Perturbation analysis analyzed—mathematical modeling of intact and perturbed gene regulatory circuits for animal development

Gene regulatory networks for animal development are the underlying mechanisms controlling cell fate specification and differentiation. The architecture of gene regulatory circuits determines their information processing properties and their developmental function. It is a major task to derive realistic network models from exceedingly advanced high throughput experimental data. Here we use mathematical modeling to study the dynamics of gene regulatory circuits to advance the ability to infer regulatory connections and logic function from experimental data. This study is guided by experimental methodologies that are commonly used to study gene regulatory networks that control cell fate specification. We study the effect of a perturbation of an input on the level of its downstream genes and compare between the cis-regulatory execution of OR and AND logics. Circuits that initiate gene activation and circuits that lock on the expression of genes are analyzed. The model improves our ability to analyze experimental data and construct from it the network topology. The model also illuminates information processing properties of gene regulatory circuits for animal development

The conserved role and divergent regulation of foxa, a pan-eumetazoan developmental regulatory gene

Foxa is a forkhead transcription factor that is expressed in the endoderm lineage across metazoans. Orthologs of foxa are expressed in cells that intercalate, polarize, and form tight junctions in the digestive tracts of the mouse, the sea urchin, and the nematode and in the chordate notochord. The loss of foxa expression eliminates these morphogenetic processes. The remarkable similarity in foxa phenotypes in these diverse organisms raises the following questions: why is the developmental role of Foxa so highly conserved? Is foxa transcriptional regulation as conserved as its developmental role? Comparison of the regulation of foxa orthologs in sea urchin and in Caenorhabditis elegans shows that foxa transcriptional regulation has diverged significantly between these two organisms, particularly in the cells that contribute to the C. elegans pharynx formation. We suggest that the similarity of foxa phenotype is due to its role in an ancestral gene regulatory network that controlled intercalation followed by mesenchymal-to-epithelial transition. foxa transcriptional regulation had evolved to support the developmental program in each species so foxa would play its role controlling morphogenesis at the necessary embryonic address

Experimentally based sea urchin gene regulatory network and the causal explanation of developmental phenomenology

Gene regulatory networks (GRNs) for development underlie cell fate specification and differentiation. Network topology, logic, and dynamics can be obtained by thorough experimental analysis. Our understanding of the GRN controlling endomesoderm specification in the sea urchin embryo has attained an advanced level such that it explains developmental phenomenology.Here we review how the network explains the mechanisms utilized in development to control the formation of dynamic expression patterns of transcription factors and signaling molecules. The network represents the genomic program controlling timely activation of specification and differentiation genes in the correct embryonic lineages. It can also be used to study evolution of body plans. We demonstrate how comparing the sea urchin GRN to that of the sea star and to that of later developmental stages in the sea urchin, reveals mechanisms underlying the origin of evolutionary novelty. The experimentally based GRN for endomesoderm specification in the sea urchin embryo provides unique insights into the system level properties of cell fate specification and its evolution

Comparative Studies of Gene Expression Kinetics: Methodologies and Insights on Development and Evolution

Across the animal kingdom, embryos of closely related species show high morphological similarity despite genetic and environmental distances. Deciphering the molecular mechanisms that underlie morphological conservation and those that support embryonic adaptation are keys to understand developmental robustness and evolution. Comparative studies of developmental gene regulatory networks can track the genetic changes that lead to evolutionary novelties. However, these studies are limited to a relatively small set of genes and demand extensive experimental efforts. An alternative approach enabled by next-generation sequencing, is to compare the expression kinetic of large sets of genes between different species. The advantages of these comparisons are that they can be done relatively easily, for any species and they provide information of all expressed genes. The challenge in these experiments is to compare the kinetic profiles of thousands of genes between species that develop in different rates. Here we review recent comparative studies that tackled the challenges of accurate staging and large-scale analyses using different computational approaches. These studies reveal how correct temporal scaling exposes the striking conservation of developmental gene expression between morphologically similar species. Different clustering approaches are used to address various comparative questions and identify the conservation and divergence of large gene sets. We discuss the unexpected contribution of housekeeping genes to the interspecies correlations and how this contribution distorts the hourglass pattern generated by developmental genes. Overall, we demonstrate how comparative studies of gene expression kinetics can provide novel insights into the developmental constraints and plasticity that shape animal body plans

Gene regulatory control in the sea urchin aboral ectoderm: Spatial initiation, signaling inputs,and cell fate lockdown

The regulation of oral–aboral ectoderm specification in the sea urchin embryo has been extensively studied in recent years. The oral–aboral polarity is initially imposed downstream of a redox gradient induced by asymmetric maternal distribution of mitochondria. Two TGF-β signaling pathways, Nodal and BMP, are then respectively utilized in the generation of oral and aboral regulatory states. However, a causal understanding of the regulation of aboral ectoderm specification has been lacking. In this work control of aboral ectoderm regulatory state specification was revealed by combining detailed regulatory gene expression studies, perturbation and cis-regulatory analyses. Our analysis illuminates a dynamic system where different factors dominate at different developmental times. We found that the initial activation of aboral genes depends directly on the redox sensitive transcription factor, hypoxia inducible factor 1α (HIF-1α). Two BMP ligands, BMP2/4 and BMP5/8, then significantly enhance aboral regulatory gene transcription. Ultimately, encoded feedback wiring lockdown the aboral ectoderm regulatory state. Our study elucidates the different regulatory mechanisms that sequentially dominate the spatial localization of aboral regulatory states

Analysis of the P. lividus sea urchin genome highlights contrasting trends of genomic and regulatory evolution in deuterostomes

Sea urchins are emblematic models in developmental biology and display several characteristics that set them apart from other deuterostomes. To uncover the genomic cues that may underlie these specificities, we generated a chromosome-scale genome assembly for the sea urchin Paracentrotus lividus and an extensive gene expression and epigenetic profiles of its embryonic development. We found that, unlike vertebrates, sea urchins retained ancestral chromosomal linkages but underwent very fast intrachromosomal gene order mixing. We identified a burst of gene duplication in the echinoid lineage and showed that some of these expanded genes have been recruited in novel structures (water vascular system, Aristotle's lantern, and skeletogenic micromere lineage). Finally, we identified gene-regulatory modules conserved between sea urchins and chordates. Our results suggest that gene-regulatory networks controlling development can be conserved despite extensive gene order rearrangement

The Evolution of Biomineralization through the Co-Option of Organic Scaffold Forming Networks

Biomineralization is the process in which organisms use minerals to generate hard structures like teeth, skeletons and shells. Biomineralization is proposed to have evolved independently in different phyla through the co-option of pre-existing developmental programs. Comparing the gene regulatory networks (GRNs) that drive biomineralization in different species could illuminate the molecular evolution of biomineralization. Skeletogenesis in the sea urchin embryo was extensively studied and the underlying GRN shows high conservation within echinoderms, larval and adult skeletogenesis. The organic scaffold in which the calcite skeletal elements form in echinoderms is a tubular compartment generated by the syncytial skeletogenic cells. This is strictly different than the organic cartilaginous scaffold that vertebrates mineralize with hydroxyapatite to make their bones. Here I compare the GRNs that drive biomineralization and tubulogenesis in echinoderms and in vertebrates. The GRN that drives skeletogenesis in the sea urchin embryo shows little similarity to the GRN that drives bone formation and high resemblance to the GRN that drives vertebrates’ vascular tubulogenesis. On the other hand, vertebrates’ bone-GRNs show high similarity to the GRNs that operate in the cells that generate the cartilage-like tissues of basal chordate and invertebrates that do not produce mineralized tissue. These comparisons suggest that biomineralization in deuterostomes evolved through the phylum specific co-option of GRNs that control distinct organic scaffolds to mineralization

<i>Sp</i> and <i>Pl</i> development.

<p><b>A,</b> Comparison of <i>Pl</i> (top) and <i>Sp</i> (bottom) embryo development up to prism stage. <b>B,</b> Schematic diagrams of sea urchin cell lineages [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005435#pgen.1005435.ref028" target="_blank">28</a>]. Red–skeletogenic mesoderm, light purple–non skeletogenic mesoderm, blue–endoderm, yellow–oral ectoderm, green–aboral ectoderm, dark purple–small micromeres.</p

Comparative Study of Regulatory Circuits in Two Sea Urchin Species Reveals Tight Control of Timing and High Conservation of Expression Dynamics

<div><p>Accurate temporal control of gene expression is essential for normal development and must be robust to natural genetic and environmental variation. Studying gene expression variation within and between related species can delineate the level of expression variability that development can tolerate. Here we exploit the comprehensive model of sea urchin gene regulatory networks and generate high-density expression profiles of key regulatory genes of the Mediterranean sea urchin, <i>Paracentrotus lividus</i> (<i>Pl</i>). The high resolution of our studies reveals highly reproducible gene initiation times that have lower variation than those of maximal mRNA levels between different individuals of the same species. This observation supports a threshold behavior of gene activation that is less sensitive to input concentrations. We then compare Mediterranean sea urchin gene expression profiles to those of its Pacific Ocean relative, <i>Strongylocentrotus purpuratus</i> (<i>Sp</i>). These species shared a common ancestor about 40 million years ago and show highly similar embryonic morphologies. Our comparative analyses of five regulatory circuits operating in different embryonic territories reveal a high conservation of the temporal order of gene activation but also some cases of divergence. A linear ratio of 1.3-fold between gene initiation times in <i>Pl</i> and <i>Sp</i> is partially explained by scaling of the developmental rates with temperature. Scaling the developmental rates according to the estimated <i>Sp-Pl</i> ratio and normalizing the expression levels reveals a striking conservation of relative dynamics of gene expression between the species. Overall, our findings demonstrate the ability of biological developmental systems to tightly control the timing of gene activation and relative dynamics and overcome expression noise induced by genetic variation and growth conditions.</p></div