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Diao, X.

Publications and source records attributed to Diao, X..

13 recordsLinked to original sources
bioRxiv2026

Cross-Species Comparison of Topologically Associating Domains (TADs) in Cereals Reveals Their Role in Genome Stability During Evolution

Topologically associating domains (TADs) are essential structural and functional modules of the genome that play a crucial role in regulating gene expression. In this study, we systematically investigated the conservation and evolution of TADs in five closely related crops, including maize, sorghum, coix, foxtail millet and broomcorn millet. Our results show that 74% of TAD boundaries are conserved between two inbred maize lines, B73 and Mo17, and that approximately 50% or more of TAD boundaries are conserved across different crop species. TAD number remains relatively stable in the face of changes in genome size. However, the length of TADs varies depending on genome size. Furthermore, we found that large-scale transposable element expansion leads to TAD expansion, while chromosomal inversions lead to TAD fusion and the formation of new TAD boundaries. Frequent chromatin interactions between subgenome chromosomes occur after whole-genome duplication. Moreover, we also found that crossovers are enriched at TAD boundaries in maize, indicating the importance of TADs as a fundamental unit during species evolution. Overall, our study provides insights into the conservation and evolution of TADs in crop genomes and their roles in genome organization and function.

plant biology↗
bioRxiv2025

Streptavidins play a multifunctional role within a biotin-pathway antibiotic network encoded in a biosynthetic supercluster

Streptavidin, the well-known biotin-binding protein extensively used in biotechnology, is naturally co-produced in Streptomyces avidinii alongside stravidins--inhibitors of biotin biosynthesis. Here, we uncover and activate a conserved genomic region flanked by two streptavidin genes, revealing multiple biosynthetic gene clusters that produce diverse biotin-related metabolites, including stravidins, acidomycin, -methyl-KAPA, -methyldesthiobiotin, and the novel non-proteinogenic amino acid ANDA. We show -methyldesthiobiotin arises from -methyl-KAPA, illustrating how methylated analogues interfere with distinct steps of biotin biosynthesis. Contrary to the view that streptavidin functions solely by sequestering biotin, our biochemical, structural, and bioactivity analyses demonstrate that it also binds acidomycin, neutralizing its antibacterial activity to protect the producer while likely facilitating compound secretion. The crystal structure of the streptavidin-acidomycin complex reveals the molecular basis for this dual functionality. Our findings establish a multifunctional streptavidin that integrates biotin sequestration and self-resistance to balance an antibiotic network targeting the biotin pathway in microbial competition.

microbiology↗
bioRxiv2025

Changing the initiation unit of nonribosomal peptide synthetases to access underexplored biosynthetic potential

Nonribosomal peptide synthases (NRPSs) are large multimodular enzymes, capable of synthesizing nonribosomal peptides (NRPs) with diverse structures and bioactivities. Genome sequencing revealed a large number of uncharacterized NRPS biosynthetic gene clusters (BGCs) and their products are underexplored. The majority of NRPSs remain silent potentially attributed to factors such as the low activity of the initiation unit or insufficient precursor supply. Exchanging the starter condensation (Cs) domains within initiation unit can change the length of acyl chains of NRPs, hinting at a promising strategy through swapping of a well-studied Cs domain to activate the initiation unit and harness primary metabolites as precursors, which may offer a new option to access silent BGCs. Here, we first pinpointed two highly efficient fusion sites for initiation unit exchanges. Subsequently, they were validated by replacing the initiation region of endopyrrole pathway with a Cs-containing initiation unit for generation of a lipo-endopyrrole derivate. We promptly leveraged this strategy of changing initiation unit to target six previously silent NRPS BGCs, three BGCs were successfully activated and five novel lipopeptides were identified, demonstrating its application to recover silent BGCs. Furthermore, we extended this strategy in more BGCs from different bacteria. Utilizing a heterologous Cs-containing unit to replace the initiation region of chitinimide biosynthetic pathway led to successful incorporation of N-terminal fatty acid chains into chitinimide to create artificial lipo-chitinimides. This study provides a feasible strategy to rationally recover silent BGCs and add fatty acid chains to NRPs, enriching the genome mining and combinatorial biosynthesis approach for bacterial natural products. SignificanceNonribosomal peptide synthetases (NRPSs) represent a valuable yet underexplored reservoir for bioactive natural products, but most of them are silent. This study introduces the concept that changing initiation unit to activate and optimize the functional expression of NRPSs, and successfully access three of six previously silent NRPS pathways, providing a feasible complement to the current genome mining approaches. Moreover, the N-terminal lipid chains are crucial for the activity of lipopeptides. We employed a heterologous Cs (starter condensation domain)-containing initiation unit to replace the original initiation region for creation of artificial nonribosomal lipopeptides, ultimately yielding three novel lipo-derivatives. This work presents a groundbreaking approach for activating and optimizing NRPSs and provides profound insight into the exploration of bacterial NRPS pathways.

synthetic biology↗
bioRxiv2025

1,3- and 1,4-linked Polysaccharides Uptake in Intestinal Cells Relies on Clathrin/Dynamin 1/Rab5-dependent Endocytosis

Natural polysaccharides are considered to be a promising feedstock for manufacturing sustainable materials. However, polysaccharides are believed by most biologists that they could not be absorbed by intestinal mucosa to enter circulation system owing to their high hydrophilicity and large molecule size. Herein, {beta}-1,3-linked glucan (GFPBW1) and -1,4-linked glucan (WGE) is used as the model polysaccharide and the transport experiments show that different linked and charged polysaccharides can permeate Caco-2 cell monolayers. The in vivo experiments demonstrate that GFPBW1 and WGE are detectable in rodent plasma and liver after oral administration the isotope (99mTc, 3H) and fluorescein labeled polysaccharides. Further study show that polysaccharide uptake requires clathrin heavy chain (CLTC) and its associated factors Rab5 and dynamin1 in intestinal cells using gene-knockdown strategies and inhibitors. Strikingly, polysaccharide absorption is attenuated in both CLTC intestine deficient mice and Rab5, dynamin1 conventional-knockout mice. Importantly, membrane receptors bone morphogenetic protein receptor type IA (BMPRIA), Dectin-1 and epidermal growth factor receptor (EGFR) are also critical for specific structural polysaccharide internalization. These findings provide novel insight to understand polysaccharide absorption mechanism and lay foundation for oral polysaccharide- based new drugs development.

pharmacology and toxicology↗
bioRxiv2024

GESIAP3.0: Sensor-based Image Analysis Program for Transmission Visualization In Vivo

Synaptic transmission mediated by various neurotransmitters influences a wide range of behaviors. However, understanding how neuromodulatory transmitters encode diverse behaviors and affect their functions remains challenging. Here, we introduce GESIAP3.0, an advanced, third-generation image analysis program based on genetically encoded sensors. This tool enables precise quantitative analysis of transmission in both awake, freely moving animals and immobilized subjects. GESIAP3.0 incorporates movement correction algorithms that effectively eliminate image displacement in behaving animals while optimizing synaptic information extraction and simplifying computations on commodity computers. Quantitative analysis of cholinergic, dopaminergic, and serotonergic transmission, corrected for tissue movement, revealed synaptic properties consistent with measurements from ex vivo wide-field and in vivo two-photon imaging under stable conditions. This validates the applicability of GESIAP3.0 for analyzing synaptic properties of neuromodulatory transmission in behaving animals.

bioengineering↗
bioRxiv2024

Single-Cell Resolution DESI Mass Spectrometry Imaging through 10-Fold Sample Expansion

Desorption electrospray ionization mass spectrometry imaging (DESI MSI) is a valuable tool for label-free, spatially resolved molecular analysis of biological tissues. However, its spatial resolution has been limited to tens of micrometers due to constraints in spray geometry and solvent flow, hindering single-cell imaging. Inspired by expansion microscopy, we developed a tenfold DESI expansion MSI (10x-DESI ExMSI) technique to achieve cellular-level resolution. In this study, tissue sections were embedded in a swellable polyelectrolyte gel, and physically enlarged by approximately tenfold without compromising molecular integrity. After drying of the sample, mass spectrometry imaging was performed using a custom-built DESI ion source coupled with an Orbitrap mass spectrometer. Using mouse brain tissues as representative samples, 10x-DESI ExMSI achieved a spatial resolution of approximately 5 {micro}m, comparable to the highest levels attained by advanced commercial MALDI-MSI systems. Enhanced resolution allowed clear visualization of fine structural details, such as the granule cell layer of the dentate gyrus and the pyramidal layer of the Cornu Ammonis, which were unresolved in unexpanded samples under a 100 m step size. Lipid profiling demonstrated high retention and detection efficiency, confirming the techniques suitability for comprehensive molecular imaging. The 10x-DESI ExMSI significantly advances DESI MSI by enabling single-cell level resolution without modifying existing mass spectrometers. The method is simple, cost-effective, and easily implementable in most laboratories, making high-resolution DESI MSI accessible for a wide range of biological applications. This approach overcomes longstanding limitations of DESI imaging and holds great potential for detailed molecular studies at the cellular level.

biochemistry↗
bioRxiv2024

Ten-Fold Expansion MALDI Mass Spectrometry Imaging of Tissues and Cells at 500 nm Resolution

Achieving high spatial resolution in matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is crucial for detailed molecular mapping in biological tissues and cells. However, conventional MALDI-MSI platforms are typically limited to spatial resolutions of 5-20 m, restricting their ability to visualize fine subcellular structures. Here, we present a novel ten-fold expansion MALDI-MSI (10X ExMSI) method that attains 500 nm spatial resolution without the need for specialized optics or customized instrumentation. By physically expanding biological samples using a swellable hydrogel matrix, our method maintains high retention and broadens the detection range of lipids, ensuring comprehensive lipid profiling. We demonstrated the effectiveness of 10X ExMSI by visualizing intricate subcellular structures within mouse brain tissues, including individual nuclei, astrocytes, Purkinje cells, and, notably, dendritic arborizations-features previously unresolvable using mass spectrometry imaging. Applying 10X ExMSI to single-cell analysis of cultured cells revealed detailed spatial distributions of lipids at the subcellular to organelle level. Notably, the method is fully compatible with standard commercial MALDI-MSI platforms, enabling widespread adoption without significant additional investment. The 10X ExMSI offers a transformative tool for high-resolution molecular imaging, opening new avenues for molecular analysis in diverse biological and biomedical fields, including neuroscience, pathology, and cellular biology.

biochemistry↗
bioRxiv2024

eProbe: a capture probe design toolkit for genetic diversity reconstructions from ancient environmental DNA

Ancient environmental DNA (aeDNA) is now commonly used in paleoecology and evolutionary ecology, yet due to difficulties in gaining sufficient genome coverage on individual species from metagenome data, its genetic perspectives remain largely uninvestigated. Hybridization capture has proven as an effective approach for enriching the DNA of target species, thus increasing the genome coverage of sequencing data and enabling population and evolutionary genetics analysis. However, to date there is no tool available for designing capture probe sets tailored for aeDNA based population genetics. Here we present eProbe, an efficient, flexible and easy-to-use program toolkit that provides a complete workflow for capture probe design, assessment and validation. By benchmarking a probe set for foxtail millet, an annual grass, made by the eProbe workflow, we demonstrate a remarkable increase of capturing efficiency, with the target taxa recovery rate improved by 577-fold, and the genome coverage achieved by soil capture-sequencing data even higher than data directly shotgun sequenced from the plant tissues. Probes that underwent our filtering panels show notably higher efficiency. The capture sequencing data enabled accurate population and evolutionary genetic analysis, by effectively inferring the fine-scale genetic structures and patterns, as well as the genotypes on functional genes.

bioinformatics↗
bioRxiv2023

Expansion Mass Spectrometry Imaging: Achieving One-Micron Level Resolution for Lipid Analysis in Tissues

A novel method for enhanced resolution, termed expansion mass spectrometry imaging (Ex-MSI), has been developed for lipid mass spectrometry imaging, utilizing existing commercially available mass spectrometers without necessitating modifications. This approach involves embedding tissue sections in a swellable polyelectrolyte gel, with the target biomolecules indirectly anchored to the gel network. By employing matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI), the method has realized an enhancement in spatial resolution that surpasses the conventional resolution limits of commercial instruments by approximately 4.5 folds. This enhancement permits the detailed visualization of intricate structures within the mouse brain at a subcellular level, with a lateral resolution nearing 1 m. As a physical technique for achieving resolution beyond standard capabilities, this approach is readily adaptable and presents a powerful tool for high-definition imaging in biological research.

biochemistry↗
bioRxiv2022

Chiral Derivatization-enabled Discrimination and Visualization of Proteinogenic Amino Acids by Ion Mobility Mass Spectrometry

The importance of chiral amino acids (AAs) in living organisms has been widely recognized since the discovery of endogenous D-AAs as potential biomarkers in several metabolic disorders. Chiral analysis by ion mobility spectrometry-mass spectrometry (IMS-MS) has the advantages of high speed and sensitivity but is still in its infancy. Here, a N-(2,4-dinitro-5-fluorophenyl)-L-alaninamide (FDAA) derivatization is combined with trapped ion mobility spectrometry-mass spectrometry (TIMS-MS) for chiral AA analysis. For the first time, we demonstrate the simultaneous separation of 19 pairs of chiral proteinogenic AAs in a single fixed condition TIMS-MS run. The utility of this approach presents for mouse brain extracts by direct-infusion TIMS-MS. The robust separation ability in complex biological sample was proven in MALDI TIMS mass spectrometry imaging (MSI) as well by directly depositing 19 pairs of AAs on a tissue slide following on-tissue derivatization. In addition, endogenous chiral amino acids were also detected and distinguished. The developed methods show compelling application prospects in biomarker discovery and biological research. Entry for the Table of Contents O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=102 SRC="FIGDIR/small/498692v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@19af718org.highwire.dtl.DTLVardef@d457d7org.highwire.dtl.DTLVardef@d2f932org.highwire.dtl.DTLVardef@e64831_HPS_FORMAT_FIGEXP M_FIG C_FIG The combination of chiral derivatization and trapped ion mobility-mass spectrometry provides the first insights into the separation of 19 pairs of chiral proteinogenic D/L-amino acids in a single run and further visualization of chiral amino acids under complex biological matrix.

biochemistry↗
bioRxiv2022

SiPLATZ12 transcript factor regulates multiple yield traits and salt tolerance in foxtail millet (Setaria italica)

Grain yield and salt tolerance are critical for crop production. However, the genetic and biochemical basis underlying the trade-off of these characters remain poorly described in crops. We show here that SiPLATZ12 transcription factor positively regulates multiple elite yield traits at the expense of salt tolerance in foxtail millet. SiPLATZ12 overexpression increases seed size, panicle length, and stem diameter, while reduces plant height and salt tolerance of foxtail millet. A 9-bp insertion in the SiPLATZ12 promoter has significant effects on the different expression of SiPLATZ12, multiple yield traits, and salt tolerance between foxtail millet and its wild ancestor, green foxtail. Moreover, SiPLATZ12 upregulates the expression of genes involved in seed development, but repressing the transcription of most NHX, SOS, and CBL genes to regulate Na+, K+ and pH homeostasis. Therefore, our results uncover a domesticated site that could be used to improve grain yield and salt tolerance in foxtail millet.

molecular biology↗
bioRxiv2021

The role of AUX1 during lateral root development in the domestication of the model C4 grass Setaria italica

C4 photosynthesis increases the efficiency of carbon fixation by spatially separating high concentrations of molecular oxygen from rubisco. The specialized leaf anatomy required for this separation evolved independently many times. C4 root systems are highly branched, an adaptation thought to support high rates of photosynthesis; however, little is known about the molecular mechanisms that have driven the evolution of C4 root system architecture (RSA). Using a mutant screen in the C4 model plant Setaria italica, we identify siaux1-1 and siaux1-2 as RSA mutants, and use CRISPR/cas9-mediated genome editing and overexpression to confirm the importance of the locus. As AUX1 is not necessary for lateral root emergence in S. viridis, the species from which S. italica was domesticated, we conducted an analysis of auxin responsive elements in the promoters of auxin-responsive gene families in S. italica, and explore the molecular basis of SiAUX1s role in seedling development using an RNAseq analysis of wild type and siaux1-1 plants. Finally, we use a root coordinate system to compare cell-by-cell meristem structures in siaux1-1 and wild type Setaria plants, observing changes in the distribution of cell volumes in all cell layers and a dependence in the frequency of protophloem and protoxylem strands on siAUX1.

plant biology↗