Origin of animal epithelia: insights from the sponge genome

Epithelial tissues are a key metazoan cell type, providing a basic structural unit for the construction of diverse animal body plans. Historically, an epithelial grade of organization was considered to be restricted to the Eumetazoa, with the majority of cell layers described for Porifera lacking any of the conserved ultrastructural characteristics of epithelia. Now with the use of genomic information from the demosponge, Amphimedon queenslandica, we identify orthologs of bilaterian genes that determine epithelial cell polarity or encode components of specialized epithelial junctions and extracellular matrix structures. Amphimedon possesses orthologs of most bilaterian epithelial polarity and adherens junction genes but few or no tight junction, septate junction, or basal lamina genes. To place this information in an evolutionary context, we extended these analyses to the completed genomes of various fungi, the choanoflagellate, Monosiga brevicollis, the placozoan, Trichoplax adhaerens, and the cnidarian, Nematostella vectensis. The results indicate that the majority of "epithelial" genes originated in metazoan or eumetazoan lineages, with only two genes, Par-1 and Discs large, antedating the choanoflagellate-metazoan split. We further explored the mechanism of evolution for each of these genes by tracking the origin of constituent domains and domain combinations. In general, domain configurations found in contemporary bilaterians are inferred to have evolved early in metazoan evolution and are identical or similar to those present in representatives of modern cnidarians, placozoans, and demosponges.     

Origin and evolution of laminin gene family diversity

Laminins are a family of multidomain glycoproteins that are important contributors to the structure of metazoan extracellular matrices. To investigate the origin and evolution of the laminin family, we characterized the full complement of laminin-related genes in the genome of the sponge, Amphimedon queenslandica. As a representative of the Demospongiae, a group consistently placed within the earliest diverging branch of animals by molecular phylogenies, Amphimedon is uniquely placed to provide insight into early steps in the evolution of metazoan gene families. Five Amphimedon laminin-related genes possess the conserved molecular features, and most of the domains found in bilaterian laminins, but all display domain architectures distinct from those of the canonical laminin chain types known from model bilaterians. This finding prompted us to perform a comparative genomic analysis of laminins and related genes from a choanoflagellate and diverse metazoans and to conduct phylogenetic analyses using the conserved Laminin N-terminal domain in order to explore the relationships between genes with distinct architectures. Laminin-like genes appear to have originated in the holozoan lineage (choanoflagellates + metazoans + several other unicellular opisthokont taxa), with several laminin domains originating later and appearing only in metazoan (animal) or eumetazoan (placozoans + ctenophores + cnidarians + bilaterians) laminins. Typical bilaterian α, β, and γ laminin chain forms arose in the eumetazoan stem and another chain type that is conserved in Amphimedon, the cnidarian, Nematostella vectensis, and the echinoderm, Strongylocentrotus purpuratus, appears to have been lost independently from the placozoan, Trichoplax adhaerens, and from multiple bilaterians. Phylogenetic analysis did not clearly reconstruct relationships between the distinct laminin chain types (with the exception of the α chains) but did reveal how several members of the netrin family were generated independently from within the laminin family by duplication and domain shuffling and by domain loss. Together, our results suggest that gene duplication and loss and domain shuffling and loss all played a role in the evolution of the laminin family and contributed to the generation of lineage-specific diversity in the laminin gene complements of extant metazoans.     

The NK homeobox gene cluster predates the origin of Hox genes

Hox and other Antennapedia (ANTP)-like homeobox gene subclasses - ParaHox, EHGbox, and NK-like - contribute to key developmental events in bilaterians [1-4]. Evidence of physical clustering of ANTP genes in multiple animal genomes [4-9] suggests that all four subclasses arose via sequential cis-duplication events. Here, we show that Hox genes' origin occurred after the divergence of sponge and eumetazoan lineages and occurred concomitantly with a major evolutionary transition in animal body-plan complexity. By using whole genome information from the demosponge Amphimedon queenslandica, we provide the first conclusive evidence that the earliest metazoans possessed multiple NK-like genes but no Hox, ParaHox, or EHGbox genes. Six of the eight NK-like genes present in the Amphimedon genome are clustered within 71 kb in an order akin to bilaterian NK clusters. We infer that the NK cluster in the last common ancestor to sponges, cnidarians, and bilaterians consisted of at least five genes. It appears that the ProtoHox gene originated from within this ancestral cluster after the divergence of sponge and eumetazoan lineages. The maintenance of the NK cluster in sponges and bilaterians for greater than 550 million years is likely to reflect regulatory constraints inherent to the organization of this ancient cluster.     

The genome of the sponge Amphimedon queenslandica provides new perspectives into the origin of Toll-like and interleukin 1 receptor pathways

Members of the Toll-like receptor (TLR) and the interleukin 1 receptor (IL1R) superfamilies activate various signaling cascades that are evolutionarily conserved in eumetazoans. In this study, we have searched the genome and expressed sequence tags of the demosponge Amphimedon queenslandica for molecules involved in TLR and IL1R signaling. Although we did not identify a conventional TLR or ILR, the Amphimedon genome encodes two related receptors, AmqIgTIRs, which are comprised of at least three extracellular IL1R-like immunoglobulins (Ig) and an intracellular TLR-like Toll/interleukin1 receptor/resistance (TIR) domain. The remainder of the TLR/IL1R pathway is mostly conserved in Amphimedon and includes genes known to interact with TLRs and IL1Rs in bilaterians, such as Toll-interacting protein (Tollip) and myeloid differentiation factor 88 (MyD88). By comparing the sponge genome to that of nonmetazoan eukaryotes and other basal animal phyla (i.e., placozoan and cnidarian representatives) we can infer that most components of the signaling cascade, including the receptors, evolved after the divergence of metazoan, and choanoflagellate lineages. In most cases, these proteins are composed of metazoan-specific domains (e.g., Pellino) or architectures (e.g., the association of a death domain with a TIR domain in the MyD88). The dynamic expression of the two AmqIgTIRs, AmqMyD88, AmqTollip, and AmqPellino during Amphimedon embryogenesis and larval development is consistent with the TLR/IL1R pathway having a role in both development and immunity in the last common metazoan ancestor.     

Animal phylogeny and the ancestry of bilaterians: inferences from morphology and 18S rDNA gene sequences

SUMMARY Insight into the origin and early evolution of the animal phyla requires an understanding of how animal groups are related to one another. Thus, we set out to explore animal phylogeny by analyzing with maximum parsimony 138 morphological characters from 40 metazoan groups, and 304 18S rDNA sequences, both separately and together. Both types of data agree that arthropods are not closely related to annelids: the former group with nematodes and other molting animals (Ecdysozoa), and the latter group with molluscs and other taxa with spiral cleavage. Furthermore, neither brachiopods nor chaetognaths group with deuterostomes; brachiopods are allied with the molluscs and annelids (Lophotrochozoa), whereas chaetognaths are allied with the ecdysozoans. The major discordance between the two types of data concerns the rooting of the bilaterians, and the bilaterian sister-taxon. Morphology suggests that the root is between deuterostomes and protostomes, with ctenophores the bilaterian sister-group, whereas 18S rDNA suggests that the root is within the Lophotrochozoa with acoel flatworms and gnathostomulids as basal bilaterians, and with cnidarians the bilaterian sister-group. We suggest that this basal position of acoels and gnathostomulids is artifactal because for 1000 replicate phylogenetic analyses with one random sequence as outgroup, the majority root with an acoel flatworm or gnathostomulid as the basal ingroup lineage. When these problematic taxa are eliminated from the matrix, the combined analysis suggests that the root lies between the deuterostomes and protostomes, and Ctenophora is the bilaterian sister-group. We suggest that because chaetognaths and lophophorates, taxa traditionally allied with deuterostomes, occupy basal positions within their respective protostomian clades, deuterostomy most likely represents a suite of characters plesiomorphic for bilaterians.     

Structure and expression of conserved Wnt pathway components in the demosponge Amphimedon queenslandica

Wnt-signalling plays a critical role in animal development, and its misregulation results in serious human diseases, including cancer. While the Wnt pathway is well studied in eumetazoan models, little is known about the evolutionary origin of its components and their functions. Here, we have identified key machinery of the Wnt-β-catenin (canonical)-signalling pathway that is encoded in the Amphimedon queenslandica (Demospongiae; Porifera) genome, namely Wnt, Fzd, SFRP, Lrp5/6, Dvl, Axin, APC, GSK3, β-catenin, Tcf, and Groucho. Most of these genes are not detected in the choanoflagellate and other nonmetazoan eukaryotic genomes. In contrast, orthologues of some of key components of bilaterian Wnt-planar cell polarity and Wnt/Ca(2+) are absent from the Amphimedon genome, suggesting these pathways evolved after demosponge and eumetazoan lineages diverged. Sequence analysis of the identified proteins of the Wnt-β-catenin pathway has revealed the presence of most of the conserved motifs and domains responsible for protein-protein and protein-DNA interactions in vertebrates and insects. However, several protein-protein interaction domains appear to be absent from the Amphimedon Axin and APC proteins. These are also missing from their orthologues in the cnidarian Nematostella vectensis, suggesting that they are bilaterian novelties. All of the analyzed Wnt pathway genes are expressed in specific patterns during Amphimedon embryogenesis. Most are expressed in especially striking and highly dynamic patterns during formation of a simple organ-like larval structure, the pigment ring. Overall, our results indicate that the Wnt-β-catenin pathway was used in embryonic patterning in the last common ancestor of living metazoans. Subsequently, gene duplications and a possible increase in complexity of protein interactions have resulted in the precisely regulated Wnt pathway observed in extant bilaterian animals.     

Cnidarian milestones in metazoan evolution

Cnidarians display most of the characters considered as milestonesof metazoan evolution. Whereas a tissue-level organization wasprobably already present in the multicellular common ancestorof all animals, the Urmetazoa, the emergence of important animalfeatures such as bilateral symmetry, triploblasty, a polarizednervous system, sense organs (eyes, statocysts), and a (chitinousor calcium-based) continuous skeleton can be traced back beforethe divergence between cnidarians and bilaterians. Modularityand metamery might be also regarded as two faces of the samemedal, likely involving conserved molecular mechanisms rulinganimal body architectures through regional specification ofiterated units. Available evidence indicates that the commonancestor of cnidarians and bilaterians, the UrEumetazoa, wasa surprisingly complex animal with nerve cell differentiation.We suggest that paedomorphic events in descendants of this ancestorled to the array of diversity seen in the main extant animalphyla. The use of molecular analyses and identifying the geneticdeterminants of anatomical organizations can provide an integrativetest of hypotheses of homologies and independent evidence ofthe evolutionary relationships among extant taxa.     

Hox and ParaHox genes in Nemertodermatida, a basal bilaterian clade

Molecular evidence suggests that Acoelomorpha, a proposed phylum composed of acoel and Nemertodermatida flatworms, are the most basal bilaterian animals. Hox and ParaHox gene complements characterised so far in acoels consist of a small set of genes, comprising representatives of anterior, central and posterior genes, altogether Hox and ParaHox, but no PG3-Xlox representatives have been reported. It has been proposed that this might be the ancestral Hox repertoire in basal bilaterians. However, no studies of the other members of the group, the Nemertodermatida, have been done. In order to get a more complete picture of the basal bilaterian Hox and ParaHox complement, we have analysed the Hox/ParaHox complement of the nemertodermatid Nemertoderma westbladi. We have found representatives of two central and one posterior Hox genes, as well as an Xlox and a Caudal ParaHox gene. From our data we conclude that a PG3-Xlox gene was present in the ancestor of bilaterians. These findings support the speculation that basal bilaterians already had the beginnings of the extended central Hox set, driving back gene duplications in the central part of the Hox cluster deeper in phylogeny than previously suggested.     

Comparative genomic and phylogenetic analysis of vitellogenin and other large lipid transfer proteins in metazoans

Vitellogenins and other large lipid transfer proteins (LLTP) are well known to play significant roles in the development, metabolism and reproduction of animals. Comparative genomics and phylogenetic analyses of LLTPs using the most comprehensive dataset in metazoans to date are carried out. Our analyses demonstrate that LLTP genes arose significantly earlier, and are more widespread than previously proposed - being present in numerous additional bilaterian and non-bilaterian lineages. A hypothesis is advanced that the most ancestral animal LLTP gene is Vtg, while loss of domains occurred at the bilaterians stem giving rise to apolipoprotein and microsomal triglyceride transfer proteins genes.     

Evolution of the TGF-β signaling pathway and its potential role in the ctenophore, Mnemiopsis leidyi

The TGF-β signaling pathway is a metazoan-specific intercellular signaling pathway known to be important in many developmental and cellular processes in a wide variety of animals. We investigated the complexity and possible functions of this pathway in a member of one of the earliest branching metazoan phyla, the ctenophore Mnemiopsis leidyi. A search of the recently sequenced Mnemiopsis genome revealed an inventory of genes encoding ligands and the rest of the components of the TGF-β superfamily signaling pathway. The Mnemiopsis genome contains nine TGF-β ligands, two TGF-β-like family members, two BMP-like family members, and five gene products that were unable to be classified with certainty. We also identified four TGF-β receptors: three Type I and a single Type II receptor. There are five genes encoding Smad proteins (Smad2, Smad4, Smad6, and two Smad1s). While we have identified many of the other components of this pathway, including Tolloid, SMURF, and Nomo, notably absent are SARA and all of the known antagonists belonging to the Chordin, Follistatin, Noggin, and CAN families. This pathway likely evolved early in metazoan evolution as nearly all components of this pathway have yet to be identified in any non-metazoan. The complement of TGF-β signaling pathway components of ctenophores is more similar to that of the sponge, Amphimedon, than to cnidarians, Trichoplax, or bilaterians. The mRNA expression patterns of key genes revealed by in situ hybridization suggests that TGF-β signaling is not involved in ctenophore early axis specification. Four ligands are expressed during gastrulation in ectodermal micromeres along all three body axes, suggesting a role in transducing earlier maternal signals. Later expression patterns and experiments with the TGF-β inhibitor SB432542 suggest roles in pharyngeal morphogenesis and comb row organization.     

The dawn of bilaterian animals: the case of acoelomorph flatworms

The origin of the bilaterian metazoans from radial ancestors is one of the biggest puzzles in animal evolution. A way to solve it is to identify the nature and main features of the last common ancestor of the bilaterians (LCB). Recent progress in molecular phylogeny has shown that many platyhelminth flatworms, regarded for a long time as basal bilaterians, now belong to the lophotrochozoan protostomates. In contrast, the LCB is now considered a complex organism bearing several features of modern bilaterians. Here we discuss an alternative view, in which acoelomorph (Acoela + Nemertodermatida) flatworms, which do not belong to the Platyhelminthes, represent the earliest extant bilaterian clade. Sequences from ribosomal and other nuclear genes, Hox cluster genes, and reinterpretation of some morphological features strongly support the basal position of acoelomorphs arguing against a complex LCB. This reconstruction backs the old planuloid-acoeloid hypothesis and may help our understanding of the evolution of body axes, Hox genes and the Cambrian explosion.     

Evolution of selenoproteins in the metazoan

ABSTRACT: BACKGROUND: The selenocysteine (Sec) containing proteins, selenoproteins, are an important group of proteins present throughout all 3 kingdoms of life. With the rapid progression of selenoprotein research in the post-genomic era, application of bioinformatics methods to the identification of selenoproteins in newly sequenced species has become increasingly important. Although selenoproteins in human and other vertebrates have been investigated, studies of primitive invertebrate selenoproteomes are rarely reported outside of insects and nematodes. Result A more integrated view of selenoprotein evolution was constructed using several representative species from different evolutionary eras. Using a SelGenAmic-based selenoprotein identification method, 178 selenoprotein genes were identified in 6 invertebrates: Amphimedon queenslandica, Trichoplax adhaerens, Nematostella vectensis, Lottia gigantean, Capitella teleta, and Branchiostoma floridae. Amphioxus was found to have the most abundant and variant selenoproteins of any animal currently characterized, including a special selenoprotein P (SelP) possessing 3 repeated Trx-like domains and Sec residues in the N-terminal and 2 Sec residues in the C-terminal. This gene structure suggests the existence of two different strategies for extension of Sec numbers in SelP for the preservation and transportation of selenium. In addition, novel eukaryotic AphC-like selenoproteins were identified in sponges. CONCLUSION: Comparison of various animal species suggests that even the most primitive animals possess a selenoproteome range and variety similar to humans. During evolutionary history, only a few new selenoproteins have emerged and few were lost. Furthermore, the massive loss of selenoproteins in nematodes and insects likely occurred independently in isolated partial evolutionary branches.     

RNA-binding proteins in human genetic disease

RNA-binding proteins (RBPs) are key components in RNA metabolism, regulating the temporal, spatial and functional dynamics of RNAs. Altering the expression of RBPs has profound implications for cellular physiology, affecting RNA processes from pre-mRNA splicing to protein translation. Recent genetic and proteomic data and evidence from animal models reveal that RBPs are involved in many human diseases ranging from neurologic disorders to cancer. Here we review the emerging evidence showing the involvement of RBPs in many disease networks and conclude that defects in RNA metabolism caused by aberrations in RBPs might underlie a broader spectrum of complex human disorders.     

Evidence for a microRNA expansion in the bilaterian ancestor

Understanding how animal complexity has arisen and identifying the key genetic components of this process is a central goal of evolutionary developmental biology. The discovery of microRNAs (miRNAs) as key regulators of development has identified a new set of candidates for this role. microRNAs are small noncoding RNAs that regulate tissue-specific or temporal gene expression through base pairing with target mRNAs. The full extent of the evolutionary distribution of miRNAs is being revealed as more genomes are scrutinized. To explore the evolutionary origins of metazoan miRNAs, we searched the genomes of diverse animals occupying key phylogenetic positions for homologs of experimentally verified human, fly, and worm miRNAs. We identify 30 miRNAs conserved across bilaterians, almost double the previous estimate. We hypothesize that this larger than previously realized core set of miRNAs was already present in the ancestor of all Bilateria and likely had key roles in allowing the evolution of diverse specialist cell types, tissues, and complex morphology. In agreement with this hypothesis, we found only three, conserved miRNA families in the genome of the sea anemone Nematostella vectensis and no convincing family members in the genome of the demosponge Reniera sp. The dramatic expansion of the miRNA repertoire in bilaterians relative to sponges and cnidarians suggests that increased miRNA-mediated gene regulation accompanied the emergence of triploblastic organ-containing body plans. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Mitochondrial genomes of two demosponges provide insights into an early stage of animal evolution

Mitochondrial DNA (mtDNA) of multicellular animals (Metazoa) is typically a small ( approximately 16 kbp), circular-mapping molecule that encodes 37 tightly packed genes. The structures of mtDNA-encoded transfer RNAs (tRNAs) and ribosomal RNAs (rRNAs) are usually highly unorthodox, and proteins are translated with multiple deviations from the standard genetic code. In contrast, mtDNA of the choanoflagellate Monosiga brevicollis, the closest unicellular relative of animals, is four times larger, contains 1.5 times as many genes, and lacks mentioned peculiarities of animal mtDNA. To investigate the evolutionary transition that led to the specific organization of metazoan mtDNA, we determined complete mitochondrial sequences from the demosponges Geodia neptuni and Tethya actinia, two representatives of the most basal animal phylum, the Porifera. We found that poriferan mtDNAs resemble those of other animals in their compact organization, lack of introns, and a well-conserved animal-like gene order. Yet, they contain several extra genes, encode bacterial-like rRNAs and tRNAs, and use a minimally derived genetic code. Our findings suggest that the evolution of the typical metazoan mtDNA has been a multistep process in which the compact genome organization and the reduced gene content were established prior to the reduction of tRNA and rRNA structures and the introduction of multiple changes of the translation code.     

The Origin of Patterning Systems in Bilateria——Insights from the Hox and ParaHox Genes in Acoelomorpha

Hox and ParaHox genes constitute two families of developmental regulators that pattern the Anterior-Posterior body axis in all bilaterians.The members of these two groups of genes are usually arranged in genomic clusters and work in a coordinated fashion,both in space and in time. While the mechanistic aspects of their action are relatively well known,it is still unclear how these systems evolved. For instance,we still need a proper model of how the Hox and ParaHox clusters were assembled over time.This problem is due to the shortage of information on gene complements for many taxa (mainly basal metazoans) and the lack of a consensus phylogenetic model of animal relationships to which we can relate our new findings.Recently, several studies have shown that the Acoelomorpha most probably represent the first offshoot of the Bilateria. This finding has prompted us,and others, to study the Hox and ParaHox complements in these animals,as well as their activity during development.In this review,we analyze how the current knowledge of Hox and ParaHox genes in the Acoelomorpha is shaping our view of bilaterian evolution.