Biology subjects

Xu, Y.

Publications and source records attributed to Xu, Y..

At least 469 records · Page 26Linked to original sources

bioRxiv2024

Large-scale multi-omics analyses identified root-microbiome associations underlying plant nitrogen nutrition

The microbiome determines the performance and fitness of the host plant. Nevertheless, the causal interaction between host genetic variation, gene regulation and the impact of the microbiome on the host phenotype remain unknown. Here we generated 1,341 paired root transcriptome, rhizosphere microbiome and root ionome samples and performed a multi-omics analyses of the host-microbe association at the root-soil interface using 175 rapeseeds (Brassica napus L.) resequenced ecotypes at two field environments. We observed the highest statistically explained variance for root nitrogen uptake among natural ionomic variation by overall transcriptome-wide gene expression and microbial abundance variation. Moreover, we identified significant genome-wide associations for 203 highly heritable amplicon sequence variants (ASVs) at multiple genetic loci regulated by eQTL hotspots associated with nitrogen metabolism components. These associations involved a central bacterial genus (Sphingopyxis), which plays a dominant role on gene regulatory effect on its variation regulated by eQTL hotspots. In addition, we performed high-throughput bacterial cultivation from rapeseed roots and subjected Sphingopyxis to whole genome sequencing. Finally, targeted metabolite profiling and confocal imaging assays demonstrated a host-microbiome regulatory effect on Sphingopyxis established by lateral root development and plant nitrogen nutrition. In summary, our integrative approach reveals the genetic basis of host-microbiome trait associations in the transcriptional, nutritional and environmental domains and suggests that the microbiome might have causal effects on root development with implications towards the breeding of nutrient-efficient crops.

plant biology↗

bioRxiv2024

SPIRED-Fitness: an end-to-end framework for the prediction of protein structure and fitness from single sequence

AO_SCPLOWBSTRACTC_SCPLOWSignificant research progress has been made in the field of protein structure and fitness prediction. Particularly, single-sequence-based structure prediction methods like ESMFold and OmegaFold achieve a balance between inference speed and prediction accuracy, showing promise for many downstream prediction tasks. Here, we propose SPIRED, a novel single-sequence-based structure prediction model that exhibits comparable performance to the state-of-the-art methods but with approximately 5-fold acceleration in inference and at least one order of magnitude reduction in training consumption. By integrating SPIRED with downstream neural networks, we compose an end-to-end framework named SPIRED-Fitness for the rapid prediction of both protein structure and fitness from single sequence. SPIRED-Fitness and its derivative SPIRED-Stab achieve state-of-the-art performance in predicting the mutational effects on protein fitness and stability metrics, respectively.

bioinformatics↗

bioRxiv2024

Scalable single-cell pooled CRISPR screens with conventional knockout vector libraries

Current methods for single-cell RNA profiling of pooled CRISPR screens are limited, either by indirect capture of single guide RNAs (sgRNAs) or by custom modification of plasmid libraries. Here, we present a direct sgRNA capture platform called Native sgRNA Capture and sequencing (NSC-seq) that enables single-cell CRISPR screens using common knockout plasmid libraries, facilitating genotype-phenotype mapping at multiple scales in vitro and in vivo. Additionally, we characterize sgRNA expression in three whole-genome knockout libraries, revealing a substantial subset of truncated (isoform) spacer reads. We provide this dataset as a reference of expressed sgRNA isoforms that may potentially have compromised CRISPR gene editing efficacy and precision.

genomics↗

bioRxiv2024

Systematic characterization of protein structural features of alternative splicing isoforms using AlphaFold 2

Alternative splicing is an important cellular process in eukaryotes, altering pre-mRNA to yield multiple protein isoforms from a single gene. However, our understanding of the impact of alternative splicing events on protein structures is currently constrained by a lack of sufficient protein structural data. To address this limitation, we employed AlphaFold 2, a cutting-edge protein structure prediction tool, to conduct a comprehensive analysis of alternative splicing for approximately 3,000 human genes, providing valuable insights into its impact on the protein structural. Our investigation employed state of the art high-performance computing infrastructure to systematically characterize structural features in alternatively spliced regions and identified changes in protein structure following alternative splicing events. Notably, we found that alternative splicing tends to alter the structure of residues primarily located in coils and beta-sheets. Our research highlighted a significant enrichment of loops and highly exposed residues within human alternatively spliced regions. Specifically, our examination of the Septin-9 protein revealed potential associations between loops and alternative splicing, providing insights into its evolutionary role. Furthermore, our analysis uncovered two missense mutations in the Tau protein that could influence alternative splicing, potentially contributing to the pathogenesis of Alzheimers disease. In summary, our work, through a thorough statistical analysis of extensive protein structural data, sheds new light on the intricate relationship between alternative splicing, evolution, and human disease.

bioinformatics↗

bioRxiv2024

Rubisco supplies pyruvate for the 2-C-methyl-D-erythritol-4-phosphate pathway in Arabidopsis

Ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) produces pyruvate in the chloroplast through beta-elimination of the aci-carbanion intermediate. Here we show that this side reaction supplies pyruvate for isoprenoid, fatty acid, and branched chain amino acid biosynthesis in photosynthetically active tissue. 13C labeling studies of whole Arabidopsis plants demonstrate that the total carbon commitment to pyruvate is too large for phosphoenolpyruvate (PEP) to serve as precursor. Low oxygen stimulates rubisco carboxylase activity and increased pyruvate production and flux through the 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway, which supplies the precursors for plastidic isoprenoid biosynthesis. Metabolome analysis of mutants defective in PEP or pyruvate import further supported rubisco as the main source of pyruvate in chloroplasts. Rubisco beta-elimination leading to pyruvate constituted 0.7% of the product profile in in vitro assays, which translates to 2% of the total carbon leaving the Calvin-Benson-Bassham cycle (CBC). These insights solve the so-called pyruvate paradox, improve the fit of metabolic models for central metabolism, and connect the MEP pathway directly to carbon assimilation.

plant biology↗

bioRxiv2024

Using split protein reassembly strategy to optically control PLD enzymatic activity

Phospholipase D (PLD) and phosphatidic acid (PA) play a spatio-temporal role in regulating diverse cellular activities. Although current methodologies enable optical control of the subcellular localization of PLD and by which influence local PLD enzyme activity, the overexpression of PLD elevates the basal PLD enzyme activity and further leads to increased PA levels in cells. In this study, we employed a split protein reassembly strategy and optogenetic techniques to modify superPLD (developed by Jeremy Baskin group). We splited this variants into two HKD domains and fused these domains with optogenetic and chemogenetic elements and by which we achieved control of the two HKD interaction and then restored the PLD enzymatic activity.

cell biology↗

bioRxiv2024

LncRNA GAS6-AS1 contributes to 5-fluorouracil resistance in colorectal cancer

5-Fluorouracil (5-FU) resistance has always been a formidable obstacle in the adjuvant treatment of advanced colorectal cancer (CRC). In recent years, long non-coding RNAs have emerged as key regulators in various pathophysiological processes including 5-FU resistance. Here, RNA-seq combined with weighted gene correlation network analysis confirmed the close association of GAS6-AS1 with TRG grades. GAS6-AS1 expression was positively correlated with advanced clinicopathological features and poor prognosis in CRC. GAS6-AS1 increased the 50% inhibiting concentration of 5-FU, enhanced cell proliferation, and accelerated G1/S transition in CRC cells, both with and without 5-FU, both in vitro and in vivo. Mechanistically, GAS6-AS1 enhanced the stability of MCM3 mRNA by recruiting PCBP1, consequently increasing MCM3 expression. Furthermore, PCBP1 and MCM3 counteracted the effects of GAS6-AS1 on 5-FU resistance. Notably, the PDX model indicated that combining chemotherapeutic drugs with GAS6-AS1 knockdown yielded superior outcomes in vivo. Together, our findings elucidate that GAS6-AS1 directly binds to PCBP1, enhancing MCM3 expression and thereby promoting 5-FU resistance. GAS6-AS1 may serve as a robust biomarker and potential therapeutic target for combination therapy in CRC.

molecular biology↗

bioRxiv2024

Phylogenetically and structurally diverse reductive dehalogenases link biogeochemical cycles in deep-sea cold seeps

Reductive dehalogenation is crucial for halogen cycling and environmental remediation, yet its ecological role is not completely understood, especially in deep-sea environments. To address this gap, we investigated the diversity of reductive dehalogenases (RDases) and ecophysiology of organohalide reducers in deep-sea cold seeps, which are environments rich in halogenated compounds. Through genome- resolved metagenomic analysis of 165 global cold seep sediment samples, we identified four types of RDases, namely prototypical respiratory, transmembrane respiratory, and cytosolic RDases, and one novel clade. Cold seeps were found to harbor a higher abundance of these RDase genes compared to other marine sediments, establishing them as unique hotspots for microbial reductive dehalogenation activity. These RDases are encoded by a wide range of microorganisms across four archaeal and 36 bacterial phyla, significantly expanding the known diversity of organohalide reducers. Halogen geochemistry, laboratory incubations with various halogenated compounds, metatranscriptomic data, and metabolomic profiling confirmed the presence of organohalides at concentrations of up to 18 mg/g in these sediments and demonstrated their active microbial reduction. This process is tightly linked to other biogeochemical cycles, including carbon, hydrogen, nitrogen, sulfur, and trace elements. RDases from cold seeps have diverse N-terminal structures across different gene groups, and reductive dehalogenase genes in these environments are mostly functionally constrained and conserved. These findings collectively suggest that reductive dehalogenation is a central process in deep-sea environments, mediated by a diverse array of microbes and novel enzymes.

microbiology↗

bioRxiv2024

Multi-state kinetics of the syringe-like injection mechanism of Tc toxins

Tc toxins are virulence factors of many insects and human pathogenic bacteria. They attach as soluble prepores to receptors on host cells and following acidification in the late endosome, perforate the cell membrane like a syringe to translocate toxic enzymes into the host cell through their pore-forming channel. Although this complex transformation has been structurally well studied, the functional aspects of this large-scale rearrangement, such as the reaction pathway with possible intermediate states and the resulting temporal evolution have remained elusive. Here, we used an integrated biophysical approach to monitor the prepore-to-pore transition and found that it takes [~]28 h when induced by high pH in the absence of other factors. In the presence of liposomes, an increasingly high pH or receptors, such as heparin or Vsg, the probability to transform prepores to pores increases by a factor of up to 4. This effect can also be mimicked by biotinylation or site-directed mutagenesis of the shell, demonstrating that shell destabilization is a crucial step in prepore-to-pore transition. We show that shell opening is a heterogeneous process with transition times ranging from 60 ms to 1.6 s and resolve three sequential intermediate states: an initial transient intermediate during shell destabilization, a first stable intermediate where the receptor-binding domains on the shell rearrange and a second stable intermediate with an open shell. In contrast, the ejection of the pore-forming channel from the open shell is highly cooperative with a transition time of < 60 ms. This detailed knowledge of the Tc toxin mechanism of action, even in the absence of receptors, is important for the future application of Tc toxins as biomedical devices or biopesticides.

biochemistry↗

bioRxiv2024

Unveiling the regulatory role of GRP7 in ABA signal-mediated mRNA translation efficiency regulation

Abscisic acid (ABA) is a crucial phytohormone involved in plant growth and stress responses. Although ABA has been implicated in the regulation of translation efficiency in Arabidopsis thaliana, the underlying mechanism remains largely unknown. In this study, we discovered that ABA treatment modulates globally translation efficiency (TE) by affecting pre-rRNA processing in the nucleolus and ribosome distribution status in the cytoplasm. The regulation of TE by ABA was largely abolished in mutants of ABA signaling core components, such as receptors PYRABACTIN RESISTANCE1/PYRABACTIN-LIKE/REGULATORY COMPONENTS OF ABA RECEPTORS (PYR/PYL/RACRs), the protein phosphatase 2Cs (PP2Cs), and the SNF1-related protein kinase 2s (SnRK2s). ABA treatment reduced the protein levels of glycine-rich RNA binding protein 7 (GRP7) in the signaling core components-dependent manner. Ribo-seq and CLIP-seq analyses unveiled GRP7s role in governing the TE of a substantial proportion of ABA-regulated genes, although independent of directly binding to the respective mRNAs. Furthermore, GRP7 directly bound to pre-rRNA and interacted with Ribosomal protein S6 (RPS6A), RPS14A and RPL36aA in the nucleolus to regulate rRNA processing. Additionally, GRP7 associated with mature polysome in the cytoplasm and is hypersensitive to translation inhibitor anisomycin when loss of function. Collectively, our study unveils the role of GRP7 in mediating translation regulation in ABA signaling, providing a novel regulatory model for plants to response to environmental stresses.

plant biology↗

bioRxiv2024

Evaluation of the effects of ripening-related genes on fruit metabolites and the associated regulatory mechanisms in tomato

Fruit ripening, which is a complex process involving dynamic changes to metabolites, is controlled by multiple factors, including transcription factors (TFs). Several TFs are reportedly essential regulators of tomato fruit ripening. To evaluate the effects of specific TFs on metabolite accumulation during fruit ripening, CRISPR/Cas9-mediated mutagenesis was combined with metabolome and transcriptome analyses to explore regulatory mechanisms. Specifically, we generated various genetically engineered tomato lines that differed regarding metabolite contents and fruit colors. The metabolite and transcript profiles indicated that the selected TFs have distinct functions that control fruit metabolite contents, especially carotenoids and sugars. Our findings may provide new insights into the regulatory mechanisms governing tomato fruit ripening. Moreover, a mutation to ELONGATED HYPOCOTYL5 (HY5) increased the tomato fruit fructose and glucose contents by approximately 20% (relative to the wild-type levels). Our in vitro assay showed that HY5 can bind directly to the G-box cis-element in the SWEET12c promoter to activate expression, thereby modulating sugar transport. Our findings have clarified the mechanism regulating fruit metabolic networks, while also providing the theoretical basis for breeding horticultural crops that produce fruit with diverse flavors and colors.g

plant biology↗

bioRxiv2024

Hydrophobic bulk conservation and genetic code determine the pattern of prokaryotic genome organization

Prokaryotic genomes in general exhibit marked organizational asymmetry, and differ substantially in GC-skew, AT-skew, and GS-bias (gene strand bias). Despite enigmatic origins, these organizational features and genomic GC-content have been described to be widely associated with each other recently, providing an opportunity to probe the evolutionary mechanisms. By analyzing sequence data of 4,012 leading strands from all available representative genomes, we found that the unusual nucleotide usage of coding genes under low GC-content contributed to a shift between two organizational patterns in the context of mutational biases. Analysis of artificially established neutral and natural models further suggests that in genes both GC-skew and AT-skew increase with decreasing GC-content, mostly contributed by synonymous substitutions because of genetic code, and by nonsynonymous substitutions because of hydrophobic bulk conservation in amino acid usage, respectively. This novel mechanistic framework in our study highlights the importance of evolutionary processes "operating at lower levels of organization" in the microbial world.

evolutionary biology↗

bioRxiv2024

Heparan sulfate selectively inhibits the collagenase activity of cathepsin K

Cathepsin K (CtsK) is a cysteine protease with potent collagenase activity. CtsK is highly expressed by bone-resorbing osteoclasts and plays an essential role in bone remodeling. Although CtsK is known to bind heparan sulfate (HS), the structural details of the interaction, and how HS ultimately regulates the biological functions of CtsK, remains largely unknown. In this report, we determined that CtsK preferably binds to larger HS oligosaccharides, such as dodecasaccharides (12mer), and that the12mer can induce monomeric CtsK to form a stable dimer in solution. Interestingly, while HS has no effect on the peptidase activity of CtsK, it greatly inhibits the collagenase activity of CtsK in a manner dependent on sulfation level. By forming a complex with CtsK, HS was able to preserve the full peptidase activity of CtsK for prolonged periods, likely by stabilizing its active conformation. Crystal structures of Ctsk with a bound 12mer, alone and in the presence of the endogenous inhibitor cystatin-C reveal the location of HS binding is remote from the active site. Mutagenesis based on these complex structures identified 6 basic residues of Ctsk that play essential roles in mediating HS-binding. At last, we show that HS 12mers can effectively block osteoclast resorption of bone in vitro. Combined, we have shown that HS can function as a multifaceted regulator of CtsK and that HS-based oligosaccharide might be explored as a new class of selective CtsK inhibitor in many diseases that involve exaggerated bone resorption.

biochemistry↗

bioRxiv2024

Fast and lightweight cell atlas approximations across organs and organisms

Omic technologies at single-cell resolution are reshaping our understanding of cellular diversity. The generation of cell atlases that capture the cellular composition of an entire individual is progressing rapidly. However, the science of organising and extracting information from these atlases is still in its infancy and for many biomedical researchers atlas exploration remains challenging. Here, we leveraged extensive experience in single-cell data analytics to pinpoint three major accessibility barriers to cell atlases, related to (i) programming skill or language, (ii) scalability, and (iii) dissemination standards. To help researchers overcome these barriers, we developed cell atlas approximations, a computational approach enabling the analysis of cell atlases across organs and organisms without programming skills, rapidly, and at scale. The web interface at https://atlasapprox.org facilitates the exploration of cell atlases in 19 species across the tree of life through a chatbot driven by frontend natural language processing. In parallel, application programming interfaces streamline data access for computational researchers and include specialised packages for Python, R, JavaScript, and Bash. Supported queries include marker gene identification, cross-organ comparisons, cell embeddings, gene sequences, searches for similar features, and bidirectional zoom between cell types and cell states. Most queries are answered in less than 1.5 seconds thanks to lossy data compression algorithms based on cell annotations and similarity graphs. Compared to traditional cell atlas analysis, this approach can reduce data size by more than 100 times and accelerate workflows by up to 100,000 times. Atlas approximations aim to make the exploration of cell atlases accessible to anyone in the world.

cell biology↗

bioRxiv2023

Node Features of Chromosome Structure Network and Their Connections to Genome Annotation

The 3D conformations of chromosomes can encode biological significance, and its implication is being increasingly appreciated recently. Certain chromosome structural features, such as A/B compartmentalization, are frequently extracted from pairwise contact information (physical association between different regions of the genome) and compared with linear annotations of the genome, such as histone modifications and lamina association. Here, we investigate how additional properties of chromosome structure can be deduced using the abstract graph representation of the contact heatmap, and how network properties can have a better connection with some of these biological annotations. We constructed chromosome structure networks (CSNs) from bulk Hi-C data and calculated a set of site-resolved (node-based) network properties of these CSNs. We found these network properties are useful for characterizing chromosome structure features. We examined the ability of network properties in differentiating several scenarios, such as haploid vs diploid cells, partially inverted nuclei vs conventional architecture, and structural changes during cell development. We also examined the connection between network properties and a series of other linear annotations, such as histone modifications and chromatin states including poised promoter and enhancer labels. We found that semi-local network properties are more capable of characterizing genome annotations than diffusive or ultra-local node features. For example, local square clustering coefficient can be a strong classifier of lamina-associated domains (LADs), whereas a path-based network property, closeness centrality, does not vary concordantly with LAD status. We demonstrated that network properties can be useful for discerning large-scale chromosome structures that emerge in different biological situations. TOC Figure O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=135 SRC="FIGDIR/small/573476v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@45df0forg.highwire.dtl.DTLVardef@13776daorg.highwire.dtl.DTLVardef@5c1519org.highwire.dtl.DTLVardef@16d447f_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2023

CentIER: accurate centromere identification for plant genomes with sequence specificity information

Centromere identification is one of the important problems in genomics, providing a foundation for the studies of centromeres in aspects of composition, functionality, evolution, inheritance, and variation. The existing wet-experiment-based method is costly and time-consuming, while the bioinformatic method can only detect tandem repeats losing non-repetitive sequence regions in the centromere. To address these shortcomings, we introduce a new pipeline, CentIER, for the automatic and accurate identification and annotation of centromere regions by taking advantage of the sequence specificity information. CentIER only requires users to input the genomic sequence, and then it can partition the centromeric region from a chromosome, identify tandem repeat monomers, annotate retrotransposons, and ultimately output visualized results. By referencing the experimentally determined centromere regions, it was discovered that the predictive accuracy of centromere recognition by CentIER exceeded 90%. Following the evaluation of CentIERs accuracy, it was applied to investigate the sequence and distribution characteristics of centromeric retrotransposons and tandem repeat sequences of different species, providing insights into these traits in monocotyledonous and dicotyledonous plants.

bioinformatics↗

bioRxiv2023

Single-cell and spatial multiomic inference of gene regulatory networks using SCRIPro

The accurate reconstruction of gene regulation networks (GRNs) from sparse and noisy single-cell or spatial multi-omics data remains a challenge. Here, we present SCRIPro, a comprehensive computational framework that robustly infers GRNs for both single-cell and spatial multi-omics data. SCRIPro first addresses sample sparseness by a density clustering approach. SCRIPro assesses transcriptional regulator (TR) importance through chromatin reconstruction and in silico deletion, referencing 1,292 human and 994 mouse TRs. It combines TR-target importance scores with expression levels for precise GRN reconstruction. Finally, we benchmarked SCRIPro on diverse datasets, it outperforms existing motif-based methods and accurately reconstructs cell type-specific, stage-specific, and region-specific GRNs.

bioinformatics↗

bioRxiv2023

DNA nicks in both leading and lagging strand templates can trigger break-induced replication

Encounters between replication forks and unrepaired single-strand DNA breaks (SSBs) generate single-ended double-strand breaks (seDSBs) that can later become double-ended (deDSBs) through fork convergence. seDSBs can be repaired by break-induced replication (BIR), which is a highly mutagenic pathway that is thought to be repsonsible for many of the mutations and genome rearrangements that drive cancer development. However, the frequency of BIRs deployment and its ability to be triggered by both leading and lagging template strand SSBs was unclear. Using site- and strand-specific SSBs generated by nicking enzymes, including CRISPR-Cas9n, we demonstrate that leading and lagging template strand SSBs in fission yeast are typically converted into deDSBs that are repaired primarily by error-free homologous recombination. However, both types of SSB can also trigger BIR, and the frequency of these events increases when the converging fork is delayed and the non-homologous end joining protein Ku70 is deleted.

molecular biology↗