Biology subjects

Du, R.

Publications and source records attributed to Du, R..

At least 19 recordsLinked to original sources

bioRxiv2026

Synthetic reaction-diffusion patterning in mammalian cells

During development, periodic patterns arise through reaction-diffusion (RD) circuits involving local activation and long-range inhibition (LALI). Natural systems are complex, typically entangling regulation and transport in the same morphogens. It remains unclear what minimal circuits are sufficient for LALI patterning in mammalian cells, and whether they can be implemented in a modular fashion, allowing independent control of regulation and transport. Here, we designed synthetic LALI circuits using three natural and synthetic signaling systems, BMP, Activin, and the SNIPR pathway, and showed that they generate spatial patterns with distinct characteristic spatial scales. Model-guided design allowed flexible morphogen reassignment, with BMP4 acting as either short-range activator or long-range inhibitor. Gene expression noise degraded long-range order but preserved the dominant local spatial scale. Together, these results provide a modular, extensible synthetic framework for engineered patterning in mammalian cells.

synthetic biology↗

bioRxiv2026

Multiomic analysis of non-glaucomatous human trabecular meshwork cells.

Glaucoma is an irreversible blinding disease that affects millions of individuals worldwide. Elevated intraocular pressure (IOP), regulated by the trabecular meshwork (TM) in the anterior eye, is the only modifiable risk factor. Human TM cells can be cultured from donor eyes, providing a precious resource for studying factors that induce or prevent glaucoma. The goal of this study was to produce datasets that define the molecular profile of human non-glaucomatous TM cells. Using 18 human TM cell strains cultured from non-glaucomatous individuals deposited from seven laboratories in the USA and UK, this study used transcriptomic, proteomic, lipidomic, and metabolomic analyses to characterize the molecular content of TM cells. The data herein provides the most comprehensive multiomic analyses of human TM cells to date and will be a useful resource for researchers and clinicians in the TM and glaucoma fields.

cell biology↗

bioRxiv2026

A global metagenomic atlas uncovers ubiquitous biosynthetic potential linked to adaptation in extreme environments

Extreme environments impose severe physicochemical stresses that drive microorganisms to evolve specialized survival strategies. Microbial secondary metabolites determined by biosynthetic gene clusters (BGCs) are recognized as important mediators of microbial adaptation to environmental stress. However, their ecological roles, particularly habitat-dependent preferences across different environments, remain poorly understood. Although extreme environments provide opportunities to mine microbiomes for unique adaptations, such research is hampered by a lack of systematic overview of its genomic diversity, BGC diversity, and the relationships between them. Here, we constructed a standardized extremophilic genomic catalogue (SEGC) from 1,462 metagenomic samples spanning seven representative extreme habitats. The catalogue comprised 54,661 metagenome-assembled genomes representing 21,805 species, 66.1% of which were previously uncharacterized. With this catalogue, we identified 162,855 BGCs distributed across 81.5% of MAGs. Gene cluster family analysis showed the strong habitat dependence largely explained by species-level habitat specificity. Terpene biosynthetic pathways illustrated habitat-linked adaptive strategies, with hopan-22-ol biosynthesis enriched in acid mine, deep sea and hydrothermal plume environments, while retinal-based phototrophy predominated in cryosphere and saline-alkaline habitats. Metatranscriptomic analyses supported in situ activity of these pathways. In conclusion, we presented a global atlas of biosynthetic potential across extreme-environment microbiota and revealed habitat-dependent patterns of secondary metabolism linked to microbial survival.

microbiology↗

bioRxiv2025

Breakthrough infection elicits synergistic broadly neutralizing antibodies targeting non-dominant epitopes on RBD and NTD in an individual of inactivated SARS-CoV-2 vaccine

Background/ObjectivesIdentification and characterization of broadly neutralizing monoclonal antibodies from individuals exposed to SARS-CoV-2, either by infection or vaccination, can inform the development of next-generation vaccines and antibody therapeutics with pan-SARS-CoV-2 protection. MethodsThrough single B cell sorting and RT-PCR, monoclonal antibodies (mAbs) were isolated from a donor who experienced BA.5 or BF.7 breakthrough infection after three doses of inactivated vaccines. Their binding and neutralizing capacities were respectively measured with ELISA and pseudovirus-based neutralization assay. Their epitopes were mapped by competition ELISA and site- directed mutation. ResultsAmong a total of 67 spike-specific mAbs cloned from the donor, four mAbs (KXD643, KXD652, KXD681, and KXD686) can neutralize all tested SARS-CoV-2 variants from wild-type to KP.3. Moreover, KXD643, KXD652, and KXD681 belong to a clonotype encoded by IGHV5-51 and IGKV1-13, and recognize the cryptic and conserved RBD-8 epitope on the receptor-binding domain (RBD). In contrast, KXD686 is encoded by IGHV1-69 and IGKV3-20, and targets a conserved epitope (NTD Site iv) outside the antigenic supersite (NTD Site i) of the N-terminal domain (NTD). Notably, antibody cocktails containing these two groups of mAbs can neutralize SARS-CoV-2 more potently due to synergistic effects. In addition, bispecific antibodies derived from KXD643 and KXD686 demonstrate further improved neutralizing potency compared to antibody cocktails. ConclusionsThese four mAbs can be developed as candidates of pan-SARS-CoV-2 antibody therapeutics through further antibody engineering. On the other hand, vaccines designed to simultaneously elicit neutralizing antibodies towards RBD-8 and NTD Site iv have the potential to provide pan-SARS-CoV-2 protection.

immunology↗

bioRxiv2025

High-resolution NMR spectroscopy of proteins in intact mitochondria

Understanding protein structure and function within mitochondria is essential for unraveling the molecular mechanisms underlying cellular energy production, stress response, and disease. Here, we present a novel approach for NMR observation of proteins within intact mitochondria by delivering proteins directly into isolated mitochondria via electroporation. Using this method, we investigate the interaction of -synuclein (-syn) with the mitochondria membrane and examine how post-translational modifications regulate this interaction. Additionally, we assessed the stability of GB1 and the dimerization of its variant within mitochondria, achieving quantitative insights into mitochondrial environmental impact on protein function. This approach offers a valuable framework for exploring mitochondrial related biomolecular events at atomic-resolution within intact mitochondria, paving the way for a more comprehensive understanding of the molecular events governing mitochondrial health and dysfunction.

biochemistry↗

bioRxiv2024

Longitudinal profiling of B cells primed by mRNA vaccine and recalled by Omicron variants uncovers antibodies broadly neutralizing sarbecoviruses

A key question on ancestral SARS-CoV-2 immune imprinting is to what extent imprinted B cells can develop neutralizing breadth and potency in immune recalls. Here, we longitudinally tracked B cells recognizing wild-type spike in two individuals, who were sequentially infected by Omicron variants after receiving mRNA vaccines. Functional and genetic analysis of 632 monoclonal antibodies (mAbs) from those B cells reveals that mAbs cloned after second infection have dramatically enhanced neutralizing breadth and potency, which is attributed to recall and maturation of pre-existing memory B cells. Among the 11 mAbs that potently neutralize SARS-CoV-2 variants from wild-type to KP.3, 5 mAbs are classified into public clonotypes encoded by IGHV3-53 or IGHV3-66, whereas the rest belong to a rarely reported clonotype encoded by IGHV3-74. Notably, IGHV3-74 mAbs can also broadly neutralize other sarbecoviruses by targeting a novel epitope on receptor-binding domain of spike. These results support that ancestral SARS-CoV-2 immune imprinting can be harnessed in developing pan-SARS-CoV-2 and even pan-sarbecovirus vaccines. SummaryChen et al. demonstrate that B cells imprinted by ancestral SARS-CoV-2 have tremendous potential to develop neutralizing breadth and potency in repeated immune recalls driven by Omicron variants, implicating that ancestral SARS-CoV-2 immune imprinting can be harnessed in developing pan-SARS-CoV-2 and even pan-sarbecovirus vaccines.

immunology↗

bioRxiv2024

Chemical Space Localization for Unknown Metabolite Annotation via Semantic Similarity of Mass Spectral Language

Untargeted analysis using liquid chromatography{square}mass spectrometry (LC-MS) allows quantification of known and unknown compounds within biological systems. However, in practical analysis of complex biological system, the majority of compounds often remain unidentified. Here, we developed a novel deep learning-based compound annotation approach via semantic similarity analysis of mass spectral language. This approach enables the prediction of structurally related compounds for unknowns. By considering the chemical space, these structurally related compounds provide valuable information about the potential location of the unknown compounds and assist in ranking candidates obtained from molecular structure databases. Validated with two independent benchmark datasets obtained by chemical standards, our method has consistently demonstrated superior performance compared to existing compound annotation methods. A case study of the tomato ripening process indicates that DeepMASS has significant potential for metabolic biomarker identification in real biological systems. Overall, the presented method shows considerable promise in annotating metabolites, particularly in revealing the "dark matter" in untargeted analysis.

bioinformatics↗

bioRxiv2024

Ancestral SARS-CoV-2 immune imprinting persists on RBD but not NTD after sequential Omicron infections

Whether Omicron exposures could overcome ancestral SARS-CoV-2 immune imprinting remains controversial. Here we analyzed B cell responses evoked by sequential Omicron infections in vaccinated and unvaccinated individuals. Plasma neutralizing antibody titers against ancestral SARS-CoV-2 and variants indicate that immune imprinting is not consistently induced by inactivated or recombinant protein vaccines. However, once induced, immune imprinting is not countered by successive Omicron challenges. We compared binding specificities, neutralizing capacities, developing origins and targeting epitopes of monoclonal antibodies from individuals with or without immune imprinting. Although receptor-binding domain (RBD) and N-terminal domain (NTD) of spike are both primary targets for neutralizing antibodies, immune imprinting only shapes antibody responses to RBD by impeding the production of Omicron-specific neutralizing antibodies while facilitating the development of broadly neutralizing antibodies. We propose that immune imprinting can be either neglected by NTD-based vaccines to induce variant-specific antibodies or leveraged by RBD-based vaccines to induce broadly neutralizing antibodies.

immunology↗

bioRxiv2024

Synthetic dosage-compensating miRNA circuits for quantitative gene therapy

A longstanding challenge in gene therapy is expressing a dosage-sensitive gene within a tight therapeutic window. For example, loss of MECP2 function causes Rett syndrome, while its duplication causes MECP2 duplication syndrome. Viral gene delivery methods generate variable numbers of gene copies in individual cells, creating a need for gene dosage-invariant expression systems. Here, we introduce a compact miRNA-based, incoherent feed-forward loop circuit that achieves precise control of Mecp2 expression in cells and brains, and improves outcomes in an AAV-based mouse model of Rett syndrome gene therapy. Single molecule analysis of endogenous and ectopic Mecp2 mRNA revealed precise, sustained expression across a broad range of gene dosages. Delivered systemically in a brain-targeting AAV capsid, the circuit strongly suppressed Rett behavioral symptoms for over 24 weeks, outperforming an unregulated gene therapy. These results demonstrate that synthetic miRNA-based regulatory circuits can enable precise in vivo expression to improve the safety and efficacy of gene therapy. One sentence description: A synthetic miRNA-based incoherent feed-forward loop circuit embedded in a gene delivery vector overcomes the challenge of overexpression toxicity in a mouse model of Rett syndrome gene therapy.

synthetic biology↗

bioRxiv2024

miRNA circuit modules for precise, tunable control of gene expression

Accurate control of transgene expression is important for research and therapy but challenging to achieve in most settings. miRNA-based regulatory circuits can be incorporated within transgenes for improved control. However, the design principles, performance limits, and applications of these circuits in research and biotechnology have not been systematically determined. Here, combining modeling and experiments, we introduce miRNA-based circuit modules, termed DIMMERs, that establish precise, tunable control of transgene expression across diverse cell types to facilitate imaging, editing, and gene therapy. The circuits use multivalent miRNA regulatory interactions to achieve nearly uniform, tunable, protein expression over two orders of magnitude variation in gene dosage. They function across diverse cell types, and can be multiplexed for independent regulation of multiple genes. DIMMERs reduce off-target CRISPR base editing, improve single-molecule imaging, and allow live tracking of AAV-delivered transgene expression in mouse cortical neurons. DIMMERs thus enable accurate regulation for research and biotechnology applications.

synthetic biology↗

bioRxiv2024

Contrasting Phyllosphere Mycobiome in two Lycopodiaceae Plant Species: Unraveling Potential HupA-Producing Fungi and Fungal Interactions

BackgroundHuperzine A (HupA) is a natural lycopodium alkaloid renowned for its efficacy in treating neurodegenerative diseases such as Alzheimers disease. It specifically occurs in the Huperzioideae subfamily of Lycopodiaceae. Fungi associated with Huperzioideae species are potential contributors to HupA biosynthesis, offering promising prospects for HupA production. However, limited knowledge of fungal diversity in lycophytes, coupled with decreased HupA production over time of fungal strains, has impeded the discovery and applications of HupA-producing fungi. Here, we investigated huperzine concentrations and the mycobiome across various tissues of two Lycopodiaceae species, Huperzia asiatica (a HupA producer) and Diphasiastrum complanatum (a non-HupA producer). Our objectives across the tissues of the two species are to unveil the distribution of potential HupA-producing fungi and elucidate fungal interactions within the mycobiome, aiming to uncover the role of HupA-producing fungi and pinpoint their potential fungal facilitators. ResultsAmong the tissues, H. asiatica exhibited the highest HupA concentration in apical shoots (360.27 g/ml) whereas D. complanatum showed no HupA presence in any tissue. We obtained 441 Amplicon Sequence Variants (ASVs) from H. asiatica and 497 ASVs from D. complanatum. The fungal communities in bulbils and apical shoots of H. asiatica were low in diversity and dominated by Sordariomycetes, a fungal class harboring the majority of reported HupA-producing fungi. Integrating bioinformatics with published experimental reports, we identified 27 potential HupA-producing fungal ASVs, primarily in H. asiatica, with 12 ASVs identified as hubs in the fungal interaction network, underscoring their pivotal roles in mycobiome stability. Members of certain fungal genera, such as Penicillium, Trichoderma, Dioszegia, Exobasidium, Lycoperdon and Cladosporium exhibited strong connections with the potential HupA producers in H. asiaticas network rather than in D. complanatums, implying their prospects as fungal facilitators in enhancing HupA production. ConclusionsThis study advances our knowledge of fungal diversity in Lycopodiaceae and provides insights into the search for potential HupA-producing fungi and fungal facilitators. It highlights the importance of exploring young tissues, and emphasizes the ecological interactions that promote the fungi-mediated production of complex bioactive compounds, offering new directions for research in fungal ecology and secondary metabolite production.

microbiology↗

bioRxiv2021

Eoulsan 2: an efficient workflow manager for reproducible bulk, long-read and single-cell transcriptomics analyses

AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSMotivationC_ST_ABSCore sequencing facilities produce huge amounts of sequencing data that need to be analysed with automated workflows to ensure reproducibility and traceability. Eoulsan is a versatile open-source workflow engine meeting the needs of core facilities, by automating the analysis of a large number of samples. Its core design separates the description of the workflow from the actual commands to be run. This originality simplifies its usage as the user does not need to handle code, while ensuring reproducibility. Eoulsan was initially developed for bulk RNA-seq data, but the transcriptomics applications have recently widened with the advent of long-read sequencing and single-cell technologies, calling for the development of new workflows. ResultWe present Eoulsan 2, a major update that (i) enhances the workflow manager itself, (ii) facilitates the development of new modules, and (iii) expands its applications to long reads RNA-seq (Oxford Nanopore Technologies) and scRNA-seq (Smart-seq2 and 10x Genomics). The workflow manager has been rewritten, with support for execution on a larger choice of computational infrastructure (workstations, Hadoop clusters, and various job schedulers for cluster usage). Eoulsan now facilitates the development of new modules, by reusing wrappers developed for the Galaxy platform, with support for container images (Docker or Singularity) packaging tools to execute. Finally, Eoulsan natively integrates novel modules for bulk RNA-seq, as well as others specifically designed for processing long read RNA-seq and scRNA-seq. Eoulsan 2 is distributed with ready-to-use workflows and companion tutorials. Availability and implementationEoulsan is implemented in Java, supported on Linux systems and distributed under the LGPL and CeCILL-C licenses at: http://outils.genomique.biologie.ens.fr/eoulsan/. The source code and sample workflows are available on GitHub: https://github.com/GenomicParisCentre/eoulsan. A GitHub repository for modules using the Galaxy tool XML syntax is further provided at: https://github.com/GenomicParisCentre/eoulsan-tools Contacteoulsan@bio.ens.psl.eu

bioinformatics↗

bioRxiv2021

The Taxus genome provides insights into paclitaxel biosynthesis

The ancient gymnosperm genus Taxus is the exclusive source of the anticancer drug paclitaxel, yet no reference genome sequences are available for comprehensively elucidating the paclitaxel biosynthesis pathway. We have completed a chromosome-level genome of Taxus chinensis var. mairei with a total length of 10.23 Gb. Taxus shared an ancestral whole-genome duplication with the coniferophyte lineage and underwent distinct transposon evolution. We discovered a unique physical and functional grouping of CYP725As (cytochrome P450) in the Taxus genome for paclitaxel biosynthesis. We also identified a gene cluster in the taxadiene biosynthesis, which was mainly formed by gene duplications. This study will facilitate the elucidation of paclitaxel biosynthesis and unleash the biotechnological potential of Taxus. One Sentence SummaryA chromosome-level genome assembly of Taxus chinensis var. mairei uncovers its unique genome evolution process and genetic architectures for the paclitaxel biosynthesis pathway.

genomics↗

bioRxiv2020

The jasmine (Jasminum sambac) genome and flower fragrances

Jasminum sambac, a world-renowned plant appreciated for its exceptional flower fragrance, is of cultural and economic importance. However, the genetic basis of its fragrance is largely unknown. Here, we present the first de novo genome of J. sambac with 550.12 Mb (scaffold N50 = 40.1 Mb) assembled into 13 pseudochromosomes. Terpene synthase genes associated with flower fragrance are significantly amplified in the form of gene clusters through tandem duplications in the genome. Eleven homolog genes within the SABATH super-family were identified as related to phenylpropanoid/benzenoid compounds. Several key genes regulating jasmonate biosynthesis were duplicated causing increased copy numbers. Furthermore, multi-omics analyses identified various aromatic compounds and the key genes involved in fragrance biosynthesis pathways. Our genome of J. sambac offers a basic genetic resource for studying floral scent biosynthesis and provides an essential foundation for functional genomic research and variety improvements in Jasminum.

plant biology↗

bioRxiv2020

Epitranscriptomic editing of the RNA N6-methyladenosine modification by dCasRx conjugated methyltransferase and demethylase

N6-methyladenosine (m6A) is the most common modification on endogenous RNA transcripts in mammalian cells. Currently, the lack of precise single-nucleotide RNA modification tools makes it challenging to understand the relationship between site-specific RNA methylation and the corresponding phenotypic outcomes. Here, we developed a bidirectional dCasRx epitranscriptome editing platform composed of a nucleus-localized dCasRx conjugated with either a methyltransferase, METTL3, or a demethylase, ALKBH5, to manipulate methylation events at targeted m6A sites in HEK293T and glioma stem cells. This platform specifically and efficiently edited m6A modifications at targeted sites, modulating both gene expression and cell proliferation. We then employed the dCasRx epitranscriptomic editor to further elucidate the molecular function of m6A-binding proteins YTH (DF1, DF2, DF3) family and found that the YTH paralogs act together to mediate RNA degradation. These findings collectively demonstrate that the dCasRx epitranscriptome perturbation platform reported in this study can be employed as site-specific m6A editors for delineating the functional roles of individual m6A modifications in the mammalian epitranscriptome.

molecular biology↗

bioRxiv2020

Loss of O-GlcNAcylation on MeCP2 Thr 203 Leads to Neurodevelopmental Disorders

Mutations of the X-linked methyl-CpG-binding protein 2 (MECP2) gene in humans are responsible for most cases of Rett syndrome (RTT), an X-linked progressive neurological disorder. While genome-wide screens in clinical trials reveal several putative RTT-associated mutations on MECP2, their causative relevance regarding the functional regulation of MeCP2 on the etiologic sites at the protein level require more evidence. In this study, we demonstrate that MeCP2 is dynamically modified by O-linked-{beta}-N-acetylglucosamine (O-GlcNAc) at threonine 203 (T203), an etiologic site in RTT patients. Disruption of the O-GlcNAcylation of MeCP2 specifically at T203 impairs dendrite development and spine maturation in cultured hippocampal neurons, and disrupts neuronal migration, dendritic spine morphogenesis and dysfunction of synaptic transmission in the developing and juvenile mouse cerebral cortex. Mechanistically, genetic disruption of O-GlcNAcylation at T203 on MeCP2 decreases neuronal activity-induced induction of Bdnf transcription. Our study highlights the critical role of MeCP2 T203 O-GlcNAcylation in neural development and synaptic transmission potentially via BDNF.

neuroscience↗

bioRxiv2020

Dropout in Neural Networks Simulates theParadoxical Effects of Deep Brain Stimulation onMemory

Neuromodulation techniques such as Deep Brain Stimulation (DBS) are a promising treatment for memory-related disorders including anxiety, addiction, and dementia. However, the outcome of these treatments appears to be paradoxical, as the use of these techniques can both disrupt and enhance memory even when applied to the same brain target. In this paper, we hypothesize that disruption and enhancement of memory through neuromodulation can be explained by the dropout of engram nodes. We used a convolutional neural network to classify handwritten digits and letters, applying dropout at different stages to simulate DBS effects on engrams. We showed that dropout applied during training improves the accuracy of prediction, whereas dropout applied during testing dramatically decreases accuracy of prediction, which mimics enhancement and disruption of memory, respectively. We further showed that transfer learning of neural networks with dropout had increased accuracy and rate of learning. Dropout during training provided a more robust "skeleton" network where transfer learning can be applied, mimicking the effects of chronic DBS on memory. Overall, we show that dropout of nodes can be a potential mechanism by which neuromodulation techniques such as DBS can both disrupt and enhance memory and provides a unique perspective on this paradox.

neuroscience↗

bioRxiv2019

Integrated Sequencing & Array Comparative Genomic Hybridization in Familial Parkinson’s Disease

BackgroundParkinsons disease (PD) is a genetically heterogeneous condition; both single nucleotide variants (SNVs) and copy number variants (CNVs) are important genetic risk factors. We examined the utility of combining exome sequencing and genome-wide array-based comparative genomic hybridization (aCGH) for identification of PD genetic risk factors. MethodsWe performed exome sequencing on 110 subjects with PD and a positive family history; 99 subjects were also evaluated using genome-wide aCGH. We interrogated exome sequencing and array comparative genomic hybridization data for pathogenic SNVs and CNVs at Mendelian PD gene loci. SNVs were confirmed via Sanger sequencing. CNVs were confirmed with custom-designed high-density aCGH, droplet digital PCR, and breakpoint sequencing. ResultsUsing exome sequencing, we discovered individuals with known pathogenic single nucleotide variants in GBA (p.E365K, p.T408M, p.N409S, p.L483P) and LRRK2 (p.R1441G and p.G2019S). Two subjects were each double heterozygotes for variants in GBA and LRRK2. Based on aCGH, we additionally discovered cases with an SNCA duplication and heterozygous intragenic GBA deletion. Five additional subjects harbored both SNVs (p.N52fs, p.T240M, p.P437L, p.W453*) and likely disrupting CNVs at the PARK2 locus, consistent with compound heterozygosity. In nearly all cases, breakpoint sequencing revealed microhomology, a mutational signature consistent with CNV formation due to DNA replication errors. ConclusionsIntegrated exome sequencing and aCGH yielded a genetic diagnosis in 19.3% of our familial PD cohort. Our analyses highlight potential mechanisms for SNCA and PARK2 CNV formation, uncover multilocus pathogenic variation, and identify novel SNVs and CNVs for further investigation as potential PD risk alleles.

genetics↗