Biology subjects

Xu, H.

Publications and source records attributed to Xu, H..

At least 307 records · Page 17Linked to original sources

bioRxiv2022

Huntingtin-KIF1A-mediated axonal transport of synaptic vesicle precursors influences synaptic transmission and motor skill learning in mice

Neurotransmitters are released at synapses by synaptic vesicles (SVs), which originate from SV precursors (SVPs) that have traveled along the axon. Because each synapse maintains a pool of SVs, only a small fraction of which are released, it is unclear whether axonal transport of SVPs modifies synaptic function. Here, studying the corticostriatal network both in microfluidic devices and in mice, we find that phosphorylation of the Huntingtin protein (HTT) causes it to recruit the kinesin motor KIF1A, which in turn increases axonal transport of SVPs and synaptic glutamate release. In mice, constitutive HTT phosphorylation leads to SV over-accumulation at synapses, increases the probability of SV release, and impairs motor skill learning on the rotating rod. Silencing KIF1A in these mice restored SV transport and motor skill learning to wild-type levels. Axonal SVP transport within the corticostriatal network thus influences synaptic plasticity and motor skill learning.

neuroscience↗

bioRxiv2022

Conserved and divergent peptide substrate binding properties of bacterial Hsp70

The 70-kDa heat shock protein (Hsp70) interacts with the polypeptide segments of abundant native proteins to fulfill various cellular activities in both stress and normal conditions. However, a non-native linear polypeptide NR (NRLLLTG) is widely used for the study of Hsp70 substrate binding properties, which is too simple to reflect the complex status of Hsp70 substrates in living organisms. To further broaden our knowledge in this area, we screened 2645 polypeptides derived from 78 biologically relevant proteins and identified eight native peptide substrates (named VP1[~]VP8) for bacterial Hsp70 DnaK. Consistent with previous findings, the amino acid distribution in VP1[~]VP8 were enriched in aliphatic and basic residues, and most of their residues were buried in folded proteins as well. Besides, the substrate binding properties for seven polypeptides were largely the same as observed in NR, suggesting their conserved binding mode to DnaK. However, VP5, which contains more percentage of positively charged residues, demonstrates divergent substrate binding properties during in-vitro biochemical studies. Moreover, VP5 efficiently inhibits the refolding activity of DnaK and bacterial viability, implying its potential to be a good lead peptide for antibacterial drug development.

biochemistry↗

bioRxiv2022

A lncRNA identifies IRF8 enhancer element in negative feedback control of dendritic cell differentiation

Transcription factors play a determining role in lineage commitment and cell differentiation. Interferon regulatory factor 8 (IRF8), a hematopoietic transcription factor, is prominently upregulated in dendritic cells (DC) by autoactivation and controls DC differentiation and function. However, it is unclear how IRF8 autoactivation is controlled and eventually limited. Here we identified a novel long non-coding RNA transcribed from the +32 kb enhancer downstream of IRF8 transcription start site and expressed specifically in plasmacytoid DC (pDC), referred to as lncIRF8. A sequence element of the lncIRF8 promoter, but not lncIRF8 itself, is crucial for pDC and classical DC type 1 (cDC1) differentiation. In DC development IRF8 autoactivation is first initiated by flanking enhancers and then second controlled by feedback inhibition through the lncIRF8 promoter element in the +32 kb enhancer. Our work reveals a previously unrecognized negative feedback loop of IRF8 that orchestrates its own expression and thereby controls DC differentiation.

cell biology↗

bioRxiv2022

Fibrillarin-1 and Fibrillarin-2 are required for divergent cell lineage development in planarian homeostasis and regeneration

Ribosome heterogeneity has been revealed to exist in different cell types during development. However, the function and regulatory mechanisms of ribosome heterogeneity in missing tissue regeneration have yet to be reported. We used the planarian Schmidtea mediterranea with whole-body regenerative capability as a model and revealed the function of the rRNA modification protein fibrillarin in cell lineage development and tissue regeneration. We identified two fibrillarin homologs in planarian, Smed-fbl-1 (fbl-1) and Smed-fbl-2 (fbl-2), with distinct expression patterns. While fbl-2 regulates stem cell proliferation and multiple progenitor cell differentiation, fbl-1 participates in epidermal lineage late-stage specification and wound response. This study indicates that fibrillarin, a nucleolar protein, can respond to wounds and function in distinct cell types, suggesting the existence and critical roles of ribosome heterogeneity in stem cells and tissue regeneration.

developmental biology↗

bioRxiv2022

Stability of SARS-CoV-2 in cold-chain transportation environments and the efficacy of disinfection measures

Cold-chain environment could extend the survival duration of SARS-CoV-2 and increases the risk of transmission. However, the effect of clod-chain environmental factors and packaging materials on SARS-CoV-2 stability and the efficacy of intervention measures to inactivate SARS-CoV-2 under cold-chain environment remains uncertain. This study aimed to unravel cold-chain environmental factors that preserved the stability of SARS-CoV-2 and disinfection measures against SARS-CoV-2 under the cold-chain environment. The spike gene of SARS-CoV-2 isolated from Wuhan hu-1 was used to construct the SARS-CoV-2 pseudovirus and used as model of the SARS-CoV-2 virus. The decay rate of SARS-CoV-2 pseudovirus in the cold-chain environment, various types of packaging material surfaces i.e., PE plastic, stainless steel, Teflon and cardboard, and in frozen seawater was investigated. The influence of LED visible light(wavelength 450 nm-780 nm) and airflow movement on the stability of SARS-CoV-2 pseudovirus at -18{degrees} C were subsequently assessed. The results show that SARS-CoV-2 pseudovirus decayed more rapidly on porous cardboard surface compared with the non-porous surfaces including PE plastic, stainless steel and Teflon. Compared with 25{degrees} C, the decay rate of SARS-CoV-2 pseudovirus was significantly lower at low temperature. Seawater preserved viral stability both at -18{degrees} C and repeated freeze-thawing cycles compared with deionized water. LED visible light illumination and airflow movement environment at -18{degrees} C reduced the SARS-CoV-2 pseudovirus stability. In conclusion, our results indicate cold-chain temperature and seawater as risk factors for SARS-CoV-2 transmission and LED visible light illumination and airflow movement as possible disinfection measures of SARS-CoV-2 under the cold-chain environment. ImportanceIt is widely recognized that low temperature is a condition for maintaining virus vitality, and cold-chain transportation spreads the events of the SARS-CoV-2 were reported. This study provides that the decay rate of the SARS-CoV-2 pseudovirus at low temperatures varies on different packaging materials, and salt ions present in frozen foods such as seafood may protect virus survival. These results provide evidence for the possibility of SARS-CoV-2 transmission through cold-chain transport and also suggest the importance for disinfection of items. However, the commonly used disinfection methods of ultraviolet radiation and chemical reagents are generally not suitable for the disinfection of frozen food. Our study shows LED visible light illumination and airflow movement as possible disinfection measures of SARS-CoV-2 under the cold-chain environment. This has implications for reducing the long-distance transmission of the virus through cold-chain transportation.

microbiology↗

bioRxiv2022

DeepST: A versatile graph contrastive learning framework for spatially informed clustering, integration, and deconvolution of spatial transcriptomics

Advances in spatial transcriptomics technologies have enabled the gene expression profiling of tissues while retaining its spatial context. Effective exploitation of this data combination requires spatially informed analysis tools to perform three key tasks, spatial clustering, multi-sample integration, and cell type deconvolution. Here, we present GraphST, a novel graph self-supervised contrastive learning method that incorporates spatial location information and gene expression profiles to accomplish all three tasks in a streamlined process while outperforming existing methods in each task. GraphST combines graph neural networks with self-supervised contrastive learning to learn informative and discriminative spot representations by minimizing the embedding distance between spatially adjacent spots and vice versa. With GraphST, we achieved 10% higher clustering accuracy on multiple datasets than competing methods, and better delineated the fine-grained structures in tissues such as the brain and embryo. Moreover, GraphST is the only method that can jointly analyze multiple tissue slices in both vertical and horizontal integration while correcting for batch effects. Lastly, compared to other methods, GraphSTs cell type deconvolution achieved higher accuracy on simulated data and better captured spatial niches such as the germinal centers of the lymph node in experimentally acquired data. We further showed that GraphST can recover the immune cell distribution in different regions of breast tumor tissue and reveal spatial niches with exhausted tumor infiltrating T cells. Through our examples, we demonstrated that GraphST is widely applicable to a broad range of tissue types and technology platforms. In summary, GraphST is a streamlined, user friendly and computationally efficient tool for characterizing tissue complexity and gaining biological insights into the spatial organization within tissues.

bioinformatics↗

bioRxiv2022

Allosteric Regulation of a Synaptic Vesicle Glutamate Transporter

Concentration of neurotransmitter inside synaptic vesicles (SVs) underlies the quantal nature of synaptic transmission. In contrast to many transporters, SV uptake of the principal excitatory neurotransmitter glutamate is driven by membrane potential. To prevent nonquantal efflux of glutamate after SV exocytosis, the vesicular glutamate transporters (VGLUTs) are allosterically inhibited by the neutral pH of the synaptic cleft. We have now determined high-resolution structures of VGLUT2 with a cyclic analog of glutamate bound that defines the mechanism of substrate recognition, a positively charged cytoplasmic vestibule that electrostatically attracts the negatively charged substrate, and modification by palmitoylation that promotes retrieval of the transporter after exocytosis. The structure also incorporates an extensive, cytoplasmic network of electrostatic interactions that acts as a gate. Functional analysis shows how this cytoplasmic gate confers the allosteric requirement for lumenal H+ required to restrict VGLUT activity to SVs.

biochemistry↗

bioRxiv2022

A CDKB/KRP/FB3 cell cycle core complex functions in rice gametes and zygotes

The cell cycle controls division and proliferation of all eukaryotic cells and is tightly regulated at multiple checkpoints by complexes of core cell cycle proteins. Due to the difficulty in accessing female gametes and zygotes of flowering plants, little is known about the molecular mechanisms underlying initiation embryogenesis despite the crucial importance of this process for seed crops. In this study, we reveal four levels of factors involved in rice zygotic cell cycle control and characterize their functions and regulation. Protein-protein interaction studies, including within zygote cells, and in vitro biochemical analyses delineate a model of the zygotic cell cycle core complex for rice. In this model, CDKB1, a major regulator of plant mitosis, is a cyclin (CYCD5)-dependent kinase; its activity is coordinately inhibited by two cell cycle inhibitors, KRP4 and KRP5; and both KRPs are regulated via F-box protein 3 (FB3)-mediated proteolysis. Supporting their critical role in controlling the rice zygotic cell cycle, mutations in KRP4, KRP5, and FB3 result in the compromised function of sperm cells and abnormal organization of female germ units, embryo and endosperm, thus significantly reducing seed-set rates. This work helps reveals regulatory mechanisms controlling the zygotic cell cycle toward seed formation in angiosperms.

cell biology↗

bioRxiv2022

Posterodorsal medial amygdala urocortin-3, GABA and glutamate mediate suppression of LH pulsatility in female mice

The posterodorsal subnucleus of the medial amygdala (MePD) is an upstream modulator of the hypothalamic-pituitary-gonadal (HPG) and hypothalamic-pituitary-adrenal (HPA) axes. Inhibition of MePD urocortin-3 (Ucn3) neurons prevents psychological stress-induced suppression of LH pulsatility while blocking the stress-induced elevations in corticosterone (CORT) secretion in female mice. We explore the neurotransmission and neural circuitry suppressing the GnRH pulse generator by MePD Ucn3 neurons and we further investigate whether MePD Ucn3 efferent projections to the PVN control CORT secretion and LH pulsatility. Ucn3-cre-tdTomato female ovariectomised (OVX) mice were unilaterally injected with AAV-ChR2 and implanted with optofluid cannulae targeting the MePD. We optically activated Ucn3 neurons in the MePD with blue light at 10Hz and monitored the effect on LH pulses. Next, we combined optogenetic stimulation of MePD Ucn3 neurons with pharmacological antagonism of GABAA or GABAB receptors with bicuculline or CGP, respectively, as well as a combination of NMDA and AMPA receptor antagonists, AP5 and CNQX respectively, and observed the effect on pulsatile LH secretion. A separate group of Ucn3-cre-tdTomato OVX mice with 17{beta}-estradiol (E2) replacement were unilaterally injected with AAV-ChR2 in the MePD and implanted with fibreoptic cannulae targeting the PVN. We optically stimulated the MePD Ucn3 efferent projections in the PVN with blue light at 20Hz and monitored the effect on CORT secretion and LH pulses. We reveal for the first time that activation of Ucn3 neurons in the MePD inhibits GnRH pulse generator frequency via GABA and glutamate signalling within the MePD, while MePD Ucn3 projections to the PVN modulate the HPG and HPA axes.

neuroscience↗

bioRxiv2022

Diversity and biogeography of plant leaf bacteria along a latitudinal gradient

Plant-associated microbes are essential for promoting plant well-being, maintaining biodiversity, and supporting ecosystem function. However, little is known about the geographic distribution of plant-microbe symbioses and how they are formed and change along latitudinal gradients. Here we identified leaf bacteria for 328 plant species sampled from 10 forests along a tropical to temperate gradient in China. We analyzed the diversity and composition of plant leaf-associated bacteria and quantified the contributions of hosts, habitats, and neighborhood plants to the plant-bacterial symbiosis. We found a strong latitudinal gradient in leaf bacterial diversity and composition. Bacterial assemblages on leaves were most strongly selected by host plants, and the selection pressure increased with latitude. In contrast, at low latitudes and at large geographical scales multiple factors were found to jointly regulate bacterial community composition. Our result also showed that plant-bacteria symbiotic networks were structured by network hub bacteria taxa with high co-occurrence network centrality, and the abundance of temperate hub taxa was more influenced by host plants than that in tropical forests. For the first time, we documented a previously unrecognized latitudinal gradient in plant-bacterial symbioses that was regulated by a joint effect of multiple factors at low latitudes but mostly by host selection at high latitudes, implying that leaf microbiomes are likely to respond differently to global change along the latitudinal gradient.

ecology↗

bioRxiv2022

Inferring CTCF insulators and anchored loops across human tissues and cell types

CCCTC-binding factor (CTCF) is a transcription regulator which is involved in many cellular processes. How CTCF recognizes DNA sequence to exert chromosome barrier or enhancer blocking effects remains to be fully interrogated. Despite many computational tools were developed to predict CTCF-mediated loops qualitatively or quantitatively, few could specially evaluate the regulatory potential of DNA sequence at CTCF binding sites (CBSs) and how it affects chromatin loop formation. Here, we developed a deep learning model, DeepAnchor, to precisely characterize the binding patterns for different types of CBSs. By incorporating base-wise genomic/epigenomic features, we revealed distinct chromatin and sequence features for CTCF-mediated insulation and looping at a high resolution, such as two sequence motifs flanking the core CTCF motif at loop-associated CBSs. Besides, we leveraged the predicted anchor score to optimize the loop extrusion model and achieved the best performance in predicting CTCF-anchored loops. We established a compendium of context-specific CTCF-anchored loops across 52 human tissue/cell types and found that genomic disruption of CTCF-anchored loops may represent a general causal mechanism of disease pathogenesis. These computational models, together with the established resource, could facilitate the mechanistic research on how the CTCF-mediated cis-regulatory elements (CREs) shapes context-specific gene regulation in cell development and disease progression.

bioinformatics↗

bioRxiv2022

KLF7 Promotes Preadipocyte Proliferation via Activation of the Akt Signaling Pathway by Cis-regulating CDKN3

Kruppel-like transcription factor 7 (KLF7) promotes preadipocyte proliferation; however, its target gene in this process has not yet been identified. Using KLF7 ChIP-seq analysis, we previously showed that a KLF7-binding peak is present upstream of the cyclin-dependent kinase inhibitor 3 gene (CDKN3) in chicken preadipocytes. In the current study, we identified CDKN3 as a target gene of KLF7 that mediates the effects of KLF7 on preadipocyte proliferation. Furthermore, 5'-truncating mutation analysis showed that the minimal promoter was located between CDKN3 nt -160 and nt -7 (relative to the translation initiation codon ATG). KLF7 overexpression increased CDKN3 promoter activity in the DF-1 and immortalized chicken preadipocyte (ICP1) cell lines. Deletion of the putative binding site of KLF7 abolished the promotive effect of KLF7 overexpression on CDKN3 promoter activity. Moreover, CDKN3-knockdown and -overexpression assays revealed that CDKN3 enhanced ICP1 cell proliferation. Flow cytometry analysis showed that CDKN3 accelerated the G1/S transition. Further, we found that KLF7 promoted ICP1 cell proliferation via Akt phosphorylation by regulating CDKN3. Taken together, these results suggest that KLF7 promotes preadipocyte proliferation via activating the Akt signaling pathway by cis-regulating CDKN3, thus driving the G1/S transition.

molecular biology↗

bioRxiv2022

Pathogen-Host Adhesion between SARS-CoV-2 S Proteins from Different Variants and Human ACE2 Probed at Single-Molecule and Single-Cell Levels

Pathogen-Host adhesion is considered the first step of infection for many pathogens such as bacteria and virus. The binding of the receptor binding domain (RBD) of SARS-CoV-2 Spike protein (S protein) onto human angiotensin-converting enzyme 2 (ACE2) is considered as the first step for the SARS-CoV-2 to adhere onto the host cells during the infection. Within three years, a number of variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been found all around the world. Here, we investigated the adhesion of S Proteins from different variants and ACE2 using atomic force microscopy (AFM)-based single-molecule force spectroscopy (SMFS) and single-cell force spectroscopy (SCFS). We found that the unbinding force and binding probability of the S protein from Delta variant to the ACE2 was the highest among the variants tested in our study at both single-molecule and single-cell levels. Molecular dynamics simulation showed that ACE2-RBD (Omicron) complex is destabilized by the E484A and Y505H mutations and stabilized by S477N and N501Y mutations, when compared with Delta variant. In addition, a neutralizing antibody, produced by immunization with wild type RBD of S protein, could effectively inhibit the binding of S proteins from wild type, Delta and Omicron variants onto ACE2. Our results provide new insight for the molecular mechanism of the adhesive interactions between S protein and ACE2 and suggest that effective monoclonal antibody can be prepared using wild type S protein against the different variants.

biophysics↗

bioRxiv2022

NetBCE: An Interpretable Deep Neural Network for Accurate Prediction of Linear B-Cell Epitopes

Identification of B-cell epitopes (BCEs) plays an essential role in the development of peptide vaccines, immuno-diagnostic reagents, and antibody design and production. In this work, we generated a large benchmark dataset comprising 126,779 experimentally-supported, linear epitope-containing regions in 3567 protein clusters from over 1.3 million B cell assays. Analysis of this curated dataset showed large pathogen diversity covering 176 different families. The accuracy in linear BCE prediction was found to strongly vary with different features, while the performance by sequence features was superior to that by structural features. To search more efficient and interpretive feature representations, a ten-layer deep learning framework for linear BCE prediction, namely NetBCE, was developed. NetBCE achieved high accuracy and robust performance with the average area under the curve (AUC) value of 0.846 in five-fold cross validation through automatically learning the informative classification features. NetBCE substantially outperformed the conventional machine learning algorithms and other tools, with an over 22.06% improvement of AUC value compared to other tools using an independent dataset. Through investigating the output of important network modules in NetBCE, epitopes and non-epitopes tended to present in distinct regions with efficient feature representation along the network layer hierarchy. The NetBCE tool will be useful for linear B-cell epitopes identification and more generally, immunological and computational biology research.

bioinformatics↗

bioRxiv2022

Non-equilibrium protein folding and activation by ATP-driven chaperones

AO_SCPLOWBSTRACTC_SCPLOWRecent experimental studies suggest that ATP-driven molecular chaperones can stabilize protein sub-strates in their native structures out of thermal equilibrium. The mechanism of such non-equilibrium protein folding is an open question. Based on available structural and biochemical evidence, I propose here a unifying principle that underlies the conversion of chemical energy from ATP hydrolysis to the conformational free energy associated with protein folding and activation. I demonstrate that non-equilibrium folding requires the chaperones to break at least one of four symmetry conditions. The Hsp70 and Hsp90 chaperones each breaks a different subset of these symmetries and thus they use different mechanisms for non-equilibrium protein folding. I derive an upper bound on the non-equilibrium elevation of the native concentration, which implies that non-equilibrium folding only occurs in slow-folding proteins that adopt an unstable intermediate conformation in binding to ATP-driven chaperones. Contrary to the long-held view of Anfinsens hypothesis that proteins fold to their conformational free energy minima, my results predict that some proteins may fold into thermo-dynamically unstable native structures with the assistance of ATP-driven chaperones, and that the native structures of some chaperone-dependent proteins may be shaped by their chaperone-mediated folding pathways.

biophysics↗

bioRxiv2022

Self-organized canals enable long range directed material transport in bacterial communities

Long-range material transport is essential to maintain the physiological functions of multicellular organisms such as animals and plants. By contrast, material transport in bacteria is often short-ranged and limited by diffusion. Here we report a unique form of actively regulated long-range directed material transport in structured bacterial communities. Using Pseudomonas aeruginosa colonies as a model system, we discover that a large-scale and temporally evolving open channel system spontaneously develops in the colony via shear-induced banding. Fluid flows in the open channels support high-speed (up to 450 {micro}m/s) transport of cells and outer membrane vesicles over centimeters, and help to eradicate colonies of a competing species Staphylococcus aureus. The open channels are reminiscent of human-made canals for cargo transport, and the channel flows are driven by interfacial tension mediated by cell-secreted biosurfactants. The spatial-temporal dynamics of fluid flows in the open channels are qualitatively described by flow profile measurement and mathematical modeling. Our findings demonstrate that mechanochemical coupling between interfacial force and biosurfactant kinetics can coordinate large-scale material transport in primitive life forms, suggesting a new principle to engineer self-organized microbial communities.

biophysics↗

bioRxiv2022

Downregulating PTBP1 fails to convert astrocytes into hippocampal neurons and to alleviate symptoms in Alzheimer's mouse models

Conversion of astroglia into functional neurons has been considered as a promising therapeutic strategy for neurodegenerative diseases. Recent studies reported that downregulation of the RNA binding protein, PTBP1, converts astrocytes into neurons in situ in multiple mouse brain regions, consequently improving pathological phenotypes associated with Parkinsons disease, RGC loss, and aging. Here, we demonstrate that PTBP1 downregulation using antisense oligonucleotides or an astrocyte specific AAV-mediated shRNA system fails to convert hippocampal astrocytes into neurons in WT, and {beta}-amyloid (5xFAD) and tau (PS19) Alzheimers disease (AD) mouse models, and fails to reverse synaptic/cognitive deficits and AD-associated pathology. Similarly, PTBP1 downregulation cannot convert astrocytes into neurons in the striatum and substantia nigra. Together, our study suggests that cell fate conversion strategy for neurodegenerative disease therapy through manipulating one single gene, such as PTBP1, warrants more rigorous scrutiny.

neuroscience↗

bioRxiv2022

Accelerated mass transfer from frozen thin films during thin-film freeze-drying

Freeze-drying, or lyophilization, is widely used to produce pharmaceutical solids from temperature-sensitive materials but the process is time and energy inefficient. Herein, using E. coli as a model live organism, whose viability in dry powders is highly sensitive to the water content in the powders, we demonstrated that the drying rate of thin-film freeze-drying (TFFD) is significantly higher than that of the conventional shelf freeze-drying, likely because the large total surface area from the loosely stacked frozen thin films and the low thickness of the thin-films enable faster and more efficient mass transfer during freeze-drying. The highly porous nature and high specific surface area of the thin-film freeze-dried powders may have contributed to the faster mass transfer as well. Moreover, we demonstrated that TFFD can be applied to produce dry powders of E. coli and L. acidophilus with minimum bacterial viability loss (i.e., within one log reduction), and the L. acidophilus dry powder is suitable for intranasal delivery. It is concluded that TFFD technology is promising in addressing the time-and cost-inefficient issue of conventional shelf freeze-drying.

pharmacology and toxicology↗