 1st, 2018 from Protein Data Bank) datasets, and scPDB and its derived dataset by rotating the samples in the dataset. DUnet displayed better performance than the current state-of-art methods during the benchmark test using independent validation sets, and enlarging the training set contributed to better accuracy. We developed a small dataset contains commonly used industrial enzymes for testing DUnet and found that it was also accurate in predicting the substrate binding sites. We experimentally characterized the substrate binding sites of microbial transglutaminase according to the prediction and showed the significance of these sites. Finally, DUnet was used to predict the ligand binding sites of Swiss-Prot annotated proteins.

bioinformatics↗

bioRxiv2022

A comprehensive single cell data analysis of in lymphoblastoid cells reveals the role of Super-enhancers in maintaining EBV latency

We probed the lifecycle of EBV on a cell-by-cell basis using single cell RNA sequencing (scRNA-seq) data from nine publicly available lymphoblastoid cell lines (LCL). While the majority of LCLs comprised cells containing EBV in the latent phase, two other clusters of cells were clearly evident and were distinguished by distinct expression of host and viral genes. Notably, both were high expressors of EBV LMP1/BNLF2 and BZLF1 compared to another cluster that expressed neither gene. The two novel clusters differed from each other in their expression of EBV lytic genes, including glycoprotein gene GP350. The first cluster, comprising GP350-LMP1hi cells, expressed high levels of HIF1A and was transcriptionally regulated by HIF1-. Treatment of LCLs with Pevonedistat, a drug that enhances HIF1- signaling, markedly induced this cluster. The second cluster, containing GP350+LMP1hi cells, expressed EBV lytic genes. Host genes that are controlled by super-enhancers (SEs), such as transcription factors MYC and IRF4, had the lowest expression in this cluster. Functionally, the expression of genes regulated by MYC and IRF4 in GP350+LMP1hi cells were lower compared to other cells. Indeed, induction of EBV lytic reactivation in EBV+ AKATA reduced the expression of these SE-regulated genes. Furthermore, CRISPR-mediated perturbation of the MYC or IRF4 SEs in LCLs induced the lytic EBV gene expression, suggesting that host SEs and/or SE target genes are required for maintenance of EBV latency. Collectively, our study revealed EBV associated heterogeneity among LCLs that may have functional consequence on host and viral biology. ImportanceEpstein-Barr virus (EBV) establishes a life-long latency program within host cells. As such, EBV immortalized lymphoblastoid cells (LCLs) often carry the latent EBV genome and only a small percentage of LCLs containing lytic EBV. However, the cellular programs that distinguish latent from lytic cells and the heterogeneity of cells in latent or lytic phases remains poorly explored. To explore these unknowns, we reanalyzed publicly available single cell RNA-seq data from nine LCLs. This approach permitted the simultaneous study of cells in both latent and lytic phases. We identified three cell populations with distinct lytic/latent activity and further characterized the transcriptomes of these cells. We also identified a new role of super-enhancers in regulating EBV lytic replication. Collectively, our studies revealed EBV associated heterogeneity among LCLs that contribute to EBV life cycle and biology.

microbiology↗

bioRxiv2022

Low photosynthetic rate under low light stress inhibited sucrose distribution and transportation to grain

Under the condition of low light, the yield of summer maize decreased significantly, but the decrease of yield under low light stress was not only caused by the lack of photosynthetic assimilates in leaves, but also the transportation and utilization of assimilates by stems and grains. In this study, we investigated the effects of low light stress on leaves, stems and grains of summer maize and the relationship between them. The results showed that the synthesis ability of sucrose and export sucrose to grains ability in leaves decreased under low light. Due to dry matter transfer, the number and area of small vascular bundles in spike node and shank decreased, which restricted the translocation of photoassimilates to grains at filling stage. The activities of SUS and AGPase was decreased in grains under low light stress, which limited the availability of sucrose. The process of leaf synthesis, sucrose loading and sucrose utilization in grains was affected under low light, resulting in relatively higher sucrose concentration in grains than in leaves, forming a "leaf low" - "grain high" sugar concentration gradient, resulting in the opposite hydrostatic pressure, and then feedback inhibition of sucrose output in leaves, reducing sucrose loading and transportation rate. HightlightThe key factor of low light stress reducing summer maize yield was the decrease of leaf photosynthetic rate, resulting in insufficient grain dry matter supply. The sugar concentration gradient between leaves and grains further restricted the sucrose transport from leaves to grains.

plant biology↗

bioRxiv2022

Bone marrow AdipoQ-lineage progenitors are a major cellular source of M-CSF that dominates bone marrow macrophage development, osteoclastogenesis and bone mass

M-CSF is a critical growth factor for myeloid lineage cells, including monocytes, macrophages and osteoclasts. Tissue resident macrophages in most organs rely on local M-CSF. However, it is unclear what specific cells in bone marrow produce M-CSF to maintain myeloid homeostasis. Here, we identify bone marrow AdipoQ-lineage progenitors, but not bone marrow mature adipocytes or peripheral adipose tissue, as a major cellular source of M-CSF, with these AdipoQ-lineage progenitors producing M-CSF at levels much higher than those produced by osteoblast lineage cells. Deficiency of M-CSF in bone marrow AdipoQ-lineage progenitors drastically reduces the generation of bone marrow macrophages and osteoclasts, leading to severe osteopetrosis in mice. Furthermore, the postmenopausal osteoporosis in a mouse model can be significantly alleviated by the lack of M-CSF in bone marrow AdipoQ-lineage progenitors. Our findings identify bone marrow AdipoQ-lineage progenitors as a major cellular source of M-CSF in bone marrow and reveal their crucial contribution to bone marrow macrophage development, osteoclastogenesis, bone homeostasis and pathological bone loss.

cell biology↗

bioRxiv2022

A multispecific antibody confers pan-reactive SARS-CoV-2 neutralization and prevents immune escape

Summary ParagraphDespite effective countermeasures, SARS-CoV-2 persists worldwide due to its ability to diversify and evade human immunity1. This evasion stems from amino-acid substitutions, particularly in the receptor-binding domain of the spike, that confer resistance to vaccines and antibodies 2-16. To constrain viral escape through resistance mutations, we combined antibody variable regions that recognize different receptor binding domain (RBD) sites17,18 into multispecific antibodies. Here, we describe multispecific antibodies, including a trispecific that prevented virus escape >3000-fold more potently than the most effective clinical antibody or mixtures of the parental antibodies. Despite being generated before the evolution of Omicron, this trispecific antibody potently neutralized all previous variants of concern and major Omicron variants, including the most recent BA.4/BA.5 strains at nanomolar concentrations. Negative stain electron microscopy revealed that synergistic neutralization was achieved by engaging different epitopes in specific orientations that facilitated inter-spike binding. An optimized trispecific antibody also protected Syrian hamsters against Omicron variants BA.1, BA.2 and BA.5, each of which uses different amino acid substitutions to mediate escape from therapeutic antibodies. Such multispecific antibodies decrease the likelihood of SARS-CoV-2 escape, simplify treatment, and maximize coverage, providing a strategy for universal antibody therapies that could help eliminate pandemic spread for this and other pathogens.

immunology↗

bioRxiv2022

The energy landscape reshaped by strain-specific mutations underlies the long-range epistasis in NS1 evolution of influenza A virus

The mechanisms underlying how individual mutations affect the protein energy landscape are crucial for understanding how proteins evolve. However, predicting mutational effects remains challenging because of epistasis--the nonadditive interactions between mutations. Here, we investigate the biophysical mechanism of strain-specific epistasis in the nonstructural protein 1 (NS1) of the influenza A virus (IAV). To understand the molecular basis of epistasis, we conducted comprehensive analyses of four NS1s of IAV strains that emerged between 1918 and 2004. We find that strain-specific mutations of NS1s are near-neutral with respect to the association with the p85{beta} subunit of PI3K. However, the individual residues on the p85{beta}-binding interface show long-range epistatic interactions with strain-specific mutations. We reveal that strain-specific mutations reshaped the energy landscape of NS1, leading to long-range epistasis. Our findings offer a high-resolution mechanism of how near-neutral mutations silently alter the biophysical energy landscapes, resulting in diverse background effects during molecular evolution.

biophysics↗

bioRxiv2022

Phosphorylation Status of B beta Subunit Acts as a Switch to Regulate the Function of Phosphatase PP2A in Ethylene-mediated Root Growth Inhibition

The various combinations and regulations of different subunits of phosphatase PP2A holoenzymes underlie their functional complexity and importance. We found that phosphorylation status of B{beta} of PP2A acts as a switch to regulate the activity of PP2A. In the absence of ethylene, phosphorylated B{beta} leads to an inactivation of PP2A; the substrate EIR1 remains to be phosphorylated, preventing the EIR1 mediated auxin transport in epidermis, leading to normal root growth. Upon the ethylene treatment, the dephosphorylated B{beta} mediates the formation of A2-C4-B{beta} protein complex to activate PP2A, resulting in the dephosphorylation of EIR1 to promote auxin transport in epidermis of elongation zone, leading to root growth inhibition. Altogether, our research revealed a novel molecular mechanism by which the dephosphorylation of B{beta} subunit switches on the PP2A activity to dephosphorylate EIR1 to establish EIR1 mediated auxin transport in epidermis in elongation zone for root growth inhibition in response to ethylene. Significance StatementRoot growth is critical to the establishment of planted seedlings. Ethylene plays an important role in the root growth. Yet, the molecular mechanisms that how ethylene regulates the root growth are largely unexplored. This research sheds light on the molecular mechanisms of the determination of the functional specificity and activity of PP2A in response to stimulus in cell type specific manner and reveals the novel molecular mechanism through which ethylene signaling regulates EIR1 dephosphorylation through B{beta} dephosphorylation to active EIR1 mediated auxin transport in epidermis in elongation zone, leading to root growth inhibition.

plant biology↗

bioRxiv2022

Convergent epitope specificities, V gene usage and public clones elicited by primary exposure to SARS-CoV-2 variants

An important consequence of infection with a SARS-CoV-2 variant is protective humoral immunity against other variants. The basis for such cross-protection at the molecular level is incompletely understood. Here we characterized the repertoire and epitope specificity of antibodies elicited by Beta, Gamma and ancestral variant infection and assessed their cross-reactivity to these and the more recent Delta and Omicron variants. We developed a high-throughput approach to obtain immunoglobulin sequences and produce monoclonal antibodies for functional assessment from single B cells. Infection with any variant elicited similar cross-binding antibody responses exhibiting a remarkably conserved hierarchy of epitope immunodominance. Furthermore, convergent V gene usage and similar public B cell clones were elicited regardless of infecting variant. These convergent responses despite antigenic variation may represent a general immunological principle that accounts for the continued efficacy of vaccines based on a single ancestral variant.

immunology↗

bioRxiv2022

Lnc956-TRIM28-HSP90B1 complex on replication forks promotes CMG helicase retention to ensure stem cell genomic stability and embryogenesis

Replication stress is a major source of endogenous DNA damage. Despite that numerous proteins have been identified on replication forks to modulate fork or replication machinery activities, it remains unexplored whether non-coding RNAs can localize on stalled forks and play critical regulatory roles. Here we identify an uncharacterized lncRNA NONMMUT028956 (Lnc956 for short) predominantly expressed in mouse embryonic stem cells. Lnc956 is recruited to stalled replication forks to prevent fork collapse and preserve genomic stability, and is essential for mouse embryogenesis. Mechanistically, it drives assembly of the Lnc956-TRIM28-HSP90B1 ribonucleoprotein (RNP) complex on stalled forks in an inter-dependent manner downstream of ATR signaling. This RNP complex physically associates with MCM2-7 hexamer via TRIM28 and directly regulates the CMG helicase retention on chromatin. The regulation of RNP on CMG retention is mediated by HSP90B1s chaperoning function. These findings reveal a novel pathway which actively regulates replisome retention to prevent fork collapse.

cell biology↗

bioRxiv2022

A Self-attention Graph Convolutional Network for Precision Multi-tumour Early Diagnostics with DNA Methylation Data

DNA methylation data-based precision tumor early diagnostics is emerging as state of the art technology, which could capture the signals of cancer occurrence 3[~]5 years in advance and clinically more homogenous groups. At present, the sensitivity of early detection for many tumors is about 30%, which needs to be significantly improved. Nevertheless, based on the genome wide DNA methylation information, one could comprehensively characterize the entire molecular genetic landscape of the tumors and subtle differences among various tumors. With the accumulation of DNA methylation data, we need to develop high-performance methods that can model and consider more unbiased information. According to the above analysis, we have designed a self-attention graph convolutional network to automatically learn key methylation sites in a data-driven way for precision multi-tumor early diagnostics. Based on the selected methylation sites, we further trained a multi-class classification support vector machine. Large amount experiments have been conducted to investigate the performance of the computational pipeline. Experimental results demonstrated the effectiveness of the selected key methylation sites which are highly relevant for blood diagnosis.

bioinformatics↗

bioRxiv2022

Genome Enrichment of Rare, Unknown Species from Complicated Microbiome by Nanopore Selective Sequencing

Rare species are vital members of a microbial community, but retrieving their genomes is difficult due to their low abundance. The ReadUntil (RU) approach allows nanopore devices to sequence specific DNA molecules selectively in real-time, which provides an opportunity for enriching rare species. However, there is still a gap in RU-based enriching of rare and unknown species in environmental samples whose community composition is unclear, and many species lack corresponding reference in public databases. Here we present metaRUpore to overcome this challenge. We applied metaRUpore to a thermophilic anaerobic digester (TAD) community, it successfully redirected the sequencing throughput from high-abundance populations to rare species while facilitating the recovery of 41 high-quality metagenome-assembled genomes (MAGs) at low sequencing effort. The simplicity and robustness of the approach make it accessible for labs with moderate computational resources and hold the potential to become the standard practice in future metagenomic sequencing of complicated microbiomes.

bioinformatics↗

bioRxiv2022

Spatially resolved profiling of protein conformation and interactions by biocompatible chemical cross-linking in living cells

The protein structures and interactions that maintain and regulate cellular processes in different subcellular organelles are heterogeneous and dynamic. However, it remains challenging to characterize the subcellular specificity and translocation of protein complexes in terms of conformation and interactions. Herein, we developed a spatially resolved protein complex profiling approach by biocompatible chemical cross-linking in living cells (SPACX) to monitor the dynamics of protein conformation, interactions and translocation. The advancement of fast capturing protein complexes in the physiological state, coupled with efficient enrichment of the cross-linked peptides, ensured deep-coverage analysis of the protein interactome in living cells. By ensemble structure refinement with cross-linking restraints, subcellular-specific conformation heterogeneity was identified for PTEN. PTEN displayed a broader range of dynamic conformation changes on the dual specificity domains in the nucleus than in the cytoplasm. Moreover, based on conformational differences, different interacting assemblies involving 25 cytoplasm-exclusively and 18 nucleus-exclusively PTEN-interacting proteins were found to account for diverse biological functions. Upon ubiquitin-proteasome system (UPS) stress, the assembly of PTEN and its interacting partners changed obviously during translocation. We newly identified 36 PTEN-interacting proteins, which were found to be highly enriched in functional pathways closely related to cell apoptosis. Inspiringly, the interactions among PTEN isoforms and their interacting proteins were accessible by the determination of sequence-unique cross-linking interfaces for direct interactions. All these results indicate the promise of SPACX to elucidate the functional heterogeneity of proteins in individual subcellular sociology.

biochemistry↗

bioRxiv2021

Two Stages of Speech Envelope Tracking in Human Auditory Cortex Modulated by Speech Intelligibility

The envelope is essential for speech perception. Recent studies have shown that cortical activity can track the acoustic envelope. However, whether the tracking strength reflects the extent of speech intelligibility processing remains controversial. Here, using stereo-electroencephalogram (sEEG) technology, we directly recorded the activity in human auditory cortex while subjects listened to either natural or noise-vocoded speech. These two stimuli have approximately identical envelopes, but the noise-vocoded speech does not have speech intelligibility. We found two stages of envelope tracking in auditory cortex: an early high-{gamma} (60-140 Hz) power stage (delay {approx} 49 ms) that preferred the noise-vocoded speech, and a late {theta} (4-8 Hz) phase stage (delay {approx} 178 ms) that preferred the natural speech. Furthermore, the decoding performance of high-{gamma} power was better in primary auditory cortex than in non-primary auditory cortex, consistent with its short tracking delay. We also found distinct lateralization effects: high-{gamma} power envelope tracking dominated left auditory cortex, while {theta} phase showed better decoding performance in right auditory cortex. In sum, we suggested a functional dissociation between high-{gamma} power and {theta} phase: the former reflects fast and automatic processing of brief acoustic features, while the latter correlates to slow build-up processing facilitated by speech intelligibility.

neuroscience↗

bioRxiv2021

Mitochondrial dysfunction promotes alternative gasdermin D-mediated inflammatory cell death and susceptibility to infection

Human mutations in mitochondrial-associated genes are associated with inflammatory diseases and susceptibility to infection. However, their mechanistic contributions to immune outcomes remain ill-defined. We discovered that the disease-associated gain-of-function allele Lrrk2G2019S (leucine-rich repeat kinase 2) promotes mitochondrial hyper-fission, depolarization, and oxidative stress in macrophages. In the presence of Lrrk2G2019S-dependent mitochondrial perturbations, AIM2 inflammasome activation promotes more cell death but not more pyroptotic IL-1b release. Instead, inflammasome activation in Lrrk2G2019S macrophages triggers gasdermin D (GSDMD)-mediated mitochondrial pores, driving up ROS-mediated RIPK1/RIPK3/MLKL dependent necroptosis. Consequently, infection of Lrrk2G2019S mice with Mycobacterium tuberculosis elicits hyperinflammation and immunopathology via enhanced neutrophil infiltration. By uncovering that GSDMD promotes non-pyroptotic cell death in Lrrk2G2019S macrophages, our findings demonstrate that altered mitochondrial function can reprogram cell death modalities to elicit distinct immune outcomes. This provides mechanistic insights into why mutations in LRRK2 are associated with susceptibility to chronic inflammatory and infectious diseases. HIGHLIGHTSO_LIAltered mitochondrial homeostasis reprograms cell death modalities C_LIO_LIGSDMD associates with and depolarizes mitochondrial membranes following AIM2 activation C_LIO_LIGSDMD initiates a shift from pyroptotic to necroptotic cell death in Lrrk2G2019S macrophages C_LIO_LILrrk2G2019S elicits hyperinflammation and susceptibility to infection in flies and mice C_LI

immunology↗

bioRxiv2021

Inactivation effect and damage of multi-irradiance by UVCLED on Acinetobacter baumannii

It is acknowledged that the inactivation of ultraviolet has been widely used in various fields. Much literature has been reported that ultraviolet C caused DNA damage to achieve inactivation of microorganisms. There is a lack of unified dose calibration and related parameters in this field. In this study, we used a device consisted of the LED of 272 nm to conduct sterilization experiments against A. baumanii. We confirmed the effectiveness of ultraviolet C sterilization for both sensitive and drug resistance strains and explored the relationship between bactericidal rate and ultraviolet doses under various irradiance. Dose requirements of various irradiance were clarified. High irradiance improved sterilization efficiency greatly. The overall damage to the total genome was observed though gel electrophoresis. Ultrastructure of damaged bacteria were investigated by transmission electron microscope in detail. The study revealed that damage to DNA and to the cytoplasm matrix and ribosomes. The study has yielded the possible effects of ultraviolet light on cells by amplifying the energy. The radiation significantly promoted the production of cell wall and cellular membrane. Significance StatementThe statistical results of bactericidal efficiency are influenced by the quantity of bacteria on the medium. Irradiance on the target surface affects the sterilization efficiency directly. And doses required in low irradiance are much more than high. A high irradiance reduced dosage efficiently to achieve the sterilization which improves the sterilization efficiency. There is a difference between low and high UVC dosage damage to the structure of bacteria. Less energy can make DNA coagulation solidified or be dispersed to the edge. Meanwhile the cytoplasm matrix is ruined. When the energy was enough, there is a boost of cell wall and cellular membrane production. The invisible light causes comprehensive damage to bacteria.

bioengineering↗

bioRxiv2021

Protection from SARS-CoV-2 Delta one year after mRNA-1273 vaccination in nonhuman primates is coincident with an anamnestic antibody response in the lower airway

mRNA-1273 vaccine efficacy against SARS-CoV-2 Delta wanes over time; however, there are limited data on the impact of durability of immune responses on protection. We immunized rhesus macaques at weeks 0 and 4 and assessed immune responses over one year in blood, upper and lower airways. Serum neutralizing titers to Delta were 280 and 34 reciprocal ID50 at weeks 6 (peak) and 48 (challenge), respectively. Antibody binding titers also decreased in bronchoalveolar lavage (BAL). Four days after challenge, virus was unculturable in BAL and subgenomic RNA declined [~]3-log10 compared to control animals. In nasal swabs, sgRNA declined 1-log10 and virus remained culturable. Anamnestic antibody responses (590-fold increase) but not T cell responses were detected in BAL by day 4 post-challenge. mRNA-1273-mediated protection in the lungs is durable but delayed and potentially dependent on anamnestic antibody responses. Rapid and sustained protection in upper and lower airways may eventually require a boost.

immunology↗

bioRxiv2021

Summarizing internal dynamics boosts differential analysis and functional interpretation of super enhancers

Super enhancers (SEs) are broad enhancer domains usually containing multiple constituent enhancers that hold elevated activities in gene regulation. Disruption in one or more constituent enhancers causes aberrant SE activities that lead to gene dysregulation in diseases. To quantify SE aberrations, differential analysis is performed to compare SE activities between cell conditions. The state-of-art strategy in estimating differential SEs relies on overall activities and neglect the changes in length and structure of SEs. Here, we propose a novel computational method to identify differential SEs by weighting the combinatorial effects of constituent-enhancer activities and locations (i.e., internal dynamics). In addition to overall activity changes, our method identified four novel classes of differential SEs with distinct enhancer structural alterations. We demonstrate that these structure alterations hold distinct regulatory impact, such as regulating different number of genes and modulating gene expression with different strengths, highlighting the differentiated regulatory roles of these unexplored SE features. When compared to the existing method, our method showed improved identification of differential SEs that were linked to better discernment of cell-type-specific SE activity and functional interpretation. We implement an R package, DASE, to facilitate the use of our method.

bioinformatics↗