Biology subjects

Peng, D.

Publications and source records attributed to Peng, D..

49 records · Page 3Linked to original sources

bioRxiv2022

Improved fluorescent proteins for dual-color post-embedding CLEM

Post-embedding correlative light and electron microscopy (CLEM) has the advantage of high-precision registration and enables light and electron microscopy imaging of the same slice. However, its broad application has been hampered by the limited available fluorescent proteins (FPs) and low signal-to-background ratio (SBR). Here, we developed a green photoswitchable FP, mEosEM-E with substantially high on/off contrast in EM samples embedded in Epon resin which maximally preserves cellular structures but quenches the fluorescence of FPs. Taking advantage of the photoswitching property of mEosEM-E, the autofluorescence background from the resin was significantly reduced by a subtraction-based CLEM (sCLEM) method. Meanwhile, we identified a red fluorescent protein (RFP) mScharlet-H that exhibited higher brightness and SBR in resin than previously reported RFPs. With mEosEM-E and mScharlet-H, dual-color post-Epon-embedding CLEM images with high SBR and no cross-talk signal were successfully performed to reveal the organization of nucleolar proteins. Moreover, a dissection of the influences of different EM sample preparation steps on the fluorescence preservation for several RFPs provides useful guidance for further probe development.

biophysics↗

bioRxiv2021

The genome of the camphor tree and the genetic and climatic relevance of the top-geoherbalism in this medicinal plant

Camphor tree (Cinnamomum camphora (L.) J. Presl), a species in the magnoliid family Lauraceae, is known for its rich volatile oils and is used as a medical cardiotonic and as a scent in many perfumed hygiene products. Here, we present a high-quality chromosome-scale genome of C. camphora with a scaffold N50 of 64.34 Mb and an assembled genome size of 755.41 Mb. Phylogenetic inference revealed that the magnoliids are a sister group to the clade of eudicots and monocots. Comparative genomic analyses identified two rounds of ancient whole-genome duplication (WGD). Tandem duplicated genes exhibited a higher evolutionary rate, a more recent evolutionary history and a more clustered distribution on chromosomes, contributing to the production of secondary metabolites, especially monoterpenes and sesquiterpenes, which are the principal essential oil components. Three-dimensional analyses of the volatile metabolites, gene expression and climate data of samples with the same genotype grown in different locations showed that low temperature and low precipitation during the cold season modulate the expression of genes in the terpenoid biosynthesis pathways, especially TPS genes, which facilitates the accumulation of volatile compounds. Our study lays a theoretical foundation for policy-making regarding the agroforestry applications of camphor tree.

genomics↗

bioRxiv2021

Previously unrecognized non-reproducible antibody-antigen interactions and their implications for diagnosis of viral infections including COVID-19

Antibody-antigen (Ab-Ag) interactions are canonically described by a model which exclusively accommodates non-interaction (0) or reproducible-interaction (RI) states, yet this model is inadequate to explain often-encountered non-reproducible signals. Here, by monitoring diverse experimental systems and confirmed COVID-19 clinical sera using a peptide microarray, we observed that non-specific interactions (NSI) comprise a substantial proportion of non-reproducible antibody-based results. This enabled our discovery and capacity to reliably identify non-reproducible Ab-Ag interactions (NRI), as well as our development of a powerful explanatory model ("0-RI-NRI-Hook four-state model") that is [mAb]-dependent, regardless of specificity, which ultimately shows that both NSI and NRI are not predictable yet certain-to-happen. In experiments using seven FDA-approved mAb drugs, we demonstrated the use of NSI counts in predicting epitope type. Beyond challenging the centrality of Ab-Ag interaction specificity data in serology and immunology, our discoveries also facilitated the rapid development of a serological test with uniquely informative COVID-19 diagnosis performance.

immunology↗

bioRxiv2021

Proper migration of lymphatic endothelial cells requires survival and guidance cues from arterial mural cells

The migration of lymphatic endothelial cells (LECs) is key for the development of the complex and vast lymphatic vascular network that pervades most of the tissues in an organism. In zebrafish, arterial intersegmental vessels together with chemokines have been shown to promote lymphatic cell migration from the horizontal myoseptum (HM). Here we found that LECs departure from HM coincides with the emergence of mural cells around the intersegmental arteries, raising the possibility that arterial mural cells promote LEC migration. Our live imaging and cell ablation experiments revealed that LECs migrate slower and fail to establish the lymphatic vascular network in the absence of arterial mural cells. We determined that mural cells are a source for the C-X-C motif chemokine 12 (Cxcl12a and Cxcl12b) and vascular endothelial growth factor C (Vegfc). We showed that ERK, a downstream component of Vegfc-Vegfr3 singling cascade, is activated in migrating LECs and that both chemokine and growth factor signalling is required for the robust migration. Furthermore, Vegfc-Vegfr3 has a pro-survival role in LECs during the migration. Together, the identification of mural cells a source for signals that guide LEC migration and survival will be important in the future design for rebuilding lymphatic vessels in the disease contexts.

developmental biology↗

bioRxiv2021

Quantitative comparison of in vitro and in vivo embryogenesis at a single cell resolution

Stem cell engineering is a powerful platform to address numerous challenges in regenerative medicine and developmental biology. Typically, engineered populations are derived by exposing pluripotent stem cells to a series of signaling cues meant to recapitulate developmental milestones, such as the induction of the primitive streak. Morphologic, genetic, transcriptomic, and functional differences between fully differentiated in vivo and engineered populations have long been recognized. However, the correspondence between engineered and native embryonic progenitors has been relatively less well characterized, largely due to their transient and highly heterogenous nature, and therefore many studies have relied on expression of a few canonical markers to ensure that their cells are on the correct path. To address this challenge, we first generated an in vivo gastrulation mouse single cell RNA sequencing (scRNA-seq) reference data set and used it to train a collection of computational tools for comparing cell types, states, regulators, and trajectories across studies. Then we used these tools to evaluate a commonly used protocol designed to promote mesoderm derivation, as well as four previously published biomimetic protocols. Finally, we diversified our toolkits to reach a broader scientific community by implementing our primary analysis tool in Python and as an interactive web application.

developmental biology↗

bioRxiv2020

Mitochondria surveillance systems trigger innate immune responses to bacterial pathogens via AMPK pathway in C. elegans

Pathogen recognition and triggering pattern of host innate immune system is critical to understanding pathogen-host interaction. It is generally accepted that the microbial infection can be recognized by host via pattern-triggered immunity (PTI) or effector-triggered immunity (ETI) responses. Recently, non-PRR-mediated cellular surveillance systems have been reported as an important supplement strategy to PTI and ETI responses. However, the mechanism of how surveillance systems sense pathogens and trigger innate immune responses is largely unknown. In the present study, using Bacillus thuringiensis-Caenorhabditis elegans as a model, we found a new approach for surveillance systems to sense the pathogens through no-PPRs patterns. We reported C. elegans can monitor intracellular energy status through the mitochondrial surveillance system to triggered innate immune responses against pathogenic attack via AMP-activated protein kinase (AMPK). Consider that the mitochondria surveillance systems and AMPK are conserved components from worms to mammals, our study suggests that disrupting mitochondrial homeostasis to activate the immune system through AMPK-dependent pathways may widely existing in animals.

immunology↗

bioRxiv2020

Strain-specific genome evolution in Trypanosoma cruzi, the agent of Chagas disease

The protozoan Trypanosoma cruzi almost invariably establishes life-long infections in humans and other mammals, despite the development of potent host immune responses that constrain parasite numbers. The consistent, decades-long persistence of T. cruzi in human hosts arises at least in part from the remarkable level of genetic diversity in multiple families of genes encoding the primary target antigens of anti-parasite immune responses. However, the highly repetitive nature of the genome - largely a result of these same extensive families of genes - have prevented a full understanding of the extent of gene diversity and its maintenance in T. cruzi. In this study, we have combined long-read sequencing and proximity ligation mapping to generate very high-quality assemblies of two T. cruzi strains representing the apparent ancestral lineages of the species. These assemblies reveal not only the full repertoire of gene family members in the two strains, demonstrating extreme diversity within and between isolates, but also provide evidence of the processes that generate and maintain that diversity, including extensive gene amplification, dispersion of copies throughout the genome and diversification via recombination and in situ mutations. These processes also impact genes not required for or involved in immune evasion, creating unique challenges with respect to preserving core genome function while maximizing genetic diversity.

microbiology↗

bioRxiv2020

SHP2 Inhibition Abrogates Adaptive Resistance to KRASG12C-Inhibition and Remodels the Tumor Microenvironment of KRAS-Mutant Tumors

KRAS is the most frequently mutated oncogene in human cancer, and KRAS inhibition has been a longtime therapeutic goal. Recently, inhibitors (G12C-Is) that bind KRASG12C-GDP and react with Cys-12 were developed. Using new affinity reagents to monitor KRASG12C activation and inhibitor engagement, we found that, reflecting its action upstream of SOS1/2, SHP2 inhibitors (SHP2-Is) increased KRAS-GDP occupancy, enhancing G12C-I efficacy. SHP2-Is abrogated feedback signaling by multiple RTKs and blocked adaptive resistance to G12C-Is in vitro, in xenografts, and in syngeneic KRASG12C-mutant pancreatic ductal adenocarcinoma (PDAC) and non-small cell lung cancer (NSCLC) models. Biochemical analysis revealed enhanced suppression of ERK-, MYC-, anti-apoptotic-, and cell-cycle genes, and increased pro-apoptotic gene expression in tumors from combination-treated mice. SHP2-I/G12C-I also evoked favorable changes in the immune microenvironment, decreasing myeloid suppressor cells, increasing CD8+ T cells, and sensitizing tumors to PD-1 blockade. Experiments using cells expressing inhibitor-resistant SHP2 showed that SHP2 inhibition in PDAC cells is required for tumor regression and remodeling of the immune microenvironment, but also revealed direct inhibitory effects on angiogenesis resulting in decreased tumor vascularity. Our results demonstrate that SHP2-I/G12C-I combinations confer a substantial survival benefit in PDAC and NSCLC and identify additional combination strategies for enhancing the efficacy of G12C-Is.

cancer biology↗

bioRxiv2020

BtToxin_Digger: a comprehensive and high-throughput pipeline for mining toxin protein genes from Bacillus thuringiensis

SummaryBacillus thuringiensis (Bt) which is a spore-forming gram-positive bacterium, has been used as the most successful microbial pesticide for decades. Its toxin genes (cry) have been successfully used for the development of GM crops against pests. We have previously developed a web-based insecticidal gene mining tool BtToxin_scanner, which has been proved to be the most important method for mining cry genes from Bt genome sequences. To facilitate efficiently mining major toxin genes and novel virulence factors from large-scale Bt genomic data, we re-design this tool with a new workflow. Here we present BtToxin_Digger, a comprehensive, high-throughput, and easy-to-use Bt toxin mining tool. It runs fast and can get rich, accurate, and useful results for downstream analysis and experiment designs. Moreover, it can also be used to mine other targeting genes from large-scale genome and metagenome data with the addition of other query sequences.Availability and Implementation The BtToxin_Digger codes and instructions are freely available at https://github.com/BMBGenomics/BtToxin_Digger. A web server of BtToxin_Digger can be found at http://bcam.hzau.edu.cn/BtToxin_Digger.Contact jszheng{at}mail.hzau.edu.cn; m98sun{at}mail.hzau.edu.cn.Competing Interest StatementThe authors have declared no competing interest.View Full Text

bioinformatics↗

bioRxiv2020

Evaluating the transcriptional fidelity of cancer models

BackgroundCancer researchers use cell lines, patient derived xenografts, engineered mice, and tumoroids as models to investigate tumor biology and to identify therapies. The generalizability and power of a model derives from the fidelity with which it represents the tumor type under investigation, however, the extent to which this is true is often unclear. The preponderance of models and the ability to readily generate new ones has created a demand for tools that can measure the extent and ways in which cancer models resemble or diverge from native tumors. MethodsWe developed a machine learning based computational tool, CancerCellNet, that measures the similarity of cancer models to 22 naturally occurring tumor types and 36 subtypes, in a platform and species agnostic manner. We applied this tool to 657 cancer cell lines, 415 patient derived xenografts, 26 distinct genetically engineered mouse models, and 131 tumoroids. We validated CancerCellNet by application to independent data, and we tested several predictions with immunofluorescence. ResultsWe have documented the cancer models with the greatest transcriptional fidelity to natural tumors, we have identified cancers underserved by adequate models, and we have found models with annotations that do not match their classification. By comparing models across modalities, we report that, on average, genetically engineered mice and tumoroids have higher transcriptional fidelity than patient derived xenografts and cell lines in four out of five tumor types. However, several patient derived xenografts and tumoroids have classification scores that are on par with native tumors, highlighting both their potential as faithful model classes and their heterogeneity. ConclusionsCancerCellNet enables the rapid assessment of transcriptional fidelity of tumor models. We have made CancerCellNet available as freely downloadable software and as a web application that can be applied to new cancer models that allows for direct comparison to the cancer models evaluated here.

bioinformatics↗

bioRxiv2019

C1orf109L promote R-loop accumulation induced DNA damage to inhibit cell growth

As a function unknown gene, C1orf109 is lower expression in various cells. Here, we reported that C1orf109L, the longest variant of C1orf109, which interacted with R-loop-regulating proteins to trigger R-loop, a three-stranded nucleic acid structure frequently mediated genome instability, accumulation. C1orf109L induce chronic DNA damage to promote P21 upregulation and strongly inhibits cell growth in vitro and in vivo by arresting the cell cycle in the G2 phase. With camptothecin (CPT), an R-loop activator, treatment, C1orf109L further triggers R-loop accumulation-induced DNA damage and promotes cell death by activating cell-death pathway. Furthermore, CPT treatment increases C1orf109L ubiquitination and turnover, which inhibits cell death and promotes the G0/G1 phase of the cell cycle. Therefore, our data illustrated the mechanisms underlying C1orf109L-related cell growth inhibition and provide feasibility and limitations for C1orf109L as a potential target for cancer therapy.

cell biology↗

bioRxiv2018

Bacillus thuringiensis targets the host intestinal epithelial junctions for successful infection of Caenorhabditis elegans

Pathogenic bacteria use different strategies to infect their hosts including the simultaneous production of pore forming toxins and several virulence factors that help to synergize their pathogenic effects. However, how the pathogenic bacteria are able to complete their life cycle and break out the host intestinal barrier is poorly understood. The infectious cycle of Bacillus thuringiensis (Bt) bacterium in Caenorhabditis elegans is a powerful model system to study the early stages of the infection process. Bt produces Cry pore-forming toxins during the sporulation phase that are key virulence factors involved in Bt pathogenesis. Here we show that during the early stages of infection, the Cry toxins disrupt the midgut epithelial tissue allowing the germination of spores. The vegetative Bt cells then trigger a quorum sensing response that is activated by PlcR regulator resulting in production of different virulence factors, such as the metalloproteinases ColB and Bmp1, that besides Cry toxins are necessary to disrupt the nematode epithelial junctions causing efficient bacterial host infection and dead of the nematode. Overall our work describes a novel mechanism for Bt infection, targeting the epithelial junctions of its host midgut cells.\n\nAuthor summaryThe entomopathogenic bacteria Bacillus thuringiensis (Bt) are used worldwide as biopesticides due to their insecticidal properties. Crystal proteins (Cry) produced by Bt during the sporulation phase of growth are mainly responsible for their insecticidal properties. The infection process of Bt includes three successive steps, virulence, necrotrophic, and sporulation processes. During the virulence process, after ingestion by the susceptible hosts, the Cry toxins form pores in the apical membrane of intestinal cells, inducing favorable conditions for bacterial spore germination. Vegetative bacteria multiply in the host and coordinate their behavior by using the quorum sensor regulator PlcR, which leads to the production of virulence factors allowing the bacteria to kill the host. However, how the bacteria are able to disrupt the host intestinal barrier during the early stages of infection remains unknown. Here we show that Bt employs the nematicidal Cry toxins and additional virulence factors controlled by the PlcR regulon to disrupt the intestinal epithelial junctions of C. elegans at the early stages of infection allowing that Bt bacteria complete its life cycle in the worms. Our work provides new insights into the pathogenesis of Bt, and highlights the importance of breaking down host epithelial junctions for a successful infection, a similar mechanism could be used by other pathogens-host interactions since epithelial junctions are conserved structures from insects to mammals.

microbiology↗

bioRxiv2018

Epitope-based vaccine design yields fusion peptide-directed antibodies that neutralize diverse strains of HIV-1

A central goal of HIV-1-vaccine research is the elicitation of antibodies capable of neutralizing diverse primary isolates of HIV-1. Here we show that focusing the immune response to exposed N-terminal residues of the fusion peptide, a critical component of the viral entry machinery and the epitope of antibodies elicited by HIV-1 infection, through immunization with fusion peptide-coupled carriers and prefusion-stabilized envelope trimers, induces cross-clade neutralizing responses. In mice, these immunogens elicited monoclonal antibodies capable of neutralizing up to 31% of a cross-clade panel of 208 HIV-1 strains. Crystal and cryo-electron microscopy structures of these antibodies revealed fusion peptide-conformational diversity as a molecular explanation for the cross-clade neutralization. Immunization of guinea pigs and rhesus macaques induced similarly broad fusion peptide-directed neutralizing responses suggesting translatability. The N terminus of the HIV-1-fusion peptide is thus a promising target of vaccine efforts aimed at eliciting broadly neutralizing antibodies.

immunology↗