Echinobase: a resource to support the echinoderm research community.

Echinobase (www.echinobase.org) is a model organism knowledgebase serving as a resource for the community that studies echinoderms, a phylum of marine invertebrates that includes sea urchins and sea stars. Echinoderms have been important experimental models for over 100 years and continue to make important contributions to environmental, evolutionary, and developmental studies, including research on developmental gene regulatory networks. As a centralized resource, Echinobase hosts genomes and collects functional genomic data, reagents, literature, and other information for the community. This third-generation site is based on the Xenbase knowledgebase design and utilizes gene-centric pages to minimize the time and effort required to access genomic information. Summary gene pages display gene symbols and names, functional data, links to the JBrowse genome browser, and orthology to other organisms and reagents, and tabs from the Summary gene page contain more detailed information concerning mRNAs, proteins, diseases, and protein-protein interactions. The gene pages also display 1:1 orthologs between the fully supported species Strongylocentrotus purpuratus (purple sea urchin), Lytechinus variegatus (green sea urchin), Patiria miniata (bat star), and Acanthaster planci (crown-of-thorns sea star). JBrowse tracks are available for visualization of functional genomic data from both fully supported species and the partially supported species Anneissia japonica (feather star), Asterias rubens (sugar star), and L. pictus (painted sea urchin). Echinobase serves a vital role by providing researchers with annotated genomes including orthology, functional genomic data aligned to the genomes, and curated reagents and data. The Echinoderm Anatomical Ontology provides a framework for standardizing developmental data across the phylum, and knowledgebase content is formatted to be findable, accessible, interoperable, and reusable by the research community.

Evolutionary analyses of genes in Echinodermata offer insights towards the origin of metazoan phyla.

Despite recent studies discussing the evolutionary impacts of gene duplications and losses among metazoans, the genomic basis for the evolution of phyla remains enigmatic. Here, we employ phylogenomic approaches to search for orthologous genes without known functions among echinoderms, and subsequently use them to guide the identification of their homologs across other metazoans. Our final set of 14 genes was obtained via a suite of homology prediction tools, gene expression data, gene ontology, and generating the Strongylocentrotus purpuratus phylome. The gene set was subjected to selection pressure analyses, which indicated that they are highly conserved and under negative selection. Their presence across broad taxonomic depths suggests that genes required to form a phylum are ancestral to that phylum. Therefore, rather than de novo gene genesis, we posit that evolutionary forces such as selection on existing genomic elements over large timescales may drive divergence and contribute to the emergence of phyla.

Regeneration of the larval sea star nervous system by wounding induced respecification to the Sox2 lineage.

The ability to restore lost body parts following traumatic injury is a fascinating area of biology that challenges current understanding of the ontogeny of differentiation. The origin of new cells needed to regenerate lost tissue, and whether they are pluripotent or have de- or trans-differentiated, remains one of the most important open questions . Additionally, it is not known whether developmental gene regulatory networks are reused or whether regeneration specific networks are deployed. Echinoderms, including sea stars, have extensive ability for regeneration, however, the technologies for obtaining transgenic echinoderms are limited and tracking cells involved in regeneration, and thus identifying the cellular sources and potencies has proven challenging. In this study, we develop new transgenic tools to follow the fate of populations of cells in the regenerating larva of the sea star Patiria miniata. We show that the larval serotonergic nervous system can regenerate following decapitation. Using a BAC-transgenesis approach we show that expression of the pan ectodermal marker, sox2, is induced in previously sox2 minus cells , even when cell division is inhibited. sox2+ cells give rise to new sox4+ neural precursors that then proceed along an embryonic neurogenesis pathway to reform the anterior nervous systems. sox2+ cells contribute to only neural and ectoderm lineages, indicating that these progenitors maintain their normal, embryonic lineage restriction. This indicates that sea star larval regeneration uses a combination of existing lineage restricted stem cells, as well as respecification of cells into neural lineages, and at least partial reuse of developmental GRNs to regenerate their nervous system.

Echinobase: leveraging an extant model organism database to build a knowledgebase supporting research on the genomics and biology of echinoderms.

Echinobase (www.echinobase.org) is a third generation web resource supporting genomic research on echinoderms. The new version was built by cloning the mature Xenopus model organism knowledgebase, Xenbase, refactoring data ingestion pipelines and modifying the user interface to adapt to multispecies echinoderm content. This approach leveraged over 15 years of previous database and web application development to generate a new fully featured informatics resource in a single year. In addition to the software stack, Echinobase uses the private cloud and physical hosts that support Xenbase. Echinobase currently supports six echinoderm species, focused on those used for genomics, developmental biology and gene regulatory network analyses. Over 38 000 gene pages, 18 000 publications, new improved genome assemblies, JBrowse genome browser and BLAST + services are available and supported by the development of a new echinoderm anatomical ontology, uniformly applied formal gene nomenclature, and consistent orthology predictions. A novel feature of Echinobase is integrating support for multiple, disparate species. New genomes from the diverse echinoderm phylum will be added and supported as data becomes available. The common code development design of the integrated knowledgebases ensures parallel improvements as each resource evolves. This approach is widely applicable for developing new model organism informatics resources.

Classifying domain-specific text documents containing ambiguous keywords.

A keyword-based search of comprehensive databases such as PubMed may return irrelevant papers, especially if the keywords are used in multiple fields of study. In such cases, domain experts (curators) need to verify the results and remove the irrelevant articles. Automating this filtering process will save time, but it has to be done well enough to ensure few relevant papers are rejected and few irrelevant papers are accepted. A good solution would be fast, work with the limited amount of data freely available (full paper body may be missing), handle ambiguous keywords and be as domain-neutral as possible. In this paper, we evaluate a number of classification algorithms for identifying a domain-specific set of papers about echinoderm species and show that the resulting tool satisfies most of the abovementioned requirements. Echinoderms consist of a number of very different organisms, including brittle stars, sea stars (starfish), sea urchins and sea cucumbers. While their taxonomic identifiers are specific, the common names are used in many other contexts, creating ambiguity and making a keyword search prone to error. We try classifiers using Linear, Naïve Bayes, Nearest Neighbor, Tree, SVM, Bagging, AdaBoost and Neural Network learning models and compare their performance. We show how effective the resulting classifiers are in filtering irrelevant articles returned from PubMed. The methodology used is more dependent on the good selection of training data and is a practical solution that can be applied to other fields of study facing similar challenges. Database URL: The code and date reported in this paper are freely available at http://xenbaseturbofrog.org/pub/Text-Topic-Classifier/.

A nomenclature for echinoderm genes.

Echinoderm embryos and larvae are prominent experimental model systems for studying developmental mechanisms. High-quality, assembled, annotated genome sequences are now available for several echinoderm species, including representatives from most classes. The increased availability of these data necessitates the development of a nomenclature that assigns universally interpretable gene symbols to echinoderm genes to facilitate cross-species comparisons of gene functions, both within echinoderms and across other phyla. This paper describes the implementation of an improved set of echinoderm gene nomenclature guidelines that both communicates meaningful orthology information in protein-coding gene symbols and names and establishes continuity with nomenclatures developed for major vertebrate model organisms, including humans. Differences between the echinoderm gene nomenclature guidelines and vertebrate guidelines are examined and explained. This nomenclature incorporates novel solutions to allow for several types of orthologous relationships, including the single echinoderm genes with multiple vertebrate co-orthologs that result from whole-genome-duplication events. The current version of the Echinoderm Gene Nomenclature Guidelines can be found at https://www.echinobase.org/gene/static/geneNomenclature.jsp Database URL https://www.echinobase.org/.

Modularity and hierarchy in biological systems: Using gene regulatory networks to understand evolutionary change.

Modularity and hierarchy are important theoretical concepts in biology, and both are useful frameworks to understand the evolution of complex systems. Gene regulatory networks (GRNs) provide a powerful mechanistic model for modularity in animal development, as they are made up of modular (or self-contained) circuits, which are deployed in a hierarchical manner over time. Over the years, studies in the sea urchin, Strongylocentrotus purpuratus, have provided an illustrative example of how these regulatory circuits are responsible for processes such as cell differentiation and cell state specificity. However, GRNs are themselves made up of a nested series of interactions, as each gene can be regulated by multiple cis-regulatory elements, which can be further broken down into distinct transcription factor binding sites (TFBS). As a result, modularity can be applied to each "level" of this complex hierarchy. Throughout the literature, there is considerable discussion about the roles modular circuits, modular enhancers, and modular TFBS play in evolution, yet there is little discussion about how these nested interactions operate as a whole. In this chapter, we discuss how modular changes at different levels of the GRN hierarchy affect animal development and aim to provide a unified framework to understand the role of modularity in evolution.

Systematic comparison of sea urchin and sea star developmental gene regulatory networks explains how novelty is incorporated in early development.

The extensive array of morphological diversity among animal taxa represents the product of millions of years of evolution. Morphology is the output of development, therefore phenotypic evolution arises from changes to the topology of the gene regulatory networks (GRNs) that control the highly coordinated process of embryogenesis. A particular challenge in understanding the origins of animal diversity lies in determining how GRNs incorporate novelty while preserving the overall stability of the network, and hence, embryonic viability. Here we assemble a comprehensive GRN for endomesoderm specification in the sea star from zygote through gastrulation that corresponds to the GRN for sea urchin development of equivalent territories and stages. Comparison of the GRNs identifies how novelty is incorporated in early development. We show how the GRN is resilient to the introduction of a transcription factor, pmar1, the inclusion of which leads to a switch between two stable modes of Delta-Notch signaling. Signaling pathways can function in multiple modes and we propose that GRN changes that lead to switches between modes may be a common evolutionary mechanism for changes in embryogenesis. Our data additionally proposes a model in which evolutionarily conserved network motifs, or kernels, may function throughout development to stabilize these signaling transitions.

Developmental transcriptomes of the sea star, Patiria miniata, illuminate how gene expression changes with evolutionary distance.

Understanding how changes in developmental gene expression alter morphogenesis is a fundamental problem in development and evolution. A promising approach to address this problem is to compare the developmental transcriptomes between related species. The echinoderm phylum consists of several model species that have significantly contributed to the understanding of gene regulation and evolution. Particularly, the regulatory networks of the sea star, Patiria miniata (P. miniata), have been extensively studied, however developmental transcriptomes for this species were lacking. Here we generated developmental transcriptomes of P. miniata and compared these with those of two sea urchins species. We demonstrate that the conservation of gene expression depends on gene function, cell type and evolutionary distance. With increasing evolutionary distance the interspecies correlations in gene expression decreases. The reduction is more severe in the correlations between morphologically equivalent stages (diagonal elements) than in the correlation between morphologically distinct stages (off-diagonal elements). This could reflect a decrease in the morphological constraints compared to other constraints that shape gene expression at large evolutionary divergence. Within this trend, the interspecies correlations of developmental control genes maintain their diagonality at large evolutionary distance, and peak at the onset of gastrulation, supporting the hourglass model of phylotypic stage conservation.

Genomic resources for the study of echinoderm development and evolution.

Echinoderms are important research models for a wide range of biological questions. In particular, echinoderm embryos are exemplary models for dissecting the molecular and cellular processes that drive development and testing how these processes can be modified through evolution to produce the extensive morphological diversity observed in the phylum. Modern attempts to characterize these processes depend on some level of genomic analysis; from querying annotated gene sets to functional genomics experiments to identify candidate cis-regulatory sequences. Given how essential these data have become, it is important that researchers using available datasets or performing their own genome-scale experiments understand the nature and limitations of echinoderm genomic analyses. In this chapter we highlight the current state of echinoderm genomic data and provide methodological considerations for common approaches, including analysis of transcriptome and functional genomics datasets.

Analysis of sea star larval regeneration reveals conserved processes of whole-body regeneration across the metazoa.

BACKGROUND: Metazoan lineages exhibit a wide range of regenerative capabilities that vary among developmental stage and tissue type. The most robust regenerative abilities are apparent in the phyla Cnidaria, Platyhelminthes, and Echinodermata, whose members are capable of whole-body regeneration (WBR). This phenomenon has been well characterized in planarian and hydra models, but the molecular mechanisms of WBR are less established within echinoderms, or any other deuterostome system. Thus, it is not clear to what degree aspects of this regenerative ability are shared among metazoa. RESULTS: We characterize regeneration in the larval stage of the Bat Star (Patiria miniata). Following bisection along the anterior-posterior axis, larvae progress through phases of wound healing and re-proportioning of larval tissues. The overall number of proliferating cells is reduced following bisection, and we find evidence for a re-deployment of genes with known roles in embryonic axial patterning. Following axial respecification, we observe a significant localization of proliferating cells to the wound region. Analyses of transcriptome data highlight the molecular signatures of functions that are common to regeneration, including specific signaling pathways and cell cycle controls. Notably, we find evidence for temporal similarities among orthologous genes involved in regeneration from published Platyhelminth and Cnidarian regeneration datasets. CONCLUSIONS: These analyses show that sea star larval regeneration includes phases of wound response, axis respecification, and wound-proximal proliferation. Commonalities of the overall process of regeneration, as well as gene usage between this deuterostome and other species with divergent evolutionary origins reveal a deep similarity of whole-body regeneration among the metazoa.

EchinoBase: Tools for Echinoderm Genome Analyses.

The echinoderms are a phylum of invertebrate deuterostome animals that constitute important research models for a number of biological disciplines. EchinoBase ( www.echinobase.org ) is an echinoderm-specific genome database and web information system that provides a platform for the interrogation and exploration of echinoderm genomic data. This chapter outlines the datasets available on EchinoBase; from assembled genomes and genome annotations, to spatial and quantitative expression data, as well as functional genomics datasets. We also highlight the bioinformatic tools available on the website to facilitate rapid inquiries using these data (genome browsers, precompiled BLAST databases, etc.), and suggest optimized strategies for performing these inquiries. We conclude with a perspective on how one could integrate various genomic resources to predict putative noncoding regulatory regions. The available datasets and analyses they permit provide the basic components required for developing an understanding of how echinoderm genomes are regulated, especially during early development, and provides a platform for comparative genomic inquiries among species in this phylum.

Embryonic neurogenesis in echinoderms.

The phylogenetic position of echinoderms is well suited to revealing shared features of deuterostomes that distinguish them from other bilaterians. Although echinoderm neurobiology remains understudied, genomic resources, molecular methods, and systems approaches have enabled progress in understanding mechanisms of embryonic neurogenesis. Even though the morphology of echinoderm larvae is diverse, larval nervous systems, which arise during gastrulation, have numerous similarities in their organization. Diverse neural subtypes and specialized sensory neurons have been identified and details of neuroanatomy using neuron-specific labels provide hypotheses for neural function. The early patterning of ectoderm and specification of axes has been well studied in several species and underlying gene regulatory networks have been established. The cells giving rise to central and peripheral neural components have been identified in urchins and sea stars. Neurogenesis includes typical metazoan features of asymmetric division of neural progenitors and in some cases limited proliferation of neural precursors. Delta/Notch signaling has been identified as having critical roles in regulating neural patterning and differentiation. Several transcription factors functioning in pro-neural phases of specification, neural differentiation, and sub-type specification have been identified and structural or functional components of neurons are used as differentiation markers. Several methods for altering expression in embryos have revealed aspects of a regulatory hierarchy of transcription factors in neurogenesis. Interfacing neurogenic gene regulatory networks to the networks regulating ectodermal domains and identifying the spatial and temporal inputs that pattern the larval nervous system is a major challenge that will contribute substantially to our understanding of the evolution of metazoan nervous systems. This article is categorized under: Comparative Development and Evolution > Model Systems Comparative Development and Evolution > Body Plan Evolution Early Embryonic Development > Gastrulation and Neurulation.

Paleogenomics of echinoids reveals an ancient origin for the double-negative specification of micromeres in sea urchins.

Establishing a timeline for the evolution of novelties is a common, unifying goal at the intersection of evolutionary and developmental biology. Analyses of gene regulatory networks (GRNs) provide the ability to understand the underlying genetic and developmental mechanisms responsible for the origin of morphological structures both in the development of an individual and across entire evolutionary lineages. Accurately dating GRN novelties, thereby establishing a timeline for GRN evolution, is necessary to answer questions about the rate at which GRNs and their subcircuits evolve, and to tie their evolution to paleoenvironmental and paleoecological changes. Paleogenomics unites the fossil record and all aspects of deep time, with modern genomics and developmental biology to understand the evolution of genomes in evolutionary time. Recent work on the regulatory genomic basis of development in cidaroid echinoids, sand dollars, heart urchins, and other nonmodel echinoderms provides an ideal dataset with which to explore GRN evolution in a comparative framework. Using divergence time estimation and ancestral state reconstructions, we have determined the age of the double-negative gate (DNG), the subcircuit which specifies micromeres and skeletogenic cells in Strongylocentrotus purpuratus We have determined that the DNG has likely been used for euechinoid echinoid micromere specification since at least the Late Triassic. The innovation of the DNG thus predates the burst of post-Paleozoic echinoid morphological diversification that began in the Early Jurassic. Paleogenomics has wide applicability for the integration of deep time and molecular developmental data, and has wide utility in rigorously establishing timelines for GRN evolution.

Genome-wide use of high- and low-affinity Tbrain transcription factor binding sites during echinoderm development.

Sea stars and sea urchins are model systems for interrogating the types of deep evolutionary changes that have restructured developmental gene regulatory networks (GRNs). Although cis-regulatory DNA evolution is likely the predominant mechanism of change, it was recently shown that Tbrain, a Tbox transcription factor protein, has evolved a changed preference for a low-affinity, secondary binding motif. The primary, high-affinity motif is conserved. To date, however, no genome-wide comparisons have been performed to provide an unbiased assessment of the evolution of GRNs between these taxa, and no study has attempted to determine the interplay between transcription factor binding motif evolution and GRN topology. The study here measures genome-wide binding of Tbrain orthologs by using ChIP-sequencing and associates these orthologs with putative target genes to assess global function. Targets of both factors are enriched for other regulatory genes, although nonoverlapping sets of functional enrichments in the two datasets suggest a much diverged function. The number of low-affinity binding motifs is significantly depressed in sea urchins compared with sea star, but both motif types are associated with genes from a range of functional categories. Only a small fraction (∼10%) of genes are predicted to be orthologous targets. Collectively, these data indicate that Tbr has evolved significantly different developmental roles in these echinoderms and that the targets and the binding motifs in associated cis-regulatory sequences are dispersed throughout the hierarchy of the GRN, rather than being biased toward terminal process or discrete functional blocks, which suggests extensive evolutionary tinkering.

Echinoderm development and evolution in the post-genomic era.

The highly recognizable animals within the phylum Echinodermata encompass an enormous disparity of adult and larval body plans. The extensive knowledge of sea urchin development has culminated in the description of the exquisitely detailed gene regulatory network (GRN) that governs the specification of various embryonic territories. This information provides a unique opportunity for comparative studies in other echinoderm taxa to understand the evolution and developmental mechanisms underlying body plan change. This review focuses on recent work that has utilized new genomic resources and systems-level experiments to address questions of evolutionary developmental biology. In particular, we synthesize the results of several recent studies from various echinoderm classes that have explored the development and evolution of the larval skeleton, which is a major feature that distinguishes the two predominant larval subtypes within the Phylum. We specifically examine the ways in which GRNs can evolve, either through cis regulatory and/or protein-level changes in transcription factors. We also examine recent work comparing evolution across shorter time scales that occur within and between species of sea urchin, and highlight the kinds of questions that can be addressed by these comparisons. The advent of new genomic and transcriptomic datasets in additional species from all classes of echinoderm will continue to empower the use of these taxa for evolutionary developmental studies.

A gene regulatory network for apical organ neurogenesis and its spatial control in sea star embryos.

How neural stem cells generate the correct number and type of differentiated neurons in appropriate places remains an important question. Although nervous systems are diverse across phyla, in many taxa the larva forms an anterior concentration of serotonergic neurons, or apical organ. The sea star embryo initially has a pan-neurogenic ectoderm, but the genetic mechanism that directs a subset of these cells to generate serotonergic neurons in a particular location is unresolved. We show that neurogenesis in sea star larvae begins with soxc-expressing multipotent progenitors. These give rise to restricted progenitors that express lhx2/9 soxc- and lhx2/9-expressing cells can undergo both asymmetric divisions, allowing for progression towards a particular neural fate, and symmetric proliferative divisions. We show that nested concentric domains of gene expression along the anterior-posterior (AP) axis, which are observed in a great diversity of metazoans, control neurogenesis in the sea star larva by promoting particular division modes and progression towards becoming a neuron. This work explains how spatial patterning in the ectoderm controls progression of neurogenesis in addition to providing spatial cues for neuron location. Modification to the sizes of these AP territories provides a simple mechanism to explain the diversity of neuron number among apical organs.

Unraveling the Tangled Skein: The Evolution of Transcriptional Regulatory Networks in Development.

The molecular and genetic basis for the evolution of anatomical diversity is a major question that has inspired evolutionary and developmental biologists for decades. Because morphology takes form during development, a true comprehension of how anatomical structures evolve requires an understanding of the evolutionary events that alter developmental genetic programs. Vast gene regulatory networks (GRNs) that connect transcription factors to their target regulatory sequences control gene expression in time and space and therefore determine the tissue-specific genetic programs that shape morphological structures. In recent years, many new examples have greatly advanced our understanding of the genetic alterations that modify GRNs to generate newly evolved morphologies. Here, we review several aspects of GRN evolution, including their deep preservation, their mechanisms of alteration, and how they originate to generate novel developmental programs.

Evolution of transcription factor function as a mechanism for changing metazoan developmental gene regulatory networks.

The form that an animal takes during development is directed by gene regulatory networks (GRNs). Developmental GRNs interpret maternally deposited molecules and externally supplied signals to direct cell-fate decisions, which ultimately leads to the arrangements of organs and tissues in the organism. Genetically encoded modifications to these networks have generated the wide range of metazoan diversity that exists today. Most studies of GRN evolution focus on changes to cis-regulatory DNA, and it was historically theorized that changes to the transcription factors that bind to these cis-regulatory modules (CRMs) contribute to this process only rarely. A growing body of evidence suggests that changes to the coding regions of transcription factors play a much larger role in the evolution of developmental gene regulatory networks than originally imagined. Just as cis-regulatory changes make use of modular binding site composition and tissue-specific modules to avoid pleiotropy, transcription factor coding regions also predominantly evolve in ways that limit the context of functional effects. Here, we review the recent works that have led to this unexpected change in the field of Evolution and Development (Evo-Devo) and consider the implications these studies have had on our understanding of the evolution of developmental processes.

Dose-dependent nuclear ß-catenin response segregates endomesoderm along the sea star primary axis.

In many invertebrates, the nuclearization of ß-catenin at one pole of the embryo initiates endomesoderm specification. An intriguing possibility is that a gradient of nuclear ß-catenin (nß-catenin), similar to that operating in vertebrate neural tube patterning, functions to distinguish cell fates in invertebrates. To test this hypothesis, we determined the function of nß-catenin during the early development of the sea star, which undergoes a basal deuterostomal mode of embryogenesis. We show that low levels of nß-catenin activity initiate bra, which is expressed in the future posterior endoderm-fated territory; intermediate levels are required for expression of foxa and gata4/5/6, which are later restricted to the endoderm; and activation of ets1 and erg in the mesoderm-fated territory requires the highest nß-catenin activity. Transcription factors acting downstream of high nß-catenin segregate the endoderm/mesoderm boundary, which is further reinforced by Delta/Notch signaling. Significantly, therefore, in sea stars, endomesoderm segregation arises through transcriptional responses to levels of nß-catenin activity. Here, we describe the first empirical evidence of a dose-dependent response to a dynamic spatiotemporal nß-catenin activity that patterns cell fates along the primary axis in an invertebrate.