Biology subjects

Nguyen, H.

Publications and source records attributed to Nguyen, H..

137 records · Page 8Linked to original sources

bioRxiv2019

Fast and precise single-cell data analysis using hierarchical autoencoder

A primary challenge in single-cell RNA sequencing (scRNA-seq) studies comes from the massive amount of data and the excess noise level. To address this challenge, we introduce a hierarchical autoencoder that reliably extracts representative information of each cell. In an extensive analysis, we demonstrate that the approach vastly outperforms state-of-the-art techniques in many research sub-fields of scRNA-seq analysis, including cell segregation through unsupervised learning, visualization of transcriptome landscape, cell classification, and pseudo-time inference.

bioinformatics↗

bioRxiv2019

Improved chemistry restraints for crystallographic refinement by integrating the Amber force field into Phenix

The refinement of biomolecular crystallographic models relies on geometric restraints to help address the paucity of experimental data typical in these experiments. Limitations in these restraints can degrade the quality of the resulting atomic models. Here we present an integration of the full all-atom Amber molecular dynamics force field into Phenix crystallographic refinement, which enables a more complete modeling of biomolecular chemistry. The advantages of the force field include a carefully derived set of torsion angle potentials, an extensive and flexible set of atom types, Lennard-Jones treatment of non-bonded interactions and a full treatment of crystalline electrostatics. The new combined method was tested against conventional geometry restraints for over twenty-two thousand protein structures. Structures refined with the new method show substantially improved model quality. On average, Ramachandran and rotamer scores are somewhat better; clash scores and MolProbity scores are significantly improved; and the modelling of electrostatics leads to structures that exhibit more, and more correct, hydrogen bonds than those refined with traditional geometry restraints. We find in general that model improvements are greatest at lower resolutions, prompting plans to add the Amber target function to real-space refinement for use in electron cryo-microscopy. This work opens the door to the future development of more advanced applications such as Amber-based ensemble refinement, quantum mechanical representation of active sites and improved geometric restraints for simulated annealing.\n\nIMPORTANTthis document contains embedded data - to preserve data integrity, please ensure where possible that the IUCr Word tools (available from http://journals.iucr.org/services/docxtemplate/) are installed when editing this document.\n\nSynopsisThe full Amber force field has been integrated into Phenix as an alternative refinement target. With a slight loss in speed, it achieves improved stereochemistry, fewer steric clashes and better hydrogen bonds.

molecular biology↗

bioRxiv2019

VIP-producing enteric neurons interact with innate lymphoid cells to regulate feeding-dependent intestinal epithelial barrier functions

The intestinal mucosa serves as both a conduit for uptake of food-derived nutrients and microbiome-derived metabolites and as a barrier that prevents tissue invasion by microbes and tempers inflammatory responses to the myriad contents of the lumen. How the intestine coordinates physiological and immune responses to food consumption to optimize nutrient uptake while maintaining barrier functions remains unclear. Here, we describe how a gut neuronal signal triggered by food intake is integrated with intestinal antimicrobial and metabolic responses controlled by type 3 innate lymphoid cells (ILC3)1-3. Food consumption rapidly activates a population of enteric neurons that express vasoactive intestinal peptide (VIP)4. Projections of VIP-producing neurons (VIPergic neurons) in the lamina propria are in close proximity to clusters of ILC3 that selectively express VIP receptor type 2 (VIPR2 or VPAC2). ILC3 production of IL-22, which is up-regulated by commensal microbes such as segmented filamentous bacteria (SFB)5-7, is inhibited upon engagement of VIPR2. As a consequence, there is a reduction in epithelial cell-derived antimicrobial peptide, but enhanced expression of lipid-binding proteins and transporters8. During food consumption, activation of VIPergic neurons thus enhances growth of epithelial-associated SFB and increases lipid absorption. Our results reveal a feeding- and circadian-regulated dynamic intestinal neuro-immune circuit that promotes a trade-off between IL-22-mediated innate immune protection and efficiency of nutrient absorption. Modulation of this pathway may hence be effective for enhancing resistance to enteropathogen2,3,9 and for treatment of metabolic diseases.

immunology↗

bioRxiv2019

Mapping of transgenic alleles in plants using a Nanopore-based sequencing strategy

Transgenic technology was developed to introduce transgenes into various organisms to validate gene function and add genetic variation for the development of beneficial input or output trait over 40 years ago. However, the identification of the transgene insertion position in the genome, while doable, can be cumbersome in the organisms with complex genomes. Here, we report a Nanopore-based sequencing method to rapidly map transgenic alleles in the soybean genome. This strategy is high-throughput, convenient, reliable, and cost-efficient. The transgenic allele mapping protocol outlined herein can be easily translated to other higher eukaryotes with complex genomes.

plant biology↗

bioRxiv2019

A contractile acto-myosin belt promotes growth anisotropy during the early stages of pectoral fin development in zebrafish.

The zebrafish pectoral fin is an in vivo model for vertebrate limb formation, well suited to investigate the integration of molecular and cellular dynamics, the results of which translate into shaping the limb bud. We used the ratio between the lengths of the anterior-posterior (AP) and dorso-ventral (DV) axes as the descriptor of how fin shape changes over time. We showed that fin shape transitions from close to hemi-spherical (ratio 1. 36 {+/-} 0.11) to semi-ellipsoid (ratio 1.64 {+/-} 0.04) between 33 and 46 hours post fertilization (hpf). This shape transition coincided with the formation of a contractile \"actin belt\" at the distal rim of the fin bud along its AP axis. The actin belt emerged from a central position and expanded on both sides along the distal rim of the fin, thus marking the DV boundary between two rows of ectodermal cells. Formation of the actin belt depended on Rac protein activity, as suggested by FRET measurements using a Rac biosensor. 3D+time imaging of the developing fin in Rac-deficient embryos showed that anisotropic growth of the fin depends on the actin belt. Indeed, actin belt formation was dramatically reduced or even absent in the embryos without proper Rac activity. This correlated with isotropic growth of the fin bud from normal shape at 33 hpf to quasi hemispherical shape with AP/DV ratio ~1 13 hours later, without affecting cell number and overall bud volume. We propose that the formation of a contractile acto-myosin belt is essential to drive the pectoral fins early anisotropic growth.

developmental biology↗

bioRxiv2018

An open-source software analysis package for Microspheres with Ratiometric Barcode Lanthanide Encoding (MRBLEs)

Multiplexed bioassays, in which multiple analytes of interest are probed in parallel within a single small volume, have greatly accelerated the pace of biological discovery. Bead-based multiplexed bioassays have many technical advantages, including near solution-phase kinetics, small sample volume requirements, many within-assay replicates to reduce measurement error, and, for some bead materials, the ability to synthesize analytes directly on beads via solid-phase synthesis. To allow bead-based multiplexing, analytes can be synthesized on spectrally encoded beads with a 1:1 linkage between analyte identity and embedded codes. Bead-bound analyte libraries can then be pooled and incubated with a fluorescently-labeled macromolecule of interest, allowing downstream quantification of interactions between the macromolecule and all analytes simultaneously via imaging alone. Extracting quantitative binding data from these images poses several computational image processing challenges, requiring the ability to identify all beads in each image, quantify bound fluorescent material associated with each bead, and determine their embedded spectral code to reveal analyte identities. Here, we present a novel open-source Python software package (the mrbles analysis package) that provides the necessary tools to: (1) find encoded beads in a bright-field microscopy image; (2) quantify bound fluorescent material associated with bead perimeters; (3) identify embedded ratiometric spectral codes within beads; and (4) return data aggregated by embedded code and for each individual bead. We demonstrate the utility of this package by applying it towards analyzing data generated via multiplexed measurement of calcineurin protein binding to MRBLEs (Microspheres with Ratiometric Barcode Lanthanide Encoding) containing known and mutant binding peptide motifs. We anticipate that this flexible package should be applicable to a wide variety of assays, including simple bead or droplet finding analysis, quantification of binding to non-encoded beads, and analysis of multiplexed assays that use ratiometric, spectrally encoded beads.

bioengineering↗

bioRxiv2018

Widespread separation of the polypyrimidine tract from 3’ AG by G tracts in association with alternative exons in metazoa and plants

At the end of introns, the polypyrimidine tract (Py) is often close to the 3 AG in a consensus (Y)20NCAGgt in humans. Interestingly, we have found that they could also be separated by purine-rich elements including G tracts in thousands of human genes. These regulatory elements between the Py and 3AG (REPA) mainly regulate alternative 3 splice sites (3SS) and intron retention. Here we show their widespread distribution and special properties across kingdoms. The purine-rich 3SS are found in up to about 60% of the introns among more than 1000 species/lineages by whole genome analysis, and up to 18% of these introns contain the REPA G tracts in about 2.4 millions of 3SS in total. In particular, they are significantly enriched over their 3SS and genome backgrounds in metazoa and plants, and highly associated with alternative splicing of genes in diverse functional clusters. They are also highly enriched (3-6 folds) in the canonical as well as aberrantly used 3 splice sites in cancer patients carrying mutations of the branch point factor SF3B1 or the 3AG binding factor U2AF35. Moreover, the REPA G tract-harbouring 3SS have significantly reduced occurrences of branch point (BP) motifs between the -24 and -4 positions, in particular absent from the -7 - -5 positions in several model organisms examined. The more distant branch points are associated with increased occurrences of alternative splicing in human and zebrafish. The branch points, REPA G tracts and associated 3SS motifs appear to have emerged differentially in a phylum- or species-specific way during evolution. Thus, there is widespread separation of the Py and 3AG by REPA G tracts, likely evolved among different species or branches of life. This special 3SS arrangement contributes to the generation of diverse transcript or protein isoforms in biological functions or diseases through alternative or aberrant splicing.

molecular biology↗

bioRxiv2018

Using a robotic fish to investigate individual differences in social responsiveness in the guppy

Responding towards the actions of others is one of the most important behavioral traits whenever animals of the same species interact. Mutual influences among interacting individuals may modulate the social responsiveness seen and thus makes it often difficult to study the level and variation of individuality in responsiveness. Here, biomimetic robots (BRs) that are accepted as conspecifics but controlled by the experimenter can be a useful tool. Studying the interactions of live animals with BRs allows pinpointing the live animals level of responsiveness by removing confounding mutuality. In this paper, we show that live guppies (Poecilia reticulata) exhibit consistent differences among each other in their responsiveness when interacting with a biomimetic fish robot - Robofish - and a live companion. It has been repeatedly suggested that social responsiveness correlates with other individual behavioral traits like risk-taking behavior ( boldness) or activity level. We tested this assumption in a second experiment. Interestingly, our detailed analysis of individual differences in social responsiveness using the Robofish, suggests that responsiveness is an independent trait, not part of a larger behavioral syndrome formed by boldness and activity.

animal behavior and cognition↗

bioRxiv2018

Increased autophagy in ephrinB2 deficient osteocytes is associated with hypermineralized, brittle bones

Mineralized bone forms when collagen-containing osteoid accrues hydroxyapatite crystals. This process has two phases: a rapid initiation (primary mineralization), followed by slower accrual of mineral (secondary mineralization) that continues until that portion of bone is renewed by remodelling. Within the bone matrix is an interconnected network of cells termed osteocytes. These cells are derived from bone-forming osteoblasts. Osteoblast differentiation requires expression of ephrinB2, and we were intrigued about why ephrinB2 continues to be expressed in mature osteocytes. To determine its function in osteocytes, we developed an osteocyte-specific ephrinB2 null mouse and found they exhibited a brittle bone phenotype. This was not caused by a change in bone mass, but by an intrinsic defect in the strength of the bone material. Although the initiation of osteoid mineralization occurred at a normal rate, the process of secondary mineralization was accelerated in these mice. The maturing mineralized bone matrix incorporated mineral and carbonate more rapidly than controls, indicating that osteocytic ephrinB2 suppresses mineral accumulation in bone. No known regulators of mineralization were modified in the bone of these mice. However, RNA sequencing showed differential expression of a group of autophagy-associated genes, and increased autophagic flux was confirmed in ephrinB2 knockdown osteocytes. This indicates that the process of secondary mineralization in bone makes use of autophagic machinery in a manner that is limited by ephrinB2 in osteocytes, and that this process may be disrupted in conditions of bone fragility.

cell biology↗

bioRxiv2017

Visual evidence accumulation behavior in unrestrained mice

A major challenge to studying the neural circuits underlying perceptual decision making has been the limited availability of tools for manipulation of neural activity. In recent years, rodents have emerged as a desirable model for overcoming the technical hurdle. However, mice, which offer abundant genetic tools for circuit manipulation, are underrepresented in perceptual evidence accumulation studies. Here we describe the behavior of mice performing a visual evidence accumulation task similar to one previously used in rats and humans. We found that although mice were capable of achieving similar accuracy levels as rats, differences in accumulation strategy were apparent. To test the engagement of cortex in the visual evidence accumulation task, we optogenetically inhibited activity in the anteromedial (AM) visual area using JAWS. Importantly, light activation biased choices in both injected and uninjected animals. Fortunately, by varying the stimulus-response contingency while holding constant the stimulated hemisphere, we surmounted this obstacle and demonstrated a role for AM in contralateral choices. Taken together, our results argue that mice accumulate visual evidence to guide decisions, an ability that is supported in part by area AM.

neuroscience↗

bioRxiv2017

Multisite Generalizability Of Schizophrenia Diagnosis Classification Based On Functional Brain Connectivity

Our objective was to assess the generalizability, across sites and cognitive contexts, of schizophrenia classification based on functional brain connectivity. We tested different training-test scenarios combining fMRI data from 191 schizophrenia patients and 191 matched healthy controls obtained at 6 scanning sites and under different task conditions. Diagnosis classification accuracy generalized well to a novel site and cognitive context provided data from multiple sites were used for classifier training. By contrast, lower classification accuracy was achieved when data from a single distinct site was used for training. These findings indicate that it is beneficial to use multisite data to train fMRI-based classifiers intended for large-scale use in the clinical realm.

neuroscience↗