Biology subjects

Stern, C. D.

Publications and source records attributed to Stern, C. D..

11 recordsLinked to original sources

bioRxiv2026

Molecular basis of competence for neural induction in the chick embryo

Competence is the capacity of a cell or tissue to respond to a specific inducing signal from a neighbouring tissue, by changing its fate in a specific direction. Neural induction is the process by which the epiblast of the early embryo responds to signals from the organizer (the tip of the primitive streak in amniotes) by forming a neural plate. Here we study why three regions of the early chick embryo lack competence to respond to neural induction by a grafted organizer: the outer anterior area opaca and the posterior area opaca at primitive streak stages (HH3+-4-), and the inner anterior area opaca at head process stage (HH5), in comparison with the competent anterior inner area opaca at HH3+-4-.Molecular analysis of these tissues, and their temporal dynamics following exposure to the organizer, reveals several differences. Among them, increased BMP and decreased ERK signalling characterise the non-competent regions. Inhibition of BMP can restore competence to HH5 epiblast; a combination of BMP-inhibition with ERK-stimulation by FGF8 can confer competence to the outer area opaca, whereas none of these can endow posterior epiblast with competence for neural induction. We conclude that spatiotemporal competence of epiblast for neural induction is regulated by several mechanisms, including extracellular signals.

developmental biology↗

bioRxiv2025

The organizer as a cooperative of signaling cells for neural induction

The "organizer", discovered 100 years ago by Hans Spemann and Hilde Mangold, is a special region of vertebrate embryos at the gastrula stage; it emits signals that can re-direct the fate of neighboring cells to acquire neural plate identity. It is generally imagined as unique population of cells producing one or a few signaling molecules, responsible for neural induction and for patterning the neural plate and the mesoderm. Here we use single cell and tissue transcriptomics to explore the expression of signaling molecules in the node (the amniote organizer). Although all organizer cells express the homeobox gene Goosecoid, node cells show a diversity of transcription factor signatures associated with expression of subsets of many signaling molecules, suggesting distinct cell sub-populations. Using a recently described Gene Regulatory Network (GRN) of 175 transcriptional responses to neural induction, we explore the activities of 22 of these signals and find that some of them regulate the expression of components of the GRN that are not responsive to previously described pathways associated with neural induction. These results suggest that rather than a single, static, homogeneous population, the organizer comprises a diverse collective of specialized cells that emit cooperating signals to instruct receiving neighbors to adopt their new identities. Significance StatementThe Spemann-Mangold organizer is an embryonic region that can induce the formation of a fully patterned nervous system from non-neural embryonic cells. Here we show that it is made up of a diversity of cell populations that emit distinct sets of signals, which cooperate to account for the repertoire of molecular responses in receiving cells. Several of these signals had not previously been associated with neural induction or patterning.

developmental biology↗

bioRxiv2024

Canonical Wnt signalling from the Area Opaca induces and maintains the Marginal Zone in pre-primitive-streak stage chick embryos

In chick embryos prior to primitive streak formation, the outermost extraembryonic region, known as the area opaca (AO), was generally thought to act only by providing nutrients and mechanical support to the embryo. Just internal to the AO is a ring of epiblast called the marginal zone (MZ), separating the former from the inner, area pellucida epiblast. The MZ does not contribute cells to any part of the embryo but is involved in determining the position of primitive streak formation from the inner epiblast. Recently it was discovered that the AO can induce a MZ from area pellucida epiblast. Here we explore the nature of this inductive signal. We find that Wnt8c is highly expressed in the AO, whereas canonical Wnt pathway targets are enriched in the MZ, along with strong nuclear {beta}-catenin localization. Using isolation and recombination experiments combined with gain- and loss-of-function by exogenous chemical modulators of the pathway, we reveal that Wnt signalling is essential for induction and maintenance of the MZ, as well as sufficient to induce MZ properties in area pellucida epiblast. We propose that canonical Wnt signalling is responsible for induction of the marginal zone by the area opaca.

developmental biology↗

bioRxiv2022

Capturing non-local through-bond effects in molecular mechanics force fields: II. Using fractional bond orders to fit torsion parameters

Accurate small molecule force fields are crucial for predicting thermodynamic and kinetic properties of drug-like molecules in biomolecular systems. Torsion parameters, in particular, are essential for determining conformational distribution of molecules. However, they are usually fit to computationally expensive quantum chemical torsion scans and generalize poorly to different chemical environments. Torsion parameters should ideally capture local through-space non-bonded interactions such as 1-4 steric and electrostatics and non-local through-bond effects such as conjugation and hyperconjugation. Non-local through-bond effects are sensitive to remote substituents and are a contributing factor to torsion parameters poor transferability. Here we show that fractional bond orders such as the Wiberg Bond Order (WBO) are sensitive to remote substituents and correctly captures extent of conjugation and hyperconjugation. We show that the relationship between WBO and torsion barrier heights are linear and can therefore serve as a surrogate to QC torsion barriers, and to interpolate torsion force constants. Using this approach we can reduce the number of computationally expensive QC torsion scans needed while maintaining accurate torsion parameters. We demonstrate this approach to a set of substituted benzene rings.

biophysics↗

bioRxiv2021

A novel role for the extraembryonic area opaca in positioning the primitive streak of the early chick embryo

Classical studies have established that the marginal zone, a ring of extraembryonic epiblast immediately surrounding the embryonic epiblast (area pellucida) of the chick embryo is important in setting embryonic polarity by positioning the primitive streak, the site of gastrulation. The more external extraembryonic region (area opaca) was only thought to have nutritive and support functions. Using experimental embryology approaches, this study reveals three separable functions for this outer region: first, juxtaposition of the area opaca directly onto the area pellucida induces a new marginal zone from the latter; this induced domain is entirely posterior in character. Second, ablation and grafting experiments using an isolated anterior half of the blastoderm and pieces of area opaca suggest that the area opaca can influence the polarity of the adjacent marginal zone. Finally, we show that the loss of the ability of such isolated anterior half-embryos to regulate (re-establish polarity spontaneously) at the early primitive streak stage can be rescued by replacing the area opaca by one from a younger stage. These results uncover new roles of chick extraembryonic tissues in early development. Summary statementTwo adjacent extraembryonic tissues, the area opaca and the marginal zone, interact to influence the polarity of the early chick embryo.

developmental biology↗

bioRxiv2021

"Neighborhood watch" model: embryonic epiblast cells assess positional information in relation to their neighbors

In many developing and regenerating systems, tissue pattern is established through gradients of informative morphogens, but we know little about how cells interpret these. Using experimental manipulation of early chick embryos including misexpression of an inducer (VG1 or ACTIVIN) and an inhibitor (BMP4), we test two alternative models for their ability to explain how the site of primitive streak formation is positioned relative to the rest of the embryo. In one model, cells read morphogen concentrations cell-autonomously. In the other, cells sense changes in morphogen status relative to their neighbourhood. We find that only the latter model can account for the experimental results, including some counter-intuitive predictions. This mechanism (which we name "neighbourhood watch" model) illuminates the classic "French Flag Problem" and how positional information is interpreted by a sheet of cells in a large developing system. Summary statementIn a large developing system, the chick embryo before gastrulation, cells interpret gradients of positional signals relative to their neighbours to position the primitive streak, establishing bilateral symmetry.

developmental biology↗

bioRxiv2021

A gene regulatory network for neural induction

During early vertebrate development, signals from a special region of the embryo, the organizer, can re-direct the fate of non-neural ectoderm cells to form a complete, patterned nervous system. This is called neural induction and has generally been imagined as a single signalling event, causing a switch of fate. Here we undertake a comprehensive analysis, in very fine time-course, of the events following exposure of ectoderm to the organizer. Using transcriptomics and epigenomics we generate a Gene Regulatory Network comprising 175 transcriptional regulators and 5,614 predicted interactions between them, with fine temporal dynamics from initial exposure to the signals to expression of mature neural plate markers. Using in situ hybridization, single-cell RNA-sequencing and reporter assays we show that neural induction by a grafted organizer closely resembles normal neural plate development. The study is accompanied by a comprehensive resource including information about conservation of the predicted enhancers in different vertebrate systems.

developmental biology↗

bioRxiv2020

The embryonic node functions as an instructive stem cell niche

In warm-blooded vertebrate embryos (mammals and birds), the body forms from a growth zone at the tail end. Hensens node, a region which induces and patterns the neural axis is located within this growth zone. The node also contains the precursors of neural, mesodermal and endodermal structures along the midline and has been suggested to contain a small population of resident stem cells. However, it is unknown whether the rest of the node constitutes an instructive stem cell niche, specifying stem cell behaviour. Here we combine transplantation of a single cell in vivo with single-cell mRNA sequencing in the chick and show that when made to enter the node, non-node-progenitor cells become resident and gain stem cell behaviour. These cells preferentially express G2/M phase cell-cycle related genes and are concentrated in posterior sub-regions of the node. The posterior part of the node therefore behaves as an instructive stem cell niche. These results demonstrate a new function for the vertebrate node during development.

developmental biology↗

bioRxiv2020

Capturing non-local through-bond effects when fragmenting molecules for quantum chemical torsion scans

Accurate molecular mechanics force fields for small molecules are essential for predicting protein-ligand binding affinities in drug discovery and understanding the biophysics of biomolecular systems. Torsion potentials derived from quantum chemical (QC) calculations are critical for determining the conformational distributions of small molecules, but are computationally expensive and scale poorly with molecular size. To reduce computational cost and avoid the complications of distal through-space intramolecular interactions, molecules are generally fragmented into smaller entities to carry out QC torsion scans. However, torsion potentials, particularly for conjugated bonds, can be strongly affected by through-bond chemistry distal to the torsion it-self. Poor fragmentation schemes have the potential to significantly disrupt electronic properties in the region around the torsion by removing important, distal chemistries, leading to poor representation of the parent molecules chemical environment and the resulting torsion energy profile. Here we show that a rapidly computable quantity, the fractional Wiberg bond order (WBO), is a sensitive reporter on whether the chemical environment around a torsion has been disrupted. We show that the WBO can be used as a surrogate to assess the robustness of fragmentation schemes and identify conjugated bond sets. We use this concept to construct a validation set by exhaustively fragmenting a set of druglike organic molecules and examine their corresponding WBO distributions derived from accessible conformations that can be used to evaluate fragmentation schemes. To illustrate the utility of the WBO in assessing fragmentation schemes that preserve the chemical environment, we propose a new fragmentation scheme that uses rapidly-computable AM1 WBOs, which are available essentially for free as part of standard AM1-BCC partial charge assignment. This approach can simultaneously maximize the chemical equivalency of the fragment and the substructure in the larger molecule while minimizing fragment size to accelerate QC torsion potential computation for small molecules and reducing undesired through-space steric interactions.

biophysics↗

bioRxiv2019

A Mechanical Model of Early Somite Segmentation

The clock-and-wavefront model (CW) hypothesizes that the formation of somites in vertebrate embryos results from the interplay of molecular oscillations with a wave traveling along the body axis. This model however does not explain how molecular information is interpreted by cells to modulate their rearrangement into somites. Here we performed Scanning Electron Microscopy (SEM) on the pre-somitic mesoderm (PSM) of chicken embryos at stages 11-12 to describe in detail the cell shape changes occurring along the axis of the PSM. This reveals a wave of epithelialization of the dorsal PSM that precedes somite segmentation. Signs of spatially periodic apical constriction appear in this layer starting at least 3-4 somite lengths caudal to the most recently formed somite. The sizes of these clusters correspond to the typical diameter of chicken somites. We propose that a mechanical instability process leads to the separation of cells into these structures and positions the future inter-somite boundaries. We present a model in which a wave of apical constriction leads to increasing tension and periodic failure of adhesion junctions within the dorsal epithelial layer of the PSM, thus positioning somite boundaries. This model can produce spatially periodic segments whose size depends on the speed of the contraction wave (W) and the rate of increase of apical contractility ({Lambda}). The {Lambda}/W ratio determines whether this mechanism produces spatially and temporally regular or irregular segments, and whether segment sizes increase with the wave speed (scaling) as in the CW model. We discuss the limitations of a purely mechanical model of somite segmentation and the role of biomechanics along with CW during somitogenesis.

developmental biology↗

bioRxiv2019

A 3D molecular atlas of the chick embryonic heart

We present a detailed analysis of gene expression in the 2-day (HH12) embryonic chick heart. RNA-seq of 13 micro-dissected regions reveals regionalised expression of 15,570 genes. Of these, 132 were studied by in situ hybridisation and a subset (38 genes) was mapped by Optical Projection Tomography or serial sectioning to build a detailed 3-dimensional atlas of expression. We display this with a novel interactive 3-D viewer and as stacks of sections, revealing the boundaries of expression domains and regions of overlap. Analysis of the expression domains also defines some sub-regions distinct from those normally recognised by anatomical criteria at this stage of development, such as a previously undescribed subdivision of the atria into two orthogonal sets of domains (dorsoventral and left-right). We also include a detailed comparison of expression in the chick with the mouse and other species.

developmental biology↗