Non-bilaterian Animals Research Articles

Prolific Origination of Eyes in Cnidaria with Co-option of Non-visual Opsins

Animal eyes vary considerably in morphology and complexity and are thus ideal for understanding the evolution of complex biological traits [1]. While eyes evolved many times in bilaterian animals with elaborate nervous systems, image-forming and simpler eyes also exist in cnidarians, which are ancient non-bilaterians with neural nets and regions with condensed neurons to process information. How often eyes of varying complexity, including image-forming eyes, arose in animals with such simple neural circuitry remains obscure. Here, we produced large-scale phylogenies of Cnidaria and their photosensitive proteins and coupled them with anextensive literature search on eyes and light-sensing behavior to show that cnidarian eyes originated at least eight times, with complex, lensed-eyes having a history separate from other eye types. Compiled data show widespread light-sensing behavior in eyeless cnidarians, and comparative analyses support ancestors without eyes that already sensed light with dispersed photoreceptor cells. The history of expression of photoreceptive opsin proteins supports the inference of distinct eye origins via separate co-option of different non-visual opsin paralogs into eyes. Overall, our results show eyes evolved repeatedly from ancestral photoreceptor cells in non-bilaterian animals with simple nervous systems, co-opting existing precursors, similar to what occurred in Bilateria. Our study underscores the potential for multiple, evolutionarily distinct visual systems even in animals with simple nervous systems.

Early metazoan cell type diversity and the evolution of multicellular gene regulation.

A hallmark of metazoan evolution is the emergence of genomic mechanisms that implement cell type-specific functions. However, the evolution of metazoan cell types and their underlying gene regulatory programs remain largely uncharacterized. Here, we use whole-organism single-cell RNA-seq to map cell type-specific transcription in Porifera (sponges), Ctenophora (comb jellies) and Placozoa species. We describe the repertoires of cell types in these non-bilaterian animals, uncovering diverse instances of previously unknown molecular signatures, such as multiple types of peptidergic cells in Placozoa. Analysis of the regulatory programs of these cell types reveal variable levels of complexity. In placozoans and poriferans, sequence motifs in the promoters are predictive of cell type-specific programs. In contrast, the generation of a higher diversity of cell types in ctenophores is associated to lower specificity of promoter sequences and to the existence of distal regulatory elements. Our findings demonstrate that metazoan cell types can be defined by networks of TFs and proximal promoters, and indicate that further genome regulatory complexity may be required for more diverse cell type repertoires.

Prolific Origination of Eyes in Cnidaria With Co-Option of Non-Visual Opsins

Animal eyes vary considerably in morphology and complexity, and are thus essential for understanding the evolution of complex biological traits. While eyes evolved many times in bilaterian animals with elaborate nervous systems, image-forming and simpler eyes also exist in cnidarians, ancient non-bilaterians with only simple neural nets to process information. How often eyes of varying complexity, including image-forming eyes, arose in animals with such simple neural circuitry remains obscure. Here, we produced large-scale phylogenies of Cnidaria and their photosensitive proteins and coupled them with an extensive literature search on eyes and light-sensing behavior to show that cnidarian eyes originated at least eight times, with complex, lensed-eyes having a history separate from other eye types. Compiled data show widespread light-sensing behavior in eyeless cnidarians and comparative analyses support ancestors without eyes that already sensed light with dispersed photoreceptor cells. The history of expression of the photoreceptive protein opsin supports the inference of distinct eye origins via separate co-options of different non-visual opsin paralogs into eyes. Overall, our results show that eyes evolved repeatedly from ancestral photoreceptor cells in non-bilaterian animals with simple nervous systems, co-opting existing precursors, similar to what occurred in Bilateria. Our study underscores the potential for multiple, evolutionarily-distinct visual systems and underlying developmental pathways, even in animals with simple nervous systems.

On the evolution of bilaterality.

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 three-dimensional 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.

Evolution of miRNA Tailing by 3' Terminal Uridylyl Transferases in Metazoa.

In bilaterian animals the 3′ ends of microRNAs (miRNAs) are frequently modified by tailing and trimming. These modifications affect miRNA-mediated gene regulation by modulating miRNA stability. Here, we analyzed data from three nonbilaterian animals: two cnidarians (Nematostella vectensis and Hydra magnipapillata) and one poriferan (Amphimedon queenslandica). Our analysis revealed that nonbilaterian miRNAs frequently undergo modifications like the bilaterian counterparts: the majority are expressed as different length isoforms and frequent modifications of the 3′ end by mono U or mono A tailing are observed. Moreover, as the factors regulating miRNA modifications are largely uncharacterized in nonbilaterian animal phyla, in present study, we investigated the evolution of 3′ terminal uridylyl transferases (TUTases) that are known to involved in miRNA 3′ nontemplated modifications in Bilateria. Phylogenetic analysis on TUTases showed that TUTase1 and TUTase6 are a result of duplication in bilaterians and that TUTase7 and TUTase4 are the result of a vertebrate-specific duplication. We also find an unexpected number of Drosophila-specific gene duplications and domain losses in most of the investigated gene families. Overall, our findings shed new light on the evolutionary history of TUTases in Metazoa, as they reveal that this core set of enzymes already existed in the last common ancestor of all animals and was probably involved in modifying small RNAs in a similar fashion to its present activity in bilaterians.

A Large and Consistent Phylogenomic Dataset Supports Sponges as the Sister Group to All Other Animals

Resolving the early diversification of animal lineages has proven difficult, even using genome-scale datasets. Several phylogenomic studies have supported the classical scenario in which sponges (Porifera) are the sister group to all other animals ("Porifera-sister" hypothesis), consistent with a single origin of the gut, nerve cells, and muscle cells in the stem lineage of eumetazoans (bilaterians+ ctenophores+ cnidarians). In contrast, several other studies have recovered an alternative topology in which ctenophores are the sister group to all other animals (including sponges). The "Ctenophora-sister" hypothesis implies that eumetazoan-specific traits, such as neurons and muscle cells, either evolved once along the metazoan stem lineage and were then lost in sponges and placozoans or evolved at least twice independently in Ctenophora and in Cnidaria+ Bilateria. Here, we report on our reconstruction of deep metazoan relationships using a 1,719-gene dataset with dense taxonomic sampling of non-bilaterian animals that was assembled using a semi-automated procedure, designed to reduce known error sources. Our dataset outperforms previous metazoan gene superalignments in terms of data quality and quantity. Analyses with a best-fitting site-heterogeneous evolutionary model provide strong statistical supportfor placing sponges as the sister-group to all other metazoans, with ctenophores emerging as the second-earliest branching animal lineage. Only those methodological settings that exacerbated long-branch attraction artifacts yielded Ctenophora-sister. These results show that methodological issues must be carefully addressed to tackle difficult phylogenetic questions and pave the road to a better understanding of how fundamental features of animal body plans have emerged.

Animal Mitochondrial DNA as We Do Not Know It: mt-Genome Organization and Evolution in Nonbilaterian Lineages.

Animal mitochondrial DNA (mtDNA) is commonly described as a small, circular molecule that is conserved in size, gene content, and organization. Data collected in the last decade have challenged this view by revealing considerable diversity in animal mitochondrial genome organization. Much of this diversity has been found in nonbilaterian animals (phyla Cnidaria, Ctenophora, Placozoa, and Porifera), which, from a phylogenetic perspective, form the main branches of the animal tree along with Bilateria. Within these groups, mt-genomes are characterized by varying numbers of both linear and circular chromosomes, extra genes (e.g. atp9, polB, tatC), large variation in the number of encoded mitochondrial transfer RNAs (tRNAs) (0–25), at least seven different genetic codes, presence/absence of introns, tRNA and mRNA editing, fragmented ribosomal RNA genes, translational frameshifting, highly variable substitution rates, and a large range of genome sizes. This newly discovered diversity allows a better understanding of the evolutionary plasticity and conservation of animal mtDNA and provides insights into the molecular and evolutionary mechanisms shaping mitochondrial genomes.

Meeting report on "Animal Evolution: New Perspectives From Early Emerging Metazoans", Tutzing, September 14-17, 2015.

This recent meeting covered non-bilaterian (e.g., cnidarians, ctenophores, and sponges) animals broadly, but with emphasis in four areas: 1) New genomic resources and tools for functional studies, 2) advances in developmental and regeneration studies, 3) the evolution and function of nervous systems, 4) symbiosis and the holobiome.

Evolution of voltage-gated ion channels at the emergence of Metazoa.

Voltage-gated ion channels are large transmembrane proteins that enable the passage of ions through their pore across the cell membrane. These channels belong to one superfamily and carry pivotal roles such as the propagation of neuronal and muscular action potentials and the promotion of neurotransmitter secretion in synapses. In this review, we describe in detail the current state of knowledge regarding the evolution of these channels with a special emphasis on the metazoan lineage. We highlight the contribution of the genomic revolution to the understanding of ion channel evolution and for revealing that these channels appeared long before the appearance of the first animal. We also explain how the elucidation of channel selectivity properties and function in non-bilaterian animals such as cnidarians (sea anemones, corals, jellyfish and hydroids) can contribute to the study of channel evolution. Finally, we point to open questions and future directions in this field of research.

Evo-devo of non-bilaterian animals.

The non-bilaterian animals comprise organisms in the phyla Porifera, Cnidaria, Ctenophora and Placozoa. These early-diverging phyla are pivotal to understanding the evolution of bilaterian animals. After the exponential increase in research in evolutionary development (evo-devo) in the last two decades, these organisms are again in the spotlight of evolutionary biology. In this work, I briefly review some aspects of the developmental biology of nonbilaterians that contribute to understanding the evolution of development and of the metazoans. The evolution of the developmental genetic toolkit, embryonic polarization, the origin of gastrulation and mesodermal cells, and the origin of neural cells are discussed. The possibility that germline and stem cell lineages have the same origin is also examined. Although a considerable number of non-bilaterian species are already being investigated, the use of species belonging to different branches of non-bilaterian lineages and functional experimentation with gene manipulation in the majority of the non-bilaterian lineages will be necessary for further progress in this field.

TRF2 and the evolution of the bilateria.

The development of a complex body plan requires a diversity of regulatory networks. Here we consider the concept of TATA-box-binding protein (TBP) family proteins as "system factors" that each supports a distinct set of transcriptional programs. For instance, TBP activates TATA-box-dependent core promoters, whereas TBP-related factor 2 (TRF2) activates TATA-less core promoters that are dependent on a TCT or downstream core promoter element (DPE) motif. These findings led us to investigate the evolution of TRF2. TBP occurs in Archaea and eukaryotes, but TRF2 evolved prior to the emergence of the bilateria and subsequent to the evolutionary split between bilaterians and nonbilaterian animals. Unlike TBP, TRF2 does not bind to the TATA box and could thus function as a new system factor that is largely independent of TBP. We postulate that this TRF2-based system served as the foundation for new transcriptional programs, such as those involved in triploblasty and body plan development, that facilitated the evolution of bilateria.

The evolution of the four subunits of voltage-gated calcium channels: ancient roots, increasing complexity, and multiple losses.

The alpha subunits of voltage-gated calcium channels (Cavs) are large transmembrane proteins responsible for crucial physiological processes in excitable cells. They are assisted by three auxiliary subunits that can modulate their electrical behavior. Little is known about the evolution and roles of the various subunits of Cavs in nonbilaterian animals and in nonanimal lineages. For this reason, we mapped the phyletic distribution of the four channel subunits and reconstructed their phylogeny. Although alpha subunits have deep evolutionary roots as ancient as the split between plants and opistokonths, beta subunits appeared in the last common ancestor of animals and their close-relatives choanoflagellates, gamma subunits are a bilaterian novelty and alpha2/delta subunits appeared in the lineage of Placozoa, Cnidaria, and Bilateria. We note that gene losses were extremely common in the evolution of Cavs, with noticeable losses in multiple clades of subfamilies and also of whole Cav families. As in vertebrates, but not protostomes, Cav channel genes duplicated in Cnidaria. We characterized by in situ hybridization the tissue distribution of alpha subunits in the sea anemone Nematostella vectensis, a nonbilaterian animal possessing all three Cav subfamilies common to Bilateria. We find that some of the alpha subunit subtypes exhibit distinct spatiotemporal expression patterns. Further, all six sea anemone alpha subunit subtypes are conserved in stony corals, which separated from anemones 500 MA. This unexpected conservation together with the expression patterns strongly supports the notion that these subtypes carry unique functional roles.

Pan-metazoan phylogeny of the DMRT gene family: a framework for functional studies

The family of Doublesex-Mab-3 Related Transcription factors (DMRTs) includes key regulators of sexual differentiation and neurogenesis. To help understand the functional diversification of this gene family, we examined DMRT gene complements from the whole genome sequences and predicted gene models of 32 animal species representing 12 different phyla and from several non-metazoan outgroups. DMRTs are present in all animals except the sponge Amphimedon queenslandica, but are not found in any of the outgroups, indicating that this gene family is specific to animals and has an ancient pre-eumetazoan origin. Our analyses suggest that DMRT genes diversified independently in bilaterian and non-bilaterian animals. Most clades in the DMRT gene tree, including those containing the well-characterized DMRT1 and doublesex genes, have phylogenetically limited distributions.

Cnidarian microRNAs frequently regulate targets by cleavage

In bilaterians, which comprise most of extant animals, microRNAs (miRNAs) regulate the majority of messenger RNAs (mRNAs) via base-pairing of a short sequence (the miRNA “seed”) to the target, subsequently promoting translational inhibition and transcript instability. In plants, many miRNAs guide endonucleolytic cleavage of highly complementary targets. Because little is known about miRNA function in nonbilaterian animals, we investigated the repertoire and biological activity of miRNAs in the sea anemone Nematostella vectensis, a representative of Cnidaria, the sister phylum of Bilateria. Our work uncovers scores of novel miRNAs in Nematostella, increasing the total miRNA gene count to 87. Yet only a handful are conserved in corals and hydras, suggesting that microRNA gene turnover in Cnidaria greatly exceeds that of other metazoan groups. We further show that Nematostella miRNAs frequently direct the cleavage of their mRNA targets via nearly perfect complementarity. This mode of action resembles that of small interfering RNAs (siRNAs) and plant miRNAs. It appears to be common in Cnidaria, as several of the miRNA target sites are conserved among distantly related anemone species, and we also detected miRNA-directed cleavage in Hydra. Unlike in bilaterians, Nematostella miRNAs are commonly coexpressed with their target transcripts. In light of these findings, we propose that post-transcriptional regulation by miRNAs functions differently in Cnidaria and Bilateria. The similar, siRNA-like mode of action of miRNAs in Cnidaria and plants suggests that this may be an ancestral state.

Specific immune priming in the invasive ctenophore Mnemiopsis leidyi

Specific immune priming enables an induced immune response upon repeated pathogen encounter. As a functional analogue to vertebrate immune memory, such adaptive plasticity has been described, for instance, in insects and crustaceans. However, towards the base of the metazoan tree our knowledge about the existence of specific immune priming becomes scattered. Here, we exposed the invasive ctenophore Mnemiopsis leidyi repeatedly to two different bacterial epitopes (Gram-positive or -negative) and measured gene expression. Ctenophores experienced either the same bacterial epitope twice (homologous treatments) or different bacterial epitopes (heterologous treatments). Our results demonstrate that immune gene expression depends on earlier bacterial exposure. We detected significantly different expression upon heterologous compared with homologous bacterial treatment at three immune activator and effector genes. This is the first experimental evidence for specific immune priming in Ctenophora and generally in non-bilaterian animals, hereby adding to our growing notion of plasticity in innate immune systems across all animal phyla.

Mitogenomics at the base of Metazoa

Unraveling the base of metazoan evolution is of crucial importance for rooting the metazoan Tree of Life. This subject has attracted substantial attention for more than a century and recently fueled a burst of modern phylogenetic studies. Conflicting scenarios from different studies and incongruent results from nuclear versus mitochondrial markers challenge current molecular phylogenetic approaches. Here we analyze the presently most comprehensive data sets of mitochondrial genomes from non-bilaterian animals to illuminate the phylogenetic relationships among early branching metazoan phyla. The results of our analyses illustrate the value of mitogenomics and support previously known topologies between animal phyla but also identify several problematic taxa, which are sensitive to long branch artifacts or missing data.

Deep metazoan phylogeny: When different genes tell different stories

Molecular phylogenetic analyses have produced a plethora of controversial hypotheses regarding the patterns of diversification of non-bilaterian animals. To unravel the causes for the patterns of extreme inconsistencies at the base of the metazoan tree of life, we constructed a novel supermatrix containing 122 genes, enriched with non-bilaterian taxa. Comparative analyses of this supermatrix and its two non-overlapping multi-gene partitions (including ribosomal and non-ribosomal genes) revealed conflicting phylogenetic signals. We show that the levels of saturation and long branch attraction artifacts in the two partitions correlate with gene sampling. The ribosomal gene partition exhibits significantly lower saturation levels than the non-ribosomal one. Additional systematic errors derive from significant variations in amino acid substitution patterns among the metazoan lineages that violate the stationarity assumption of evolutionary models frequently used to reconstruct phylogenies. By modifying gene sampling and the taxonomic composition of the outgroup, we were able to construct three different yet well-supported phylogenies. These results show that the accuracy of phylogenetic inference may be substantially improved by selecting genes that evolve slowly across the Metazoa and applying more realistic substitution models. Additional sequence-independent genomic markers are also necessary to assess the validity of the phylogenetic hypotheses.

Muscle's dual origins

Jellyfish move using a set of muscles that look remarkably similar to striated muscles in vertebrates. However, new data show that the two muscle types contain different molecules, implying that they evolved independently. See Letter p.231 The ultrastructural similarity between striated skeletal muscles in higher bilaterian animals, such as humans and insects, and non-bilaterian animals, such as jellyfish, suggests a common evolutionary origin of this important contractile cell type. However, the precise evolution of striated muscles has not been confirmed. In this phylogenomic study, Ulrich Technau and colleagues show that core muscle proteins are present in unicellular organisms, such as sponges, that do not have true muscles. Their analysis supports a convergent-evolution model for striated muscles. According to this model, novel proteins were added to the ancient contractile apparatus during the independent evolution of the cnidarians and bilaterians, resulting in strikingly similar ultrastructures.

A "Neural" Enzyme in Nonbilaterian Animals and Algae: Preneural Origins for Peptidylglycine -Amidating Monooxygenase

Secreted peptides, produced by enzymatic processing of larger precursor molecules, are found throughout the animal kingdom and play important regulatory roles as neurotransmitters and hormones. Many require a carboxy-terminal modification, involving the conversion of a glycine residue into an α-amide, for their biological activity. Two sequential enzymatic activities catalyze this conversion: a monooxygenase (peptidylglycine α-hydroxylating monooxygenase or PHM) and an amidating lyase (peptidyl-α-hydroxyglycine α-amidating lyase or PAL). In vertebrates, these activities reside in a single polypeptide known as peptidylglycine α-amidating monooxygenase (PAM), which has been extensively studied in the context of neuropeptide modification. Bifunctional PAMs have been reported from some invertebrates, but the phylogenetic distribution of PAMs and their evolutionary relationship to PALs and PHMs is unclear. Here, we report sequence and expression data for two PAMs from the coral Acropora millepora (Anthozoa, Cnidaria), as well as providing a comprehensive survey of the available sequence data from other organisms. These analyses indicate that bifunctional PAMs predate the origins of the nervous and endocrine systems, consistent with the idea that within the Metazoa their ancestral function may have been to amidate epitheliopeptides. More surprisingly, the phylogenomic survey also revealed the presence of PAMs in green algae (but not in higher plants or fungi), implying that the bifunctional enzyme either predates the plant/animal divergence and has subsequently been lost in a number of lineages or perhaps that convergent evolution or lateral gene transfer has occurred. This finding is consistent with recent discoveries that other molecules once thought of as "neural" predate nervous systems.

Premetazoan Origin of the Hippo Signaling Pathway

Nonaggregative multicellularity requires strict control of cell number. The Hippo signaling pathway coordinates cell proliferation and apoptosis and is a central regulator of organ size in animals. Recent studies have shown the presence of key members of the Hippo pathway in nonbilaterian animals, but failed to identify this pathway outside Metazoa. Through comparative analyses of recently sequenced holozoan genomes, we show that Hippo pathway components, such as the kinases Hippo and Warts, the coactivator Yorkie, and the transcription factor Scalloped, were already present in the unicellular ancestors of animals. Remarkably, functional analysis of Hippo components of the amoeboid holozoan Capsaspora owczarzaki, performed in Drosophila melanogaster, demonstrate that the growth-regulatory activity of the Hippo pathway is conserved in this unicellular lineage. Our findings show that the Hippo pathway evolved well before the origin of Metazoa and highlight the importance of Hippo signaling as a key developmental mechanism predating the origin of Metazoa.