Search bioRxiv⌕ Search

Biology subjects

Zhang, M.

Publications and source records attributed to Zhang, M..

At least 433 records · Page 24Linked to original sources

Functional interrogation of twenty type 2 diabetesassociated genes using isogenic hESC-derived β-like cells

Genetic studies have identified numerous loci associated with type 2 diabetes (T2D), but the functional role of many loci has remained unexplored. In this study, we engineered isogenic knockout human embryonic stem cell (hESC) lines for 20 genes associated with T2D risk. We systematically examined {beta}-cell differentiation, insulin production and secretion, and survival. We performed RNA-seq and ATAC-seq on hESC-{beta} cells from each knockout line. Analyses of T2D GWAS signals overlapping with HNF4A-dependent ATAC peaks identified a specific SNP as a likely causal variant. In addition, we performed integrative association analyses and identified four genes (CP, RNASE1, PCSK1N and GSTA2) associated with insulin production, and two genes (TAGLN3 and DHRS2) associated with sensitivity to lipotoxicity. Finally, we leveraged deep ATAC-seq read coverage to assess allele-specific imbalance at variants heterozygous in the parental hESC line, to identify a single likely functional variant at each of 23 T2D GWAS signals.

genetics↗

Complement C3 reduces apoptosis in human cardiomyocytes

Complement C3 is a key factor in complement system. Our recently animal study found that C3 may regulate myocardial apoptosis through the intrinsic apoptosis pathway. The current work investigated if C3 regulation of apoptosis occurred in human cardiomyocytes. Our results showed that incubation of exogenous C3 reduced apoptosis in a cell culture system of human cardiomyocytes which did not inherently express C3. In addition, C3 inhibited intrinsic apoptosis pathway in a cell-free apoptosis system. Furthermore, pro-C3 was found to bind with an apoptotic factor, pro-caspase 3, in a cell-free system. Thus, we presented firsthand evidence that exogenous C3 is readily reduce apoptosis in human cardiomyocytes via interaction with the intrinsic apoptotic pathway.

cell biology↗

LSM14B is essential for mitochondrial clustering in the oocyte meiosis

As oocyte meiotic maturation, they undergo two successive meiotic M phases, notably lacking an intervening interphase phase. During these M phases, oocytes remain transcriptionally quiescent, and we now know that "translational repressed mRNAs" are stored in a structure called the mitochondria associated ribonucleoprotein domain (MARDO). LSM14B is one of the abundant proteins of MARDO, and is predicted to bind mRNA, but its function(s) remain elusive. Here, we demonstrate that LSM14B functions to promote MARDO assembly in mouse oocytes. We also found that LSM14B knockout female mice are infertile, and show that the knockout oocytes fail to enter meiosis II, instead entering an aberrant interphase-like stage. Finally, we show that the failure of oocyte maturation results from decreased expression of Cyclin B1. Our study has revealed that the RNA-binding protein LSM14B modulates MARDO assembly and is essential for oocyte meiotic maturation.

developmental biology↗

Improving the solubility of pseudo-hydrophobic Alzheimer's Disease medicinal chemicals through co-crystal formulation

Natural products are ligands and potential inhibitors of Alzheimers disease (AD) tau. Dihydromyricetin (DHM) is a CNS active natural product. Despite having signature polyphenolic character, DHM is ostensibly hydrophobic owing to intermolecular hydrogen bonds that shield hydrophilic phenols. Our research shows DHM becomes ionized at near-neutral pH allowing formulation of salts with transformed solubility. The MicroED co-crystal structure with trolamine reveals DHM salts as metastable solids with unlocked hydrogen bonding and a thermodynamic bent to solubilize in water. All salt formulations show better inhibitory activity against AD tau than the non-salt form, with efficacies correlating to enhanced solubilities. These results underscore the role of structural chemistry in guiding selection of solubilizing agents for chemical formulation. We propose DHM salts are appropriate formulations for research as dietary supplements to promote healthy aging by combating protein misfolding. Additionally, DHM is a suitable lead for medicinal chemistry and possible development of CNS pharmaceuticals.

pharmacology and toxicology↗

Meiotic recombination dynamics in plants with repeat-based holocentromeres shed light on the primary drivers of crossover patterning

Centromeres strongly affect (epi)genomic architecture and meiotic recombination dynamics influencing the overall distribution and frequency of crossovers. Here, we studied how recombination is regulated and distributed in the holocentric plant Rhynchospora breviuscula, a species lacking localised centromeres. Combining immunocytochemistry, chromatin analysis and high-throughput single-pollen sequencing, we discovered that crossover frequency is higher at ends related to centred chromosomal regions. Contrasting the diffused distribution of (epi)genetic features and hundreds of repeat-based centromeric units. Remarkably, we found that crossovers were abolished at core centromeric units but not at their vicinity indicating the absence of a centromere effect across repeat-based holocentromeres. We further show that telomere-led pairing and synapsis of homologous chromosomes appear to be the primary force determining the observed U-shaped recombination landscape. While centromere and (epi)genetic properties only affect crossover positioning locally. Our results suggest that the conserved U-shaped crossover distribution of eukaryotes is independent of chromosome compartmentalisation and centromere organisation.

genomics↗

Discovery and Validation of RUNX1 DNA Methylation in Differentiating Papillary Thyroid Cancer from Benign Nodules

Although most thyroid nodules can be diagnosed preoperatively by thyroid ultrasonography and fine-needle aspiration biopsy, it remains a challenge to accurately identify malignancy of thyroid nodules when the biopsy is indeterminate. This study aims to explore a novel biomarker to distinguish benign and malignant thyroid nodules. Tissue samples from patients with Stage I&II papillary thyroid carcinoma (PTC) and benign thyroid nodules (BTN) were collected for genome profiling by methylation EPIC 850K array and RNA-Sequencing. Genes with significantly differential DNA methylation and inverse mRNA expression were filtered out. The altered methylation of RUNX1 gene was validated in two independent case-control studies with a total of 699 formalin fixed paraffin-embedded (FFPE) samples using mass spectrometry and calculated by binary logistic regression analysis. Hypomethylation of RUNX1 gene in PTC patients compared to BTN subjects was verified in Validation (140 PTC vs. 189 BTN, ORs [≥] 1.50 per-10% methylation, P [≤] 4.40E-05, for all measurable CpG sites) and Validation (184 PTC vs. 186 BTN, ORs [≥] 1.72 per-10% methylation, P [≤] 2.38E-11, for all measurable CpG sites). Besides, RUNX1 methylation achieved good accuracy in differentiating early-stage PTC from BTN in Validation (AUC: 0.74) and Validation (AUC: 0.79). Gender- and age-stratified analysis revealed RUNX1 hypomethylation as an important risk factor for thyroid disease in younger women. We disclosed a significant association between RUNX1 hypomethylation and PTC, suggesting RUNX1 methylation based on FFPE tissue samples as a potential biomarker for predicting malignancy of thyroid nodules.

genetics↗

APART-QSM: an improved sub-voxel quantitative susceptibility mapping for susceptibility source separation using an iterative data fitting method

The brain tissue phase contrast in MRI sequences reflects the spatial distributions of multiple substances, such as iron, myelin, calcium, and proteins. These substances with paramagnetic and diamagnetic susceptibilities often colocalize in one voxel in brain regions. Both opposing susceptibilities play vital roles in brain development and neurodegenerative diseases. Conventional QSM methods only provide voxel-averaged susceptibility value and cannot disentangle intravoxel susceptibilities with opposite signs. Advanced susceptibility imaging methods have been recently developed to distinguish the contributions of opposing susceptibility sources for QSM. The basic concept of separating paramagnetic and diamagnetic susceptibility proportions is to include the relaxation rate [Formula] with [Formula] in QSM. The magnitude decay kernel, describing the proportionality coefficient between [Formula] and susceptibility, is an essential reconstruction coefficient for QSM separation methods. In this study, we proposed a more comprehensive complex signal model that describes the relationship between 3D GRE signal and the contributions of paramagnetic and diamagnetic susceptibility to the frequency shift and [Formula] relaxation. The algorithm is implemented as a constrained minimization problem in which the voxel-wise magnitude decay kernel and sub-voxel susceptibilities are determined alternately in each iteration until convergence. The calculated voxel-wise magnitude decay kernel could realistically model the relationship between the [Formula] relaxation and the volume susceptibility. Thus, the proposed method effectively prevents the errors of the magnitude decay kernel from propagating to the final susceptibility separation reconstruction. Phantom studies, ex vivo macaque brain experiments, and in vivo human brain imaging studies were conducted to evaluate the ability of the proposed method to distinguish paramagnetic and diamagnetic susceptibility sources. The results demonstrate that the proposed method provides state-of-the-art performances for quantifying brain iron and myelin compared to previous QSM separation methods. Our results show that the proposed method has the potential to simultaneously quantify whole brain iron and myelin during brain development and aging. The proposed model was also deployed with multiple-orientation complex GRE data input measurements, resulting in high-quality QSM separation maps with more faithful tissue delineation between brain structures compared to those reconstructed by single-orientation QSM separation methods.

neuroscience↗

Convergence of resistance and evolutionary responses in Escherichia coli and Salmonella enterica co-inhabiting chicken farms in China

Sharing among different pathogens and commensals inhabiting same hosts and environments has significant implications for antimicrobial resistance (AMR), especially in settings with high antimicrobial exposure. We analysed 661 E. coli and S. enterica isolates collected within and across hosts and environments, in 10 Chinese chicken farms over 2.5 years using novel data-mining methods. Most isolates within same hosts possessed same clinically relevant AMR-carrying mobile genetic elements (plasmids: 70.6%, transposons: 78%), which also showed recent common evolution. Machine learning revealed known and novel AMR-associated mutations and genes underlying resistance to 28 antimicrobials and primarily associated with resistance in Escherichia coli and susceptibility in Salmonella enterica. Many of these genes were essential and affected same metabolic processes in both species, albeit with varying degrees of phylogenetic penetration. Multi-modal strategies are crucial to investigate the interplay of mobilome, resistance and metabolism in cohabiting bacteria, especially in ecological settings where community-driven resistance selection occurs.

genomics↗

Optics-aware super-resolution light-field microscopy for long-term volumetric imaging of dynamic intracellular processes at millisecond timescales

Long-term and high-spatiotemporal-resolution 3D imaging of living cells remains an unmet challenge for super-resolution microscopy, owing to the noticeable phototoxicity and limited scanning speed. While emerging light-field microscopy can mitigate this issue through three-dimensionally capturing biological dynamics with merely single snapshot, it suffers from suboptimal resolution insufficient for resolving subcellular structures. Here we propose an Adaptive Learning PHysics-Aware Light-Field Microscopy (Alpha-LFM) with a physics-aware deep learning framework and adaptive-tuning strategies capable for highly-generalizable light-field reconstruction of diverse subcellular dynamics. Alpha-LFM delivers sub-diffraction-limit spatial resolution ([~]120 nm) while maintaining high temporal resolution and low phototoxicity. It enables rapid (at hundreds of volumes per second), long-term (up to 60 hours) 3D super-resolution imaging of diverse intracellular dynamics with exceptional details. Using Alpha-LFM approach, we finely resolve the lysosome-mitochondrial interactions, capture rapid motion of peroxisome and the endoplasmic reticulum, and reveal the variations in mitochondrial fission activity throughout two complete cell cycles.

microbiology↗

Evaluating performance and applications of sample-wise cell deconvolution methods on human brain transcriptomic data

Sample-wise deconvolution methods have been developed to estimate cell-type proportions and gene expressions in bulk-tissue samples. However, the performance of these methods and their biological applications has not been evaluated, particularly on human brain transcriptomic data. Here, nine deconvolution methods were evaluated with sample-matched data from bulk-tissue RNAseq, single-cell/nuclei (sc/sn) RNAseq, and immunohistochemistry. A total of 1,130,767 nuclei/cells from 149 adult postmortem brains and 72 organoid samples were used. The results showed the best performance of dtangle for estimating cell proportions and bMIND for estimating sample-wise cell-type gene expression. For eight brain cell types, 25,273 cell-type eQTLs were identified with deconvoluted expressions (decon-eQTLs). The results showed that decon-eQTLs explained more schizophrenia GWAS heritability than bulk-tissue or single-cell eQTLs alone. Differential gene expression associated with multiple phenotypes were also examined using the deconvoluted data. Our findings, which were replicated in bulk-tissue RNAseq and sc/snRNAseq data, provided new insights into the biological applications of deconvoluted data.

genomics↗

Complement C3 interacts with cytochrome c to influence myocardial apoptosis during heart ischemia/reperfusion

Myocardial ischemia/reperfusion (I/R) elicits an acute inflammatory response involving complement factors. Previous animal studies showed that circulation complement C3 was deposited in the ischemic myocardium flooded with oxygenated blood upon reperfusion. Recently, we reported that myocardial necrosis was decreased in C3-/- mice after heart I/R. The current study used in the same heart model to test the effect of C3 on myocardial apoptosis. Our results showed that myocardial apoptosis was increased in C3-/- mice after heart I/R. Further, comparative proteomics analyses found that cytochrome c was present in the myocardial C3-complex following I/R. These results indicate that C3 can interact with cytochrome c in the cytosol of cardiomyocytes during myocardial I/R, which may sequester cytochrome c and thus reduce the number of cells undergoing apoptosis. In summary, our findings raise the possibility of a new mechanism affecting cell death relevant to pathologic conditions such as ischemia: a circulating innate immune factor, i.e. complement, can interact with intracellular factor(s), and influence the types of cell death that occur.

molecular biology↗

A molecularly defined and spatially resolved cell atlas of the whole mouse brain

In mammalian brains, tens of millions to billions of cells form complex interaction networks to enable a wide range of functions. The enormous diversity and intricate organization of cells in the brain have so far hindered our understanding of the molecular and cellular basis of its functions. Recent advances in spatially resolved single-cell transcriptomics have allowed systematic mapping of the spatial organization of molecularly defined cell types in complex tissues1-3. However, these approaches have only been applied to a few brain regions1-11 and a comprehensive cell atlas of the whole brain is still missing. Here, we imaged a panel of >1,100 genes in [~]8 million cells across the entire adult mouse brain using multiplexed error-robust fluorescence in situ hybridization (MERFISH)12 and performed spatially resolved, single-cell expression profiling at the whole-transcriptome scale by integrating MERFISH and single-cell RNA-sequencing (scRNA-seq) data. Using this approach, we generated a comprehensive cell atlas of >5,000 transcriptionally distinct cell clusters, belonging to [~]300 major cell types, in the whole mouse brain with high molecular and spatial resolution. Registration of the MERFISH images to the common coordinate framework (CCF) of the mouse brain further allowed systematic quantifications of the cell composition and organization in individual brain regions defined in the CCF. We further identified spatial modules characterized by distinct cell-type compositions and spatial gradients featuring gradual changes in the gene-expression profiles of cells. Finally, this high-resolution spatial map of cells, with a transcriptome-wide expression profile associated with each cell, allowed us to infer cell-type-specific interactions between several hundred pairs of molecularly defined cell types and predict potential molecular (ligand-receptor) basis and functional implications of these cell-cell interactions. These results provide rich insights into the molecular and cellular architecture of the brain and a valuable resource for future functional investigations of neural circuits and their dysfunction in diseases.

neuroscience↗

A high-resolution transcriptomic and spatial atlas of cell types in the whole mouse brain

The mammalian brain is composed of millions to billions of cells that are organized into numerous cell types with specific spatial distribution patterns and structural and functional properties. An essential step towards understanding brain function is to obtain a parts list, i.e., a catalog of cell types, of the brain. Here, we report a comprehensive and high-resolution transcriptomic and spatial cell type atlas for the whole adult mouse brain. The cell type atlas was created based on the combination of two single-cell-level, whole-brain-scale datasets: a single- cell RNA-sequencing (scRNA-seq) dataset of [~]7 million cells profiled, and a spatially resolved transcriptomic dataset of [~]4.3 million cells using MERFISH. The atlas is hierarchically organized into five nested levels of classification: 7 divisions, 32 classes, 306 subclasses, 1,045 supertypes and 5,200 clusters. We systematically analyzed the neuronal, non-neuronal, and immature neuronal cell types across the brain and identified a high degree of correspondence between transcriptomic identity and spatial specificity for each cell type. The results reveal unique features of cell type organization in different brain regions, in particular, a dichotomy between the dorsal and ventral parts of the brain: the dorsal part contains relatively fewer yet highly divergent neuronal types, whereas the ventral part contains more numerous neuronal types that are more closely related to each other. We also systematically characterized cell-type specific expression of neurotransmitters, neuropeptides, and transcription factors. The study uncovered extraordinary diversity and heterogeneity in neurotransmitter and neuropeptide expression and co-expression patterns in different cell types across the brain, suggesting they mediate a myriad of modes of intercellular communications. Finally, we found that transcription factors are major determinants of cell type classification in the adult mouse brain and identified a combinatorial transcription factor code that defines cell types across all parts of the brain. The whole-mouse-brain transcriptomic and spatial cell type atlas establishes a benchmark reference atlas and a foundational resource for deep and integrative investigations of cell type and circuit function, development, and evolution of the mammalian brain.

neuroscience↗

Video-rate Mid-infrared Photothermal Imaging by Single Pulse Photothermal Detection per Pixel

By optically sensing the mid-infrared absorption induced photothermal effect, midinfrared photothermal (MIP) microscope enables super-resolution IR imaging and scrutinizing of biological systems in an aqueous environment. However, the speed of current lock-in based sample-scanning MIP system is limited to 1.0 millisecond or longer per pixel, which is insufficient for capturing dynamics inside living systems. Here, we report a single pulse laserscanning MIP microscope that dramatically increases the imaging speed by three orders of magnitude. We harness a lock-in free demodulation scheme which uses high-speed digitization to resolve single IR pulse induced contrast at nanosecond time scale. To realize single pulse photothermal detection at each pixel, we employ two sets of galvo mirrors for synchronized scanning of mid-infrared and probe beams to achieve an imaging line rate over 2 kHz. With video-rate imaging capability, we observed two types of distinct dynamics of lipids in living cells. Furthermore, by hyperspectral imaging, we chemically dissected a single cell wall at nanometer scale. Finally, with a uniform field of view over 200 by 200 m2 and 2 Hz frame rate, we mapped fat storage in free-moving C. elegans and live embryos.

bioengineering↗

Cryo-EM-guided engineering of T-box-tRNA modules with enhanced selectivity and sensitivity in translational regulation

Riboswitches are non-coding RNA elements that play vital roles in regulating gene expression. Their specific ligand-dependent structural reorganization facilitates their use as templates for design of engineered RNA switches for therapeutics, nanotechnology and synthetic biology. T-box riboswitches bind tRNAs to sense aminoacylation and control gene expression via transcription attenuation or translation inhibition. Here we determine the cryo-EM structure of the wild-type Mycobacterium smegmatis ileS T-box in complex with its cognate tRNAIle. This structure shows a very flexible antisequestrator region that tolerates both 3-OH and 2,3-cyclic phosphate modification at the 3 end of tRNAIle. Elongation of one helical turn (11-base pair) in both the tRNA acceptor arm and T-box Stem III maintains T-box-tRNA complex formation and increases the selectivity for tRNA 3 end modification. Moreover, elongation of Stem III results in [~]6-fold tighter binding to tRNA, which leads to increased sensitivity of downstream translational regulation indicated by precedent translation. Our results demonstrate that cryo-EM can guide RNA engineering to design improved riboswitch modules for translational regulation, and potentially a variety of additional functions.

synthetic biology↗

Drug-tolerant persister B-cell precursor acute lymphoblastic leukemia cells

Reduced responsiveness of precursor B-acute lymphoblastic leukemia (BCP-ALL) to chemotherapy can be inferred when leukemia cells persist after 28 days of initial treatment. Survival of these long-term persister (LTP) / minimal residual disease (MRD) cells is partly due to bone marrow stromal cells that protect them under conditions of chemotherapy stress. We used RNA-seq to analyse BCP-ALL cells that survived a long-term, 30-day vincristine chemotherapy treatment while in co-culture with bone marrow stromal cells. RNAs of as many as 10% of the protein-encoding genes were differentially expressed. There was substantial overlap with genes associated with MRD cell persistence reported in other studies. The top pathway regulated in the LTP cells was that involving p53, a master regulator of a spectrum of responses relevant to drug resistance and cytotoxic drug exposure including control of autophagy. We tested a select number of genes for contribution to BCP-ALL cell survival using Cas9/CRISPR in a 2-step selection, initially for overall effect on cell fitness, followed by 21 days of exposure to vincristine. Many genes involved in autophagy and lysosomal function were found to contribute to survival both at steady-state and during drug treatment. We also identified MYH9, NCSTN and KIAA2013 as specific genes contributing to fitness of BCP-ALL cells. CD44 was not essential for growth under steady state conditions but was needed for survival of vincristine treatment. Finally, although the drug transporter ABCC1/MRP1 is not overexpressed in BCP-ALL, a functional gene was needed for DTP cells to survive treatment with vincristine. This suggests that addition of possible ABCC1 inhibitors during induction therapy could provide benefit in eradication of minimal residual disease in patients treated with a chemotherapy regimen that includes vincristine.

cancer biology↗

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

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

cell biology↗

Integrated Cell Landscape and Dynamics in the Progression of Bone Repair

Bone homeostasis and repair is a systematic progress with spatiotemporal interaction of multiple cell types involved in skeletal and immune system. Precise spatiotemporal regulation of cell type-specific functions in bone repair contributes to further development of tissue engineering and regenerative medicine. Here, we utilized single-cell RNA sequencing to illustrate a map of cell landscape and dynamics in the progression of rodent bone self-healing and a perturbation by lymphoid cell-deficiency. We identified different functions of myeloid cell and lymphoid cell to osteogenesis and angiogenesis during bone repair and their mutual complementation under lymphoid cell-deficient condition. Additionally, we used CD34+ humanized reconstituted mice to reveal further insights into the mechanism of human bone homeostasis and repair. Our integrated cellular analysis of bone repair explores the functional diversity and complementation between myeloid cells and lymphoid cells during bone healing process and provides further therapeutic implications for the treatment of bone disease and degeneration following ageing.

cell biology↗