Biology subjects

Zheng, T.

Publications and source records attributed to Zheng, T..

At least 73 records · Page 4Linked to original sources

bioRxiv2023

Single-cell transcriptome analysis on the anatomic positional heterogeneity of pig skin

Different anatomic locations of the body skin dermis come from different origins, and its positional hereditary information can be maintained in adults, while highly resolvable cellular specialization is less well characterized in different anatomical regions. Pig is regarded as excellent model for human research in view of its similar physiology to human. In this study, we performed single-cell RNA sequencing of six different anatomical skin regions from the Chenghua pig with superior skin thickness trait. We obtained 215,274 cells, representing seven cell types, among which we primarily characterized the heterogeneity of smooth muscle cells, endothelial cells and fibroblasts. We identified several phenotypes of smooth muscle cell and endothelial cell and presented genes expression of pathways such as the immune response in different skin regions. By comparing differentially expressed fibroblast genes among different skin regions, we considered TNN, COL11A1, and INHBA as candidate genes for facilitating ECM accumulation. These findings of heterogeneity in the main three cell types from different anatomic skin sites will contribute to a better understanding of hereditary information and places the potential focus on skin generation, transmission and transplantation, paving the foundation for human skin priming.

cell biology↗

bioRxiv2023

Variability in Forest Plant Traits along the Western Ghats of India and Their Environmental Drivers at Different Resolutions

Identifying key environmental drivers for plant functional traits is an important step to understanding and predicting ecosystem responses to a changing climate. Imaging spectroscopy offers great potential to map plant traits at fine resolution across broad regions and then assess controls on their variation across spatial resolutions. We applied permutational partial least-squares regression to map seven key foliar chemical and morphological traits using NASAs Airborne Visible/Infrared Imaging Spectrometer-Next Generation (AVIRIS-NG) for six sites spanning the Western Ghats of India. We studied the variation of trait space using principal components analysis at spatial resolutions from the plot level (4m), community level (30m and 100m) to the ecosystem level (1000m). We observed a consistent pattern of trait space across different resolutions, with one axis representing the traditional leaf economic spectrum defined by foliar nitrogen concentration and leaf mass per area (LMA) and another axis representing leaf structure and defense defined by fiber, lignin, and total phenolics. We also observed consistent directionality of environment-trait correlations across resolutions with generally higher predictive capacity of our environment-traits models at coarser resolutions. Among the seven traits, total phenolics, fiber, and lignin showed strong environmental dependencies across sites, while calcium, sugar, and nitrogen were significantly affected by site conditions. Models incorporating site as a fixed effect explained more than 50% of the trait variance at 1000m resolution. LMA showed little dependence on both environment and site conditions, implying other factors such as species composition and perhaps site history strongly affect variation in LMA. Our results show that reliable trait-trait relationships can be identified in coarse resolution imagery, but that local scale trait-trait relationships (resolutions finer than 30m) are not sensitive to broad-scale abiotic/biotc factors.

ecology↗

bioRxiv2022

Promoting Axon Regeneration By Inhibiting RNA N6-methyladenosine Demethylase ALKBH5

A key limiting factor of successful axon regeneration is the intrinsic regenerative ability in both the peripheral (PNS) and central nervous system (CNS). Previous studies have identified axon regeneration regulators that act on gene expression in injured neurons. However, it is less known if RNA modifications play a role in this process. Here, we systematically screened the functions of all common m6A modification-related enzymes in axon regeneration and report ALKBH5, an evolutionarily conserved RNA m6A demethylase, as a regulator of axonal regeneration. In PNS, knockdown of ALKBH5 enhanced sensory axonal regeneration, whereas overexpressing ALKBH5 impaired axonal regeneration in an m6A-dependent manner. Mechanistically, ALKBH5 increased the stability of Lpin2 mRNA and thus limited regenerative growth associated lipid metabolism in DRG neurons. Moreover, in CNS, knockdown of ALKBH5 enhanced the survival and axonal regeneration of retinal ganglion cells after optic nerve injury. Together, our results suggest a novel mechanism regulating axon regeneration and point ALKBH5 as a potential target for promoting axon regeneration in both PNS and CNS.

neuroscience↗

bioRxiv2022

A stable reference human transcriptome and proteome as a standard for reproducible omics experiments

In recent years, the development of high-throughput omics technology has greatly promoted the development of biomedicine. However, the poor reproducibility of omics techniques limits its application. It is necessary to use standard reference materials of complex RNAs or proteins to test and calibrate the accuracy and reproducibility of omics workflows. However, the transcriptome and proteome of most cell lines shift during culturing, which limits their applicability to serve as standard samples. In this study, we demonstrated that the human hepatocellular cell line MHCC97H has a very stable transcriptome (R2=0.966-0.995) and proteome (R2=0.934-0.976 for DDA, R2=0.942-0.986 for DIA) after 9 subculturing generations, which allows this stable standard sample to be stably produced on an industrial scale for several decades. Moreover, this stability was maintained across labs and platforms. In sum, our results justified a omics standard reference material and reference datasets for transcriptomic and proteomics research. This helps to further standardize the workflow and data quality of omics techniques and thus promotes the application of omics technology in precision medicine.

cell biology↗

bioRxiv2022

Self-regulation of the nuclear pore complex enables clogging-free crowded transport

Nuclear pore complexes (NPCs) are the main conduits for macromolecular transport into and out of the nucleus of eukaryotic cells. The central component of the NPC transport mechanism is an assembly of intrinsically disordered proteins (IDPs) that fills the NPC channel. The channel interior is further crowded by large numbers of simultaneously translocating cargo-carrying and free transport proteins. How the NPC can efficiently, rapidly and selectively transport varied cargoes in such crowded conditions remains ill understood. Past experimental results suggest that the NPC is surprisingly resistant to clogging and that transport may even become faster and more efficient as the concentration of transport protein increases. To understand the mechanisms behind these puzzling observations, we construct a computational model of the NPC comprising only a minimal set of commonly-accepted consensus features. This model qualitatively reproduces the previous experimental results and identifies self-regulating mechanisms that relieve crowding. We show that some of the crowding-alleviating mechanisms - such as preventing saturation of the bulk flux - are "robust" and rely on very general properties of crowded dynamics in confined channels, pertaining to a broad class of selective transport nanopores. By contrast, the counter-intuitive ability of the NPC to leverage crowding to achieve more efficient single molecule translocation is "fine-tuned" and relies on the particular spatial architecture of the IDP assembly in the NPC channel.

biophysics↗

bioRxiv2022

Multiplexed and millimeter-scale superresolution imaging of cells and tissue sections via prism-illumination and microfluidics-enhanced DNA-PAINT

Fluorescence nanoscopy has become increasingly powerful for biomedical research, but it has historically afforded a small field-of-view (FOV) around 50 {micro}m x 50 {micro}m at once and more recently up to ~200 {micro}m x 200 {micro}m. Efforts to further increase the FOV in fluorescence nanoscopy have thus far relied on the use of fabricated waveguide substrates, adding cost and sample constraints on the applications. Here we report PRism-Illumination and Microfluidics-Enhanced DNA-PAINT (PRIME-PAINT) for multiplexed fluorescence nanoscopy across millimeter-scale FOVs. Built upon the well-established prism-type total internal reflection microscopy, PRIME-PAINT achieves robust single-molecule localization with up to ~520 {micro}m x 520 {micro}m single FOVs and 25-40 nm lateral resolutions. Through stitching, nanoscopic imaging over mm2 sample areas can be completed in as little as 40 minutes per target. An on-stage microfluidics chamber facilitates probe exchange for multiplexing and enhances image quality particularly for formalin-fixed paraffin-embedded (FFPE) tissue sections. We demonstrate the utility of PRIME-PAINT by analyzing ~106 caveolae structures in ~1,000 cells and imaging entire pancreatic cancer lesions from patient tissue biopsies. By imaging from nanometers to millimeters with multiplexity and broad sample compatibility, PRIME-PAINT will be useful for building multiscale, Google-Earth-like views of biological systems.

biophysics↗

bioRxiv2022

Neural Implementation of Precise Temporal Patterns in Motor Cortex

One of the most concerned problems in neuroscience is how neurons communicate and convey information through spikes. There is abundant evidence in sensory systems to support the use of precise timing of spikes to encode information. However, it remains unknown whether precise temporal patterns could be generated to drive output in the primary motor cortex (M1), a brain area containing ample recurrent connections that may destroy temporal fidelity. Here, we used a novel brain-machine interface that mapped the temporal order and precision of motor cortex activity to the auditory cursor and reward to guide the generation of precise temporal patterns in M1. During the course of learning, rats performed the "temporal neuroprosthetics" in a goal-directed manner with increasing proficiency. Precisely timed spiking activity in M1 was volitionally and robustly produced under this "temporal neuroprosthetics", demonstrating the feasibility of M1 implementing temporal codes. Population analysis showed that the local network was coordinated in a fine time scale as the overall excitation heightened. Furthermore, we found that the directed connection between neurons assigned to directly control the output ("direct neurons") was strengthened throughout learning, as well as connections in the subnetwork that contains direct neurons. Network models revealed that excitatory gain and strengthening of subnetwork connectivity transitioned neural states to a more synchronous regime, which improved the sensitivity for coincidence detection and, thus, the precision of spike patterns. Therefore, our results suggested the recurrent connections facilitate the implementation of precise temporal patterns instead of impairing them, which provided new perspectives on the fine-timescale activity and dynamics of M1.

neuroscience↗

bioRxiv2021

SoyFGB v2.0: a unique access to variations of Chinese Soybean Gene Bank (CNSGB) germplasm

In Chinese National Soybean GeneBank (CNSGB), we have collected more than 30,000 soybean accessions. However, data sharing for soybean remains an especially sensitive question, and how to share the genome variations within rule frame has been bothering the soybean germplasm workers for a long time. Here we release a big data source named Soybean Functional Genomics & Breeding database (SoyFGB v2.0) (https://sfgb.rmbreeding.cn/), which embed a core collection of 2,214 soybean resequencing genome (2K-SG) from the CNSGB germplasm. This source presents a unique example which may help elucidating the following three major functions for multiple genome data mining with general interests for plant researchers. 1) On-line analysis tools are provided by the Analysis module for haplotype mining in high-throughput genotyped germplasms with different methods. 2) Variations for 2K-SG are provided in SoyFGB v2.0 by Browse module which contains two functions of SNP and InDel. Together with the Gene (SNP & InDel) function embedded in Search module, the genotypic information of 2K-SG for targeting gene / region is accessible. 3) Scaled phenotype data of 42 traits, including 9 quality and 33 quantitative traits are provided by SoyFGB v2.0. With the scaled-phenotype data search and seed request tools under a control list, the germplasm information could be shared without direct downloading the unpublished phenotypic data or information of sensitive germplasms. In a word, the mode of data mining and sharing underlies SoyFGB v2.0 may inspire more ideas for works on genome resources of not only soybean but also the other plants.

genomics↗

bioRxiv2021

Cell free extrachromosomal circular DNA is common in human urine

Cell free extrachromosomal circular DNA (eccDNA) is evolving as a potential biomarker in liquid biopsies for disease diagnosis. In this study, an optimized next generation sequencing-based Circle-Seq method was developed to investigate urinary cell free eccDNA (ucf-eccDNA) from 28 adult healthy volunteers (mean age = 28, 19 males/ 9 females). The genomic distributions and sequence compositions of ucf-eccDNAs were comprehensively characterized. Approximately 1.2 million unique ucf-eccDNAs are identified, covering 14.9% of the human genome. Comprehensive characterization of ucf-eccDNAs show that ucf-eccDNAs contain higher GC content than flanking genomic regions. Most eccDNAs are less than 1000 bp and present four pronounced peaks at 203, 361, 550 and 728 bp, indicating the association between eccDNAs and the numbers of intact nucleosomes. Analysis of genomic distribution of ucf-eccDNAs show that eccDNAs are found in all chromosomes but enriched in chromosomes i.e. chr.17, 19 and 20 with high density of protein-codding genes, CpG islands, SINE and simple repeat elements. Lastly, analysis of sequence motif signatures at eccDNA junction sites reveal that direct repeats (DRs) are commonly found, indicating a potential role of DRs in eccDNA biogenesis. This work underscores the deep sequencing analysis of ucf-eccDNAs and provides a valuable reference resource for exploring potential applications of ucf-eccDNA as diagnostic biomarkers of urogenital disorders in the future. Significance StatementExtrachromosomal circular DNA (eccDNA) is an important genetic element and a biomarker for disease diagnosis and treatment. In this study, we conduct a comprehensive characterization of urinary cell free eccDNA (ucf-eccDNA) in 28 heathy subjects. Over one million ucf-eccDNAs are identified. Ucf-eccDNAs are characterized as high GC content. The size of most ucf-eccDNAs is less than 1000 bp and enriched in four peaks resembling the size of single, double, triple, and quadruple nucleosomes. The genomic distribution of ucf-eccDNAs is enriched in generic regions, protein-coding genes, Alu, CpG islands, SINE and simple repeats. Sequence motif analysis of ucf-eccDNA junctions identified simple direct repeats (DRs) commonly presented in most eccDNAs, suggesting potential roles of DRs in eccDNA biogenesis.

genomics↗

bioRxiv2021

A framework for multiplex imaging optimization and reproducible analysis

Multiplex imaging technologies are increasingly used for single-cell phenotyping and spatial characterization of tissues; however, transparent methods are needed for comparing the performance of platforms, protocols and analytical pipelines. We developed a python software, mplexable, for reproducible image processing and utilize Jupyter notebooks to share our optimization of signal removal, antibody specificity, background correction and batch normalization of the multiplex imaging with a focus on cyclic immunofluorescence (CyCIF). Our work both improves the CyCIF methodology and provides a framework for multiplexed image analytics that can be easily shared and reproduced.

cancer biology↗

bioRxiv2021

A tandem-repeat dimeric RBD protein-based COVID-19 vaccine ZF2001 protects mice and nonhuman primates

A safe, efficacious and deployable vaccine is urgently needed to control COVID-19 pandemic. We report here the preclinical development of a COVID-19 vaccine candidate, ZF2001, which contains tandem-repeat dimeric receptor-binding domain (RBD) protein with alum-based adjuvant. We assessed vaccine immunogenicity and efficacy in both mice and non-human primates (NHPs). ZF2001 induced high levels of RBD-binding and SARS-CoV-2 neutralizing antibody in both mice and NHPs, and also elicited balanced TH1/TH2 cellular responses in NHPs. Two doses of ZF2001 protected Ad-hACE2-transduced mice against SARS-CoV-2 infection, as detected by reduced viral RNA and relieved lung injuries. In NHPs, vaccination of either 25 g or 50 g ZF2001 prevented infection with SARS-CoV-2 in lung, trachea and bronchi, with milder lung lesions. No evidence of disease enhancement is observed in both models. ZF2001 is being evaluated in the ongoing international multi-center Phase 3 trials (NCT04646590) and has been approved for emergency use in Uzbekistan.

immunology↗

bioRxiv2021

Previously uncharacterized interactions between the folded and intrinsically disordered domains impart asymmetric effects on UBQLN2 phase separation

Shuttle protein UBQLN2 functions in protein quality control (PQC) by binding to proteasomal receptors and ubiquitinated substrates via its N-terminal ubiquitin-like (UBL) and C-terminal ubiquitin-associated (UBA) domains, respectively. Between these two folded domains are intrinsically disordered STI1-I and STI1-II regions, connected by disordered linkers. The STI1 regions bind other components, such as HSP70, that are important to the PQC functions of UBQLN2. We recently determined that the STI1-II region enables UBQLN2 to undergo liquid-liquid phase separation (LLPS) to form liquid droplets in vitro and biomolecular condensates in cells. However, how the interplay between the folded (UBL/UBA) domains and the intrinsically-disordered regions mediates phase separation is largely unknown. Using engineered domain deletion constructs, we found that removing the UBA domain inhibits UBQLN2 LLPS while removing the UBL domain enhances LLPS, suggesting that UBA and UBL domains contribute asymmetrically in modulating UBQLN2 LLPS. To explain these differential effects, we interrogated the interactions that involve the UBA and UBL domains across the entire UBQLN2 molecule using NMR spectroscopy. To our surprise, aside from well-studied canonical UBL:UBA interactions, there also exist moderate and weak interactions between the UBL and STI1-I/STI1-II domains, and between the UBA domain and the linker connecting the two STI1 regions, respectively. Our findings are essential for the understanding of both the molecular driving forces of UBQLN2 LLPS and the effects of ligand binding to UBL, UBA, or STI1 domains on the phase behavior and physiological functions of UBQLN2. Impact of Work StatementZheng and Castaneda show that interplay between the folded domains and intrinsically disordered regions regulates liquid-liquid phase separation behavior of UBQLN2, a protein quality control (PQC) shuttle protein. Despite their similar size, the folded UBL and UBA domains inhibit and promote phase separation, respectively, due to their previously uncharacterized, asymmetric interactions with the middle intrinsically-disordered region. These results strongly suggest that PQC components, including proteasomal receptors, are likely to modulate UBQLN2 phase separation behavior in cells.

biophysics↗

bioRxiv2021

Recombinant chimpanzee adenovirus AdC7 expressing dimeric tandem-repeat RBD of SARS-CoV-2 spike protein protects mice against COVID-19

A safe and effective vaccine is urgently needed to control the unprecedented COVID-19 pandemic. Four adenovirus vectored vaccines expressing spike (S) protein have advanced into phase 3 trials, with three approved for use. Here, we generated several recombinant chimpanzee adenovirus (AdC7) vaccines expressing S, receptor-binding domain (RBD) or dimeric tandem-repeat RBD (RBD-tr2). We found vaccination via either intramuscular or intranasal route was highly immunogenic in mice to elicit both humoral and cellular (Th1-based) immune responses. AdC7-RBD-tr2 showed higher antibody responses compared with both AdC7-S and AdC7-RBD. Intranasal administration of AdC7-RBD-tr2 additionally induced mucosal immunity with neutralizing activity in bronchoalveolar lavage fluid. Either single-dose or two-dose mucosal administration of AdC7-RBD-tr2 protected mice against SARS-CoV-2 challenge, with undetectable subgenomic RNA in lung and relieved lung injury. These results support AdC7-RBD-tr2 as a promising COVID-19 vaccine candidate.

immunology↗

bioRxiv2021

Genome recombination-mediated tRNA up-regulation conducts general antibiotic resistance of bacteria at early stage

Bacterial antibiotic resistance sets a great challenge to human health. It seems that the bacteria can spontaneously evolve resistance against any antibiotic within short time without the horizontal transfer of heterologous genes and before accumulating drug-resistant mutations. We have shown that the tRNA-mediated translational regulation counteracts the reactive oxygen species in bacteria. In this study, we demonstrated that isolated and subcultured Escherichia coli elevated its tRNAs under antibiotic stress to rapidly provide antibiotic resistance, especially at the early stage, before upregulating the efflux pump and evolving resistance mutations. The DNA recombination system repaired the antibiotic-induced DNA breakage in the genome, causing numerous structural variations. These structural variations are overrepresented near the tRNA genes, which indicated the cause of tRNA up-regulation. The strains knocking out the recombination system could not up-regulate tRNAs, and coincidently, they could hardly evolve antibiotic resistance in multiple antibiotics, respectively. With these results, we proposed a multi-stage model of bacterial antibiotic resistance in an isolated scenario: the early stage (recombination - tRNA up-regulation - translational regulation); the medium stage (up-regulation of efflux pump); the late stage (resistance mutations). These results also indicated that the bacterial DNA recombination system and tRNA could be targeted to retard the bacterial spontaneous drug resistance.

microbiology↗

bioRxiv2021

The repertoire of testicular extracellular vesicles cargoes and their involvement in inter-compartmental communication required for spermatogenesis

Extracellular vesicles (EVs) secreted by the epididymis and prostate are involved in sperm functions and epigenetic inheritance. However, the EVs in the testis remains unexplored. Here, we have established a testis dissociation protocol that allows the isolation of testicular EVs by minimizing the disruption of fragile sperm cells. We showed that testicular EVs were specifically and efficiently uptaken by somatic cells and germ cells in both interstitial space and seminiferous tubules compartments, including the spermatozoa. We profiled the proteome of testicular EVs and probed the cell types that release them. Moreover, we sequenced the small RNAs cargos of testicular EVs and identified sets of small non-coding RNAs that were overlooked in the testis transcriptome. Selected miRNA candidates in testicular EVs were found in sperm RNA payload and demonstrated specific resistance towards ribonuclease A independent of the vesicle membrane. Small molecule inhibition of EVs secretion increased the apoptosis of germ cells via inter-compartmental communication. Together, our study has provided valuable resources on the cargoes of testicular EVs and revealed the inter-compartmental communication that regulates spermatogenesis and may implicate in paternal inheritance.

cell biology↗

bioRxiv2020

Quantitative proteomics links the LRRC59 interactome to mRNA translation on the ER membrane

Protein synthesis on the endoplasmic reticulum (ER) requires the dynamic coordination of resident membrane proteins and cytoplasmic translation factors. While ER membrane proteins functioning in ribosome association, mRNA anchoring, and protein translocation, have been identified, little is known regarding the higher order organization of ER-localized translation. Here we utilized proximity proteomics to identify neighboring protein networks for the ribosome interactors SEC61{beta}, RPN1, SEC62, and LRRC59. Whereas the SEC61{beta} and RPN1 BioID reporters revealed translocon-associated networks, the SEC62 and LRRC59 reporters identified divergent interactome networks of previously unexplored functions. Notably, the SEC62 interactome is enriched in redox-linked proteins and ER luminal chaperones, whereas the LRRC59 interactome is enriched in SRP pathway components, translation factors, and ER-localized RNA-binding proteins. Analysis of the LRRC59 interactome by native immunoprecipitation identified similar protein and functional enrichments. Combined, these data reveal a functional domain organization for the ER and suggest a key role for LRRC59 in the organization of mRNA translation on the ER. SummaryHannigan et al. characterize the protein interactomes of four ER ribosome-binding proteins, providing evidence that ER-bound ribosomes reside in distinct molecular environments. Their data link SEC62 to ER redox regulation and chaperone trafficking, and suggest a role for LRRC59 in SRP-coupled protein synthesis.

cell biology↗

bioRxiv2020

The dynamic surface proteomes of allergenic fungal conidia

Fungal spores and hyphal fragments play an important role as allergens in respiratory diseases. In this study, we performed trypsin shaving and secretome analyses to identify the surface-exposed proteins and secreted/shed proteins of Aspergillus fumigatus conidia, respectively. We investigated the surface proteome under different conditions, including temperature variation and germination. We found that the surface proteome of resting A. fumigatus conidia is not static, but instead unexpectedly dynamic, as evidenced by drastically different surface proteomes under different growth conditions. Knockouts of two abundant A. fumigatus surface proteins, ScwA and CweA, were found to function only in fine-tuning the cell wall stress response, implying that the conidial surface is very robust against perturbations. We then compared the surface proteome of A. fumigatus to other allergy-inducing molds, including Alternaria alternata, Penicillium rubens, and Cladosporium herbarum, and performed comparative proteomics on resting and swollen conidia, as well as secreted proteins from germinating conidia. We detected 125 protein ortholog groups, including 80 with putative catalytic activity, in the extracellular region of all four molds, and 42 nonorthologous proteins produced solely by A. fumigatus. Ultimately, this study highlights the dynamic nature of the A. fumigatus conidial surface and provides targets for future diagnostics and immunotherapy.

microbiology↗

bioRxiv2019

Alternative polyadenylation mediates genetic regulation of gene expression

With the exception of mRNA splicing, little is known about co-transcriptional or post-transcriptional regulatory mechanisms that link noncoding variation to variation in organismal traits. To begin addressing this gap, we used 3 Seq to characterize alternative polyadenylation (APA) in the nuclear and total RNA fractions of 52 HapMap Yoruba lymphoblastoid cell lines, which we have studied extensively in the past. We identified thousands of polyadenylation sites that are differentially detected in nuclear mRNA and whole cell mRNA, and found that APA is an important mediator of genetic effects on gene regulation and complex traits. Specifically, we mapped 602 apaQTLs at 10% FDR, of which 152 were found only in the nuclear fraction. Nuclear-specific apaQTLs are highly enriched in introns and are also often associated with changes in steady-state expression levels, suggesting a widespread mechanism whereby genetic variants decrease mRNA expression levels by increasing usage of intronic PAS. We identified 24 apaQTLs associated with protein expression levels, but not mRNA expression, and found that eQTLs that are not associated with chromatin QTLs are enriched in apaQTLs. These findings support multiple independent pathways through which genetic effects on APA can impact gene regulation. Finally, we found that 19% of apaQTLs were also previously associated with disease. Thus, our work demonstrates that APA links genetic variation to variation in gene expression levels, protein expression levels, and disease risk, and reveals uncharted modes of genetic regulation.

genomics↗