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Bilateria encompass the vast majority of the animal phyla. As the name states, they are bilaterally symmetric, that is with a morphologically clear main body axis connecting their anterior and posterior ends, a second axis running between their dorsal and ventral surfaces, and with a left side being roughly a mirror image of their right side. Bone morphogenetic protein (BMP) signalling has widely conserved functions in the formation and patterning of the second, dorso-ventral (DV) body axis, albeit to different extents in different bilaterian species. Whilst initial findings in the fruit fly Drosophila and the frog Xenopus highlighted similarities amongst these evolutionarily very distant species, more recent analyses featuring other models revealed considerable diversity in the mechanisms underlying dorsoventral patterning. In fact, as phylogenetic sampling becomes broader, we find that this axis patterning system is so evolvable that even its core components can be deployed differently or lost in different model organisms. In this review, we will try to highlight the diversity of ways by which BMP signalling controls bilaterality in different animals, some of which do not belong to Bilateria. Future research combining functional analyses and modelling is bound to give us some understanding as to where the limits to the extent of the evolvability of BMP-dependent axial patterning may lie. [ABSTRACT FROM AUTHOR]

There is currently little information about the evolution of gene clusters, genome architectures and karyotypes in early branching animals. Slowly evolving anthozoan cnidarians can be particularly informative about the evolution of these genome features. Here we report chromosome-level genome assemblies of two related anthozoans, the sea anemones Nematostella vectensis and Scolanthus callimorphus. We find a robust set of 15 chromosomes with a clear one-to-one correspondence between the two species. Both genomes show chromosomal conservation, allowing us to reconstruct ancestral cnidarian and metazoan chromosomal blocks, consisting of at least 19 and 16 ancestral linkage groups, respectively. We show that, in contrast to Bilateria, the Hox and NK clusters of investigated cnidarians are largely disintegrated, despite the presence of staggered hox/gbx expression in Nematostella. This loss of microsynteny conservation may be facilitated by shorter distances between cis-regulatory sequences and their cognate transcriptional start sites. We find no clear evidence for topologically associated domains, suggesting fundamental differences in long-range gene regulation compared to vertebrates. These data suggest that large sets of ancestral metazoan genes have been retained in ancestral linkage groups of some extant lineages; yet, higher order gene regulation with associated 3D architecture may have evolved only after the cnidarian-bilaterian split.Slowly evolving cnidarians are useful models to study genome architecture. This study shows that sea anemones have a high degree of chromosomal macrosynteny, but poor microsynteny conservation. This is correlated with a small genome size and short distances of cis-regulatory elements to genes. [ABSTRACT FROM AUTHOR]

Canonical Wnt (cWnt) signalling is involved in a plethora of basic developmental processes such as endomesoderm specification, gastrulation and patterning the main body axis. To activate the signal, Wnt ligands form complexes with LRP5/6 and Frizzled receptors, which leads to nuclear translocation of ß-catenin and a transcriptional response. In Bilateria, the expression of different Frizzled genes is often partially overlapping, and their functions are known to be redundant in several developmental contexts. Here, we demonstrate that all four Frizzled receptors take part in the cWnt-mediated oralaboral axis patterning in the cnidarian Nematostella vectensis but show partially redundant functions. However, we do not see evidence for their involvement in the specification of the endoderm - an earlier event likely relying on maternal intracellular ß-catenin signalling components. Finally, we demonstrate that the main Wnt ligands crucial for the early oral-aboral patterning are Wnt1, Wnt3 and Wnt4. Comparison of our data with knowledge from other models suggests that distinct but overlapping expression domains and partial functional redundancy of cnidarian and bilaterian Frizzled genes may represent a shared ancestral trait. [ABSTRACT FROM AUTHOR]

Cnidarians are the only non-bilaterian group to evolve ciliated larvae with an apical sensory organ, which is possibly homologous to the apical organs of bilaterian primary larvae. Here, we generated transcriptomes of the apical tissue in the sea anemone Nematostella vectensis and showed that it has a unique neuronal signature. By integrating previously published larval single-cell data with our apical transcriptomes, we discovered that the apical domain comprises a minimum of six distinct cell types. We show that the apical organ is compartmentalised into apical tuft cells (spot) and larval-specific neurons (ring). Finally, we identify ISX-like (NVE14554), a PRD class homeobox gene specifically expressed in apical tuft cells, as an FGF signalling-dependent transcription factor responsible for the formation of the apical tuft domain via repression of the neural ring fate in apical cells. With this study, we contribute a comparison of the molecular anatomy of apical organs, which must be carried out across phyla to determine whether this crucial larval structure evolved once or multiple times. [ABSTRACT FROM AUTHOR]

In animals, body axis patterning is based on the concentration-dependent interpretation of graded morphogen signals, which enables correct positioning of the anatomical structures. The most ancient axis patterning system acting across animal phyla relies on β-catenin signaling, which directs gastrulation, and patterns the main body axis. However, within Bilateria, the patterning logic varies significantly between protostomes and deuterostomes. To deduce the ancestral principles of β-catenin-dependent axial patterning, we investigate the oral–aboral axis patterning in the sea anemone Nematostella—a member of the bilaterian sister group Cnidaria. Here we elucidate the regulatory logic by which more orally expressed β-catenin targets repress more aborally expressed β-catenin targets, and progressively restrict the initially global, maternally provided aboral identity. Similar regulatory logic of β-catenin-dependent patterning in Nematostella and deuterostomes suggests a common evolutionary origin of these processes and the equivalence of the cnidarian oral–aboral and the bilaterian posterior–anterior body axes. The authors show in Nematostella that the more orally expressed β-catenin targets repress the more aborally expressed β-catenin targets, thus patterning the oral-aboral axis. This likely represents the common mechanism of β-catenin-dependent axial patterning shared by Cnidaria and Bilateria. [ABSTRACT FROM AUTHOR]

Robust morphogenetic events are pivotal for animal embryogenesis. However, comparison of the modes of development of different members of a phylum suggests that the spectrum of developmental trajectories accessible for a species might be far broader than can be concluded from the observation of normal development. Here, by using a combination of microsurgery and transgenic reporter gene expression, we show that, facing a new developmental context, the aggregates of dissociated embryonic cells of the sea anemone Nematostella vectensis take an alternative developmental trajectory. The self-organizing aggregates rely on Wnt signals produced by the cells of the original blastopore lip organizer to form body axes but employ morphogenetic events typical for normal development of distantly related cnidarians to re-establish the germ layers. The reaggregated cells show enormous plasticity including the capacity of the ectodermal cells to convert into endoderm. Our results suggest that new developmental trajectories may evolve relatively easily when highly plastic embryonic cells face new constraints. [ABSTRACT FROM AUTHOR]

PIWI-interacting RNAs (piRNAs) and associated proteins comprise a conserved pathway for silencing transposons in metazoan germlines. piRNA pathway components are also expressed in multipotent somatic stem cells in various organisms. piRNA functions have been extensively explored in bilaterian model systems, however, comprehensive studies in non-bilaterian phyla remain limited. Here we investigate the piRNA pathway during the development ofNematostella vectensis, a well-established model system belonging to Cnidaria, the sister group to Bilateria. To date, no population of somatic stem cells has been identified in this organism, despite its long life-span and regenerative capacities that require a constant cell-renewal. We show thatNematostellapiRNA pathway components are broadly expressed in early developmental stages, while piRNAs themselves show differential expression, suggesting specific developmental roles of distinct piRNA families. In adults, piRNA associated proteins are enriched in the germline but also expressed in somatic cells, indicating putative stem cell properties. Furthermore, we provide experimental evidence thatNematostellapiRNAs cleave transposable elements as well as protein-coding genes. Our results demonstrate that somatic expression of piRNA associated proteins as well as the roles of piRNAs in transposon repression and gene regulation are likely ancestral features that evolved before the split between Cnidaria and Bilateria. [ABSTRACT FROM PUBLISHER]

Bilaterality - the possession of two orthogonal body axes - is the name-giving trait of all bilaterian animals. These body axes are established during early embryogenesis and serve as a threedimensional coordinate system that provides crucial spatial cues for developing cells, tissues, organs and appendages. The emergence of bilaterality was a major evolutionary transition, as it allowed animals to evolve more complex body plans. Therefore, how bilaterality evolved and whether it evolved once or several times independently is a fundamental issue in evolutionary developmental biology. Recent findings from non-bilaterian animals, in particular from Cnidaria, the sister group to Bilateria, have shed new light into the evolutionary origin of bilaterality. Here, we compare the molecular control of body axes in radially and bilaterally symmetric cnidarians and bilaterians, identify the minimal set of traits common for Bilateria, and evaluate whether bilaterality arose once or more than once during evolution. [ABSTRACT FROM AUTHOR]

ABSTRACT Matrotrophy, the continuous extra-vitelline supply of nutrients from the parent to the progeny during gestation, is one of the masterpieces of nature, contributing to offspring fitness and often correlated with evolutionary diversification. The most elaborate form of matrotrophy-placentotrophy-is well known for its broad occurrence among vertebrates, but the comparative distribution and structural diversity of matrotrophic expression among invertebrates is wanting. In the first comprehensive analysis of matrotrophy across the animal kingdom, we report that regardless of the degree of expression, it is established or inferred in at least 21 of 34 animal phyla, significantly exceeding previous accounts and changing the old paradigm that these phenomena are infrequent among invertebrates. In 10 phyla, matrotrophy is represented by only one or a few species, whereas in 11 it is either not uncommon or widespread and even pervasive. Among invertebrate phyla, Platyhelminthes, Arthropoda and Bryozoa dominate, with 162, 83 and 53 partly or wholly matrotrophic families, respectively. In comparison, Chordata has more than 220 families that include or consist entirely of matrotrophic species. We analysed the distribution of reproductive patterns among and within invertebrate phyla using recently published molecular phylogenies: matrotrophy has seemingly evolved at least 140 times in all major superclades: Parazoa and Eumetazoa, Radiata and Bilateria, Protostomia and Deuterostomia, Lophotrochozoa and Ecdysozoa. In Cycliophora and some Digenea, it may have evolved twice in the same life cycle. The provisioning of developing young is associated with almost all known types of incubation chambers, with matrotrophic viviparity more widespread (20 phyla) than brooding (10 phyla). In nine phyla, both matrotrophic incubation types are present. Matrotrophy is expressed in five nutritive modes, of which histotrophy and placentotrophy are most prevalent. Oophagy, embryophagy and histophagy are rarer, plausibly evolving through heterochronous development of the embryonic mouthparts and digestive system. During gestation, matrotrophic modes can shift, intergrade, and be performed simultaneously. Invertebrate matrotrophic adaptations are less complex structurally than in chordates, but they are more diverse, being formed either by a parent, embryo, or both. In a broad and still preliminary sense, there are indications of trends or grades of evolutionarily increasing complexity of nutritive structures: formation of ( i) local zones of enhanced nutritional transport (placental analogues), including specialized parent-offspring cell complexes and various appendages increasing the entire secreting and absorbing surfaces as well as the contact surface between embryo and parent, ( ii) compartmentalization of the common incubatory space into more compact and 'isolated' chambers with presumably more effective nutritional relationships, and ( iii) internal secretory ('milk') glands. Some placental analogues in onychophorans and arthropods mimic the simplest placental variants in vertebrates, comprising striking examples of convergent evolution acting at all levels-positional, structural and physiological. [ABSTRACT FROM AUTHOR]

Communication in bilaterian nervous systems is mediated by electrical and secreted signals; however, the evolutionary origin and relation of neurons to other secretory cell types has not been elucidated. Here, we use developmental single-cell RNA sequencing in the cnidarian Nematostella vectensis , representing an early evolutionary lineage with a simple nervous system. Validated by transgenics, we demonstrate that neurons, stinging cells, and gland cells arise from a common multipotent progenitor population. We identify the conserved transcription factor gene SoxC as a key upstream regulator of all neuroglandular lineages and demonstrate that SoxC knockdown eliminates both neuronal and secretory cell types. While in vertebrates and many other bilaterians neurogenesis is largely restricted to early developmental stages, we show that in the sea anemone, differentiation of neuroglandular cells is maintained throughout all life stages, and follows the same molecular trajectories from embryo to adulthood, ensuring lifelong homeostasis of neuroglandular cell lineages. [Display omitted] • Development and homeostasis of a sea anemone is captured at single-cell resolution • Developmental differentiation trajectories are maintained throughout the life cycle • Neurons, gland, and stinging cells derive from a common pool of putative stem cells • SoxC is an upstream regulator of neural progenitor cell specification Steger et al. present a temporal and spatially resolved single-cell RNA-seq atlas of a sea anemone that recapitulates development and homeostasis. Differentiation trajectories are shown to be maintained throughout the life cycle of Nematostella vectensis , in which SoxC specifies a neuroglandular progenitor population that differentiates into neurons, gland cells, and cnidocytes. [ABSTRACT FROM AUTHOR]

Summary BMP signaling plays a crucial role in the establishment of the dorso-ventral body axis in bilaterally symmetric animals. However, the topologies of the bone morphogenetic protein (BMP) signaling networks vary drastically in different animal groups, raising questions about the evolutionary constraints and evolvability of BMP signaling systems. Using loss-of-function analysis and mathematical modeling, we show that two signaling centers expressing different BMPs and BMP antagonists maintain the secondary axis of the sea anemone Nematostella . We demonstrate that BMP signaling is required for asymmetric Hox gene expression and mesentery formation. Computational analysis reveals that network parameters related to BMP4 and Chordin are constrained both in Nematostella and Xenopus , while those describing the BMP signaling modulators can vary significantly. Notably, only chordin , but not bmp4 expression needs to be spatially restricted for robust signaling gradient formation. Our data provide an explanation of the evolvability of BMP signaling systems in axis formation throughout Eumetazoa. [ABSTRACT FROM AUTHOR]

In triploblastic animals, mesoderm gives rise to many tissues and organs, including muscle. By contrast, the representatives of the diploblastic phylum Cnidaria (corals, sea anemones, jellyfish and hydroids) lack mesoderm but possess muscle. In vertebrates and insects, the transcription factor Mef2 plays a pivotal role in muscle differentiation; however, it is also an important regulator of neuron differentiation and survival. In the sea anemone Nematostella vectensis, an organism that lacks mesoderm but has muscles and neurons, Mef2 (Nvmef2) has been reported in single ectodermal cells of likely neural origin. To our surprise, we found that Nvmef2 is alternatively spliced, forming differentially expressed variants. Using morpholino-mediated knockdown and mRNA injection, we demonstrate that specific splice variants of Nvmef2 are required for the proliferation and differentiation of endodermal cells and for the development of ectodermal nematocytes, a neuronal cell type. Moreover, we identified a small conserved motif in the transactivation domain that is crucially involved in the endodermal function of Nvmef2. The identification of a crucial and conserved motif in the transactivation domain predicts a similarly important role in vertebrate Mef2 function. This is the first functional study of a determinant of several mesodermal derivatives in a diploblastic animal. Our data suggest that the involvement of alternative splice variants of Mef2 in endomesoderm and neuron differentiation predates the cnidarian-bilaterian split. [ABSTRACT FROM AUTHOR]

Novel organismal structures in metazoans are often undergirded by complex gene regulatory networks; as such, understanding the emergence of new structures through evolution requires reconstructing the series of evolutionary steps leading to these underlying networks. Here, we reconstruct the step-by-step assembly of the vertebrate splicing network regulated by Nova, a splicing factor that modulates alternative splicing in the vertebrate central nervous system by binding to clusters of YCAY motifs on pre-RNA transcripts. Transfection of human HEK293T cells with Nova orthologs indicated vertebrate-like splicing regulatory activity in bilaterian invertebrates, thus Nova acquired the ability to bind YCAY clusters and perform vertebrate-like splicing modulation at least before the last common ancestor of bilaterians. In situ hybridization studies in several species showed that Nova expression became restricted to CNS later on, during chordate evolution. Finally, comparative genomics studies revealed a diverse history for Nova-regulated exons, with target exons arising through both de novo exon creation and acquisition of YCAY motifs by preexisting exons throughout chordate and vertebrate history. In addition, we find that tissue-specific Nova expression patterns emerged independently in other lineages, suggesting independent assembly of tissue-specific regulatory networks. [ABSTRACT FROM AUTHOR]

The TGF-β molecules Dpp/BMP2/4/7 and their antagonist Sog/Chd play a conserved role in establishing the dorso-ventral (DV) axis in bilaterians. Homologues of BMP5 and the antagonist. Chordin, have been isolated from Cnidaria and show a striking asymmetric expression pattern with respect to the primary oral-aboral (OA) axis. We used Morpholino knockdowns of Nematostella dpp (bmp2/4), bmp5-8, chordin, and tolloid to investigate their function during early development of the sea anemone Nematostella vectensis. Molecular analysis of the BMP Morpholino phenotypes revealed an upregulated and radialized expression of bmps and chordin in ectoderm and endoderm indicating a negative feedback loop. Our data further suggest that BMP signaling is required for symmetry breaking of bmp and chordin expression during gastrulation. While bmps and chordin marker genes of the ectodermal OA axis extended aborally, other ectodermal markers of the OA axis were not significantly affected. By contrast, expression of other endodermal marker genes marking both the OA and the directive axis were abolished. Our data suggest that the logic of BMP2/4 signaling and the BMP antagonist, Chordin, differs significantly between Cnidaria and Bilateria, yet the double negative feedback loop detected in Nematostella bears systemic similarities with part of the regulatory network of the DV axis patterning system in amphibians. [ABSTRACT FROM AUTHOR]

The chicken talpid³ mutant, with polydactyly and defects in other embryonic regions that depend on hedgehog (Hh) signalling (e.g. the neural tube), has a mutation in KIAA0568. Similar phenotypes are seen in mice and in human syndromes with mutations in genes that encode centrosomal or intraflagella transport proteins. Such mutations lead to defects in primary cilia, sites where Hh signalling occurs. Here, we show that cells of talpid³ mutant embryos lack primary cilia and that primary cilia can be rescued with constructs encoding Talpid3. talpid³ mutant embryos also develop polycystic kidneys, consistent with widespread failure of ciliogenesis. Ultrastructural studies of talpid³ mutant neural tube show that basal bodies mature but fail to dock with the apical cell membrane, are misorientated and almost completely lack ciliary axonemes. We also detected marked changes in actin organisation in talpid³ mutant cells, which may explain misorientation of basal bodies. KIAA0586 was identified in the human centrosomal proteome and, using an antibody against chicken Talpid3, we detected Talpid3 in the centrosome of wild-type chicken cells but not in mutant cells. Cloning and bioinformatic analysis of the Talpid3 homolog from the sea anemone Nematostella vectensis identified a highly conserved region in the Talpid3 protein, including a predicted coiled-coil domain. We show that this region is required to rescue primary cilia formation and neural tube patterning in talpid³ mutant embryos, and is sufficient for centrosomal localisation. Thus, Talpid3 is one of a growing number of centrosomal proteins that affect both ciliogenesis and Hh signalling. [ABSTRACT FROM AUTHOR]