Biology subjects

Huang, S.

Publications and source records attributed to Huang, S..

At least 235 records · Page 13Linked to original sources

bioRxiv2023

Tetravalent SARS-CoV-2 S1 Subunit Protein Vaccination Elicits Robust Humoral and Cellular Immune Responses in SIV-Infected Rhesus Macaque Controllers

The COVID-19 pandemic has highlighted the need for safe and effective vaccines to be rapidly developed and distributed worldwide, especially considering the emergence of new SARS-CoV-2 variants. Protein subunit vaccines have emerged as a promising approach due to their proven safety record and ability to elicit robust immune responses. In this study, we evaluated the immunogenicity and efficacy of an adjuvanted tetravalent S1 subunit protein COVID-19 vaccine candidate composed of the Wuhan, B.1.1.7 variant, B.1.351 variant, and P.1 variant spike proteins in a nonhuman primate model with controlled SIVsab infection. The vaccine candidate induced both humoral and cellular immune responses, with T- and B cell responses mainly peaking post-boost immunization. The vaccine also elicited neutralizing and cross-reactive antibodies, ACE2 blocking antibodies, and T-cell responses, including spike specific CD4+ T cells. Importantly, the vaccine candidate was able to generate Omicron variant spike binding and ACE2 blocking antibodies without specifically vaccinating with Omicron, suggesting potential broad protection against emerging variants. The tetravalent composition of the vaccine candidate has significant implications for COVID-19 vaccine development and implementation, providing broad antibody responses against numerous SARS-CoV-2 variants.

immunology↗

bioRxiv2023

High Resolution Salmonella Subtyping by the MLSTnext Method

The food industry needs a straightforward, efficient, widely applicable and cost-efficient method to not only detect, but also determine serovar and potential sources of multiple strains in food and environment. While Whole Genome Sequencing (WGS) can generate complete genomic profile of food pathogens, it is a laborious, time-consuming, and expensive method that necessitates pure isolates. As a result, it is unsuitable for samples with complex background, limiting its widespread application by food industry. However, traditional multilocus sequence typing (MLST) approaches do not provide sufficient single nucleotide polymorphism (SNP) information to effectively track- and-trace sources of contamination. In contrast, ChapterDx MLSTnext-NGS (next-generation sequencing) Salmonella assay amplifies and sequences 47 polymorphic loci, evenly spanning the Salmonella enterica genome. As demonstrated in this study, the ChapterDx MLSTnext-NGS Salmonella can identify serovar with high resolution, distinguishing between various strains of the same serovar and resolve co-presence of different strains in a single sample. Additionally, this assay can analyze up to thousand samples in a single sequencing run within 22 hours, making it a highly efficient and scalable method for the food industry. Moreover, the cost per sample of the ChapterDx MLSTnext-NGS Salmonella assay is comparable to that of quantitative Polymerase Chain Reaction (qPCR), making it an affordable option. The assay uses ChapterDx amplification technology, allowing for the amplification of all 47 loci in a single-tube, single-step PCR reaction. This makes it one of the simplest NGS applications available. In summary, ChapterDx MLSTnext-NGS Salmonella assay can be implemented in many diagnostic laboratories to address increasingly complex food safety issues.

microbiology↗

bioRxiv2023

Quantitative Nanohistology of aging dermal collagen

While the external signs of skin aging have been well-defined throughout history, much less is known about aging within the ultrastructure of our skin. Our skin, the largest organ in our body, is structured by collagen through fibrils or large sheets. With the increased use of nanometrology tools in histology, it is now possible to explore how the aging process affects collagen at its most fundamental level, the collagen fibril. Here, we show how atomic force microscopy-based quantitative nanohistology can differentiate skin from different age groups and anatomical sites. Following the definition of specific collagen biomarkers at the nanoscale, we used a segmentation approach to quantify the prevalence of 4 structural biomarkers over a dataset of 42,000 images (30 donors) complemented by extensive nanomechanical analyses (30,000 indentation curves) on histological sections. Our results demonstrate that specific age-related collagen fingerprints could be found when comparing the % prevalence of each marker between the papillary and reticular dermis. A case of abnormal biological aging validated our markers and nanohistology approach. This first extensive study focusing on defining signs of dermal aging at the nanoscale proves we are all unique to our dermal collagen ultrastructure.

biophysics↗

bioRxiv2023

ARF-family global interactome mapping uncovers spatial organization of cellular signaling pathways

The ADP-ribosylation factors (ARFs) and ARF-like (ARLs) GTPases serve as essential molecular switches governing a wide array of cellular processes. In this study, we utilized proximity-dependent biotin identification (BioID) to comprehensively map the interactome of 28 out of 29 ARF and ARL proteins in two cellular models. Through this approach, we identified [~]3000 high-confidence proximal interactors, enabling us to assign subcellular localizations to the family members. Notably, we uncovered previously undefined localizations for ARL4D and ARL10. Clustering analyses further exposed the distinctiveness of the interactors identified with these two GTPases. We also reveal that the expression of the understudied member ARL14 is confined to the stomach and intestines. We identified phospholipase D1 (PLD1) and the ESCPE-1 complex, more precisely SNX1, as proximity interactors. Functional assays demonstrated that ARL14 can activate PLD1 in cellulo and is involved in cargo trafficking via the ESCPE-1 complex. Overall, the BioID data generated in this study provide a valuable resource for dissecting the complexities of ARF and ARL spatial organization and signaling. SUMMARY STATEMENTGeneration of the ARF family interactome allowed the attribution of potential localizations and functions to previously understudied members. We found that ARL14 activates PLD1 and contributes to ESCPE-1-mediated trafficking.

molecular biology↗

bioRxiv2023

Single cell and spatial alternative splicing analysis with long read sequencing

Long-read sequencing has become a powerful tool for alternative splicing analysis. However, technical and computational challenges have limited our ability to explore alternative splicing at single cell and spatial resolution. The higher sequencing error of long reads, especially high indel rates, have limited the accuracy of cell barcode and unique molecular identifier (UMI) recovery. Read truncation and mapping errors, the latter exacerbated by the higher sequencing error rates, can cause the false detection of spurious new isoforms. Downstream, there is yet no rigorous statistical framework to quantify splicing variation within and between cells/spots. In light of these challenges, we developed Longcell, a statistical framework and computational pipeline for accurate isoform quantification for single cell and spatial spot barcoded long read sequencing data. Longcell performs computationally efficient cell/spot barcode extraction, UMI recovery, and UMI-based truncation- and mapping-error correction. Through a statistical model that accounts for varying read coverage across cells/spots, Longcell rigorously quantifies the level of inter-cell/spot versus intra-cell/ spot diversity in exon-usage and detects changes in splicing distributions between cell populations. Applying Longcell to single cell long-read data from multiple contexts, we found that intra-cell splicing heterogeneity, where multiple isoforms co-exist within the same cell, is ubiquitous for highly expressed genes. On matched single cell and Visium long read sequencing for a tissue of colorectal cancer metastasis to the liver, Longcell found concordant signals between the two data modalities. Finally, on a perturbation experiment for 9 splicing factors, Longcell identified regulatory targets that are validated by targeted sequencing.

bioinformatics↗

bioRxiv2023

Intraperitoneal transfer of wild-type bone marrow cells in the Csf1r knockout rat repopulates resident tissue macrophages without contributing to monocytopoiesis.

Homozygous null mutation of the macrophage colony-stimulating factor receptor (Csf1r) gene in rats leads to the loss of most tissue macrophage populations and has pleiotropic impacts on postnatal growth and organ maturation leading to mortality by 8-12 weeks of age. The phenotype of the Csf1r knockout (Csf1rko) can be reversed by intraperitoneal transfer of wild-type bone marrow cells (BMT) at weaning. Here we used a Csf1r-mApple transgenic reporter, which is expressed in neutrophils and B cells as well as monocytes and macrophages, to track the fate of donor-derived cells. Following BMT into Csf1r recipients, wild-type mApple+ve cells restored IBA1+ tissue macrophage populations in every tissue donor-derived cells also completely replaced recipient macrophages in organs such as spleen, lung and liver that were only partly macrophage-deficient in the Csf1rko. However, monocytes, neutrophils and B cells in bone marrow, blood and lymphoid tissues remained of recipient (mApple-ve) origin. An mApple+ve cell population expanded in the peritoneal cavity and invaded locally in the mesentery, fat pads, omentum and diaphragm. One week after BMT, distal organs contained foci of mApple+ve, IBA1-ve immature progenitors that appeared to proliferate, migrate and differentiate locally. We conclude that rat bone marrow contains progenitor cells that are able to restore and maintain all tissue macrophage populations in a Csf1rko rat directly without contributing to the bone marrow progenitor or blood monocyte populations.

immunology↗

bioRxiv2023

Cell population growth as manifestation of stochastic heterogeneity of cell phenotype

Recent studies at individual cell resolution have revealed phenotypic heterogeneity in nominally clonal tumor cell populations. The heterogeneity affects cell growth behaviors, which can result in departure from the idealized exponential growth. Here we measured the stochastic time courses of growth of an ensemble of populations of HL60 leukemia cells in cultures, starting with distinct initial cell numbers to capture the departure from the exponential growth model in the initial growth phase. Despite being derived from the same cell clone, we observed significant variations in the early growth patterns of individual cultures with statistically significant differences in growth kinetics and the presence of subpopulations with different growth rates that endured for many generations. Based on the hypothesis of existence of multiple inter-converting subpopulations, we developed a branching process model that captures the experimental observations.

cancer biology↗

bioRxiv2023

Metabolomics reveals nucleoside analogs for regulating mucosal-associated invariant T cell responses

Although mucosal-associated invariant T (MAIT) cells recognize riboflavin-like metabolites from Gram-negative bacteria, MAIT cell stimulation by broad bacterial families and mammalian cells suggests the existence of novel ligands from different biological sources. Here we established a comparative platform of functional metabolomics and used Mycobacterium tuberculosis as a model to characterize novel metabolites for MAIT cell activation. We extracted and fractionated small metabolites of M. tuberculosis using high-performance liquid chromatography, showing a different MAIT cell stimulation pattern of M. tuberculosis metabolite fractions in comparison with Escherichia coli fractions. Mass profiling predicted multiple nucleoside analogs enriched in a biologically active fraction of M. tuberculosis. Whereas the synthetic forms of these predicted M. tuberculosis nucleosides were unavailable, structural-based autodocking of analogous nucleosides conserved in mammals supported potential binding with MR1 protein. Indeed, functional assays of these conserved nucleosides demonstrated guanosine as a stimulator and deoxyformyluridine as an inhibitor of MAIT cell activation. Identification of bioactive nucleoside metabolites broadly conserved in bacterial and mammalian systems will facilitate an understanding of the regulatory roles of MAIT cells in infectious and inflammatory conditions.

immunology↗

bioRxiv2023

Interactive computational and experimental approaches improve the sensitivity of periplasmic binding protein-based nicotine biosensors for measurements in biofluids

We developed fluorescent protein sensors for nicotine with improved sensitivity. For iNicSnFR12 at pH 7.4, the proportionality constant for {Delta}F/F0 vs [nicotine] ({delta}-slope, 2.7 M-1) is 6.1-fold higher than the previously reported iNicSnFR3a. The activated state of iNicSnFR12 has a fluorescence quantum yield of at least 0.6. We measured similar dose-response relations for the nicotine-induced absorbance increase and fluorescence increase, suggesting that the absorbance increase leads to the fluorescence increase via the previously described nicotine-induced conformational change, the "candle snuffer" mechanism. Molecular dynamics (MD) simulations identified a binding pose for nicotine, previously indeterminate from experimental data. MD simulations also showed that Helix 4 of the periplasmic binding protein (PBP) domain appears tilted in iNicSnFR12 relative to iNicSnFR3a, likely altering allosteric network(s) that link the ligand binding site to the fluorophore. In thermal melt experiments, nicotine stabilized the PBP of the tested iNicSnFR variants. iNicSnFR12 resolved nicotine in diluted mouse and human serum at 100 nM, the peak [nicotine] that occurs during smoking or vaping, and possibly at the decreasing levels during intervals between sessions. NicSnFR12 was also partially activated by unidentified endogenous ligand(s) in biofluids. Improved iNicSnFR12 variants could become the molecular sensors in continuous nicotine monitors for animal and human biofluids.

bioengineering↗

bioRxiv2023

Integration of spatial and single-cell data across modalities with weak linkage

single-cell sequencing methods have enabled the profiling of multiple types of molecular readouts at cellular resolution, and recent developments in spatial barcoding, in situ hybridization, and in situ sequencing allow such molecular readouts to retain their spatial context. Since no technology can provide complete characterization across all layers of biological modalities within the same cell, there is pervasive need for computational cross-modal integration (also called diagonal integration) of single-cell and spatial omics data. For current methods, the feasibility of cross-modal integration relies on the existence of highly correlated, a priori "linked" features. When such linked features are few or uninformative, a scenario that we call "weak linkage", existing methods fail. We developed MaxFuse, a cross-modal data integration method that, through iterative co-embedding, data smoothing, and cell matching, leverages all information in each modality to obtain high-quality integration. MaxFuse is modality-agnostic and, through comprehensive benchmarks on single-cell and spatial ground-truth multiome datasets, demonstrates high robustness and accuracy in the weak linkage scenario. A prototypical example of weak linkage is the integration of spatial proteomic data with single-cell sequencing data. On two example analyses of this type, we demonstrate how MaxFuse enables the spatial consolidation of proteomic, transcriptomic and epigenomic information at single-cell resolution on the same tissue section.

bioinformatics↗

bioRxiv2023

A new label-free optical imaging method for the lymphatic system enhanced by deep learning

Our understanding of the lymphatic vascular system lags far behind that of the blood vascular system, limited by available imaging technologies. We present a label-free optical imaging method that visualizes the lymphatic system with high contrast. We developed an orthogonal polarization imaging (OPI) in the shortwave infrared range (SWIR) and imaged both lymph nodes and lymphatic vessels of mice and rats in vivo through intact skin, as well as human mesenteric lymph nodes in colectomy specimens. By integrating SWIR-OPI with U-Net, a deep learning image segmentation algorithm, we automated the lymph node size measurement process. Changes in lymph nodes in response to cancer progression were monitored in two separate mouse cancer models, through which we obtained insights into pre-metastatic niches and correlation between lymph node masses and many important biomarkers. In a human pilot study, we demonstrated the effectiveness of SWIR-OPI to detect human lymph nodes in real time with clinical colectomy specimens. One Sentence SummaryWe develop a real-time high contrast optical technique for imaging the lymphatic system, and apply it to anatomical pathology gross examination in a clinical setting, as well as real-time monitoring of tumor microenvironment in animal studies.

bioengineering↗

bioRxiv2023

ASTK: a software for comprehensive analysis of alternative splicing

Alternative splicing (AS) is a fundamental mechanism that regulates gene expression. Splicing dynamics is involved in both physiological and pathological processes. In this paper, we introduce ASTK, a software package covering upstream and downstream analysis of AS. Initially, ASTK offers a module to perform enrichment analysis at both the gene- and exon-level to incorporate various impacts by different spliced events on a single gene. We further cluster AS genes and alternative exons into three groups based on spliced exon sizes (micro-, mid-, and macro-), which are preferentially associated with distinct biological pathways. A major challenge in the field has been decoding the regulatory codes of splicing. ASTK adeptly extracts both sequence features and epigenetic marks associated with AS events. Through the application of machine learning algorithms, we identified pivotal features influencing the inclusion levels of most AS types. Notably, the splice site strength is a primary determinant for the inclusion levels in alternative 3/5 splice sites (A3/A5). For the alternative first exon (AF) and skipping exon (SE) classes, a combination of sequence and epigenetic features collaboratively dictate exon inclusion/exclusion. Our findings underscore ASTKs capability to enhance the functional understanding of AS events and shed light on the intricacies of splicing regulation.

bioinformatics↗

bioRxiv2022

METTL3 maintains epithelial homeostasis through m6A-dependent regulation of chromatin modifiers

The balance between epithelial stemness and differentiation requires the precise regulation of gene expression programs. Epitranscriptomic RNA modifications have been implicated in both epithelial development as well as cancers. However, the underlying mechanisms are poorly understood. Here, we show that deletion of the m6A methyltransferase, METTL3, impairs the m6A-mediated degradation of numerous mRNA transcripts encoding critical chromatin modifying enzymes, resulting in widespread gene expression abnormalities as well as both aberrant cutaneous and oral epithelial phenotypes in vivo. Collectively, these results offer new insights into a new layer of gene regulation within epithelial surface tissues and will inform future epitranscriptomic studies within epithelial cancer and developmental biology.

molecular biology↗

bioRxiv2022

Identification of a key genetic factor governing arabinan utilization in the gut microbiome suggests a novel therapeutic target for constipation

Probiotics have been widely used to improve impaired gastro-intestinal motility, yet their efficacy varied substantially across strains. Here, by a large-scale genetic screen plus in vivo measurements, we identified a key genetic factor (abfA cluster governing arabinan utilization) in probiotic Bifidobacterium longum harnessing the treatment efficacy against functional constipation (FC). Intriguingly, it also presents in a range of gut resident microbiota and played a protective role against FC. Next, our longitudinal multi-omics study in humans revealed that the exogenous abfA-cluster- carrying B. longum can well establish itself in the gut, and enrich arabinan-utilization residents and beneficial metabolites (e.g., acetate, butyrate, chenodeoxycholic acid and uracil). Finally, transplantation of abfA-cluster-enriched human microbiota to FC- induced germ-free mice recapitulated the marked gut-motility improvement and elevated production of beneficial metabolites. Collectively, our proof-of-concept study actively demonstrated a critical yet underexplored role of microbial abfA cluster in ameliorating FC, establishing generalizable principles for developing functional-genomics-directed probiotic therapies.

microbiology↗

bioRxiv2022

Discovering Haematoma-Stimulated Circuits for Secondary Brain Injury after Intraventricular Haemorrhage by Spatial Transcriptome Analysis

Central nervous system (CNS) diseases, such as neurodegenerative disorders and brain diseases caused by acute injuries, are important yet challenging to study due to disease leisions locations and other complexities. Utilizing the powerful spatial transcriptome analysis together with novel algorithms we developed for the study, we report here for the first time a 3D trajectory map of gene expression changes in the brain following acute neural injury using a mouse model of intraventricular haemorrhage (IVH). IVH is a common and representative complication after various acute brain injuries with severe mortality and mobility implications. Our data identified three main 3D global pseudospace-time trajectory bundles which represent the main neural circuits from the lateral ventricle to the hippocampus and primary cortex induced by experimental intraventricular haematoma stimulation. Further analysis indicated a rapid response in the primary cortex, as well as a direct and integrated effect on the hippocampus after IVH. These results are informative in understanding the pathophysiological changes, including the spatial and temporal patterns of gene expression changes, in IVH patients after acute brain injuries and strategizing more effective clinical management regimens. The bioinformatics strategies will also be useful for the study of other CNS diseases.

bioinformatics↗

bioRxiv2022

Gene Set Scoring on the Nearest Neighbor Graph (gssnng) for Single Cell RNA-seq (scRNA-seq)

Gene set scoring (or enrichment) is a common dimension reduction task in bioinformatics that can be focused on differences between groups or at the single sample level. Gene sets can represent biological functions, molecular pathways, cell identities, and more. Gene set scores are context dependent values that are useful for interpreting biological changes following experiments or perturbations. Single sample scoring produces a set of scores, one for each member of a group, which can be analyzed with statistical models that can include additional clinically important factors such as gender or age. However, the sparsity and technical noise of single cell expression measures create difficulties for these methods, which were originally designed for bulk expression profiling (microarrays, RNAseq). This can be greatly remedied by first applying a smoothing transformation that shares gene measure information within transcriptomic neighborhoods. In this work, we use the nearest neighbor graph of cells for matrix smoothing to produce high quality gene set scores on a per-cell, per-group, level which is useful for visualization and statistical analysis. Availability and implementationThe gssnng software is available using the python package index (PyPI) and works with Scanpy AnnData objects. It can be installed using pip install gssnng. More information and demo notebooks: See https://github.com/IlyaLab/gssnng

bioinformatics↗

bioRxiv2022

Field testing of an enzymatic quorum quencher coating additive to reduce biocorrosion of steel

Microbial colonization can be detrimental to the integrity of metal surfaces and lead to microbiologically influenced corrosion (MIC). Biocorrosion is a serious problem for aquatic and marine industries in the world and severely affects the maritime transportation industry by destroying port infrastructure and increasing fuel usage and the time and cost required for maintenance of transport vessels. Here, we evaluate the potential of a stable quorum quenching lactonase enzyme to reduce biocorrosion in the field. Over the course of 21 months, steel samples coated with lactonase-containing acrylic paint were submerged at two different sites and depths in the Duluth Superior Harbor (DSH; Lake Superior, MN, USA) and benchmarked against controls, including the biological biocide surfactin. In this experiment, the lactonase treatment outperformed the surfactin biocide treatment and significantly reduced the number of corrosion tubercles (37% (p<0.01)) and the corroded surface area (39% (p<0.01)) as compared to the acrylic-coated control coupons. In an attempt to evaluate the effects of signal disruption of surface microbial communities and the reasons for lower corrosion levels, 16S rRNA sequencing was performed and community populations were analyzed. Interestingly, surface communities were similar between all treatments, and only minor changes could be observed. Among these changes, several groups including sulfate-reducing bacteria (SRBs) appeared to correlate with corrosion levels, and more specifically, SRBs abundance levels were lower on lactonase treated steel coupons. We surmise that these minute community changes may have large impacts on corrosion rates. Overall, these results highlight the potential use of stable quorum quenching lactonases as an eco-friendly antifouling coating additive.

microbiology↗

bioRxiv2022

The GxcM-Fbp17/RacC-WASP signaling cascade regulates polarized cortex assembly in migrating cells

The actin-rich cortex plays a fundamental role in many cellular processes. Its architecture and molecular composition vary across cell types and physiological states. The full complement of actin assembly factors driving cortex formation and how their activities are spatiotemporally regulated remain to be fully elucidated. Using Dictyostelium as a model for polarized and rapidly migrating cells, we show that GxcM, a RhoGEF localized specifically in the rear of migrating cells, functions together with F-BAR protein Fbp17, a small GTPase RacC, and the actin nucleation-promoting factor WASP to coordinately promote Arp2/3 complex-mediated cortical actin assembly. Over-activation of this signaling cascade leads to excessive actin polymerization in the rear cortex, whereas its disruption causes defects in cortical integrity and function. Therefore, different from its well-defined role in the formation of the front protrusions, the Arp2/3 complex-based actin carries out a previously unappreciated function in building the rear cortical subcompartment in rapidly migrating cells.

cell biology↗