Bernard M. Degnan

The Amphimedon queenslandica genome and the evolution of animal complexity

Sponges are an ancient group of animals that diverged from other metazoans over 600 million years ago. Here we present the draft genome sequence of Amphimedon queenslandica, a demosponge from the Great Barrier Reef, and show that it is remarkably similar to other animal genomes in content, structure and organization. Comparative analysis enabled by the sequencing of the sponge genome reveals genomic events linked to the origin and early evolution of animals, including the appearance, expansion and diversification of pan-metazoan transcription factor, signalling pathway and structural genes. This diverse ‘toolkit’ of genes correlates with critical aspects of all metazoan body plans, and comprises cell cycle control and growth, development, somatic- and germ-cell specification, cell adhesion, innate immunity and allorecognition. Notably, many of the genes associated with the emergence of animals are also implicated in cancer, which arises from defects in basic processes associated with metazoan multicellularity.

Early origins and evolution of microRNAs and Piwi-interacting RNAs in animals

In bilaterian animals, such as humans, flies and worms, hundreds of microRNAs (miRNAs), some conserved throughout bilaterian evolution, collectively regulate a substantial fraction of the transcriptome. In addition to miRNAs, other bilaterian small RNAs, known as Piwi-interacting RNAs (piRNAs), protect the genome from transposons. Here we identify small RNAs from animal phyla that diverged before the emergence of the Bilateria. The cnidarian Nematostella vectensis (starlet sea anemone), a close relative to the Bilateria, possesses an extensive repertoire of miRNA genes, two classes of piRNAs and a complement of proteins specific to small-RNA biology comparable to that of humans. The poriferan Amphimedon queenslandica (sponge), one of the simplest animals and a distant relative of the Bilateria, also possesses miRNAs, both classes of piRNAs and a full complement of the small-RNA machinery. Animal miRNA evolution seems to have been relatively dynamic, with precursor sizes and mature miRNA sequences differing greatly between poriferans, cnidarians and bilaterians. Nonetheless, miRNAs and piRNAs have been available as classes of riboregulators to shape gene expression throughout the evolution and radiation of animal phyla.

A post-synaptic scaffold at the origin of the animal kingdom

Background The evolution of complex sub-cellular structures such as the synapse requires the assembly of multiple proteins, each conferring added functionality to the integrated structure. Tracking the early evolution of synapses has not been possible without genomic information from the earliest branching animals. As the closest extant relatives to the Eumetazoa, Porifera (sponges) represent a pivotal group for understanding the evolution of nervous systems, because sponges lack neurons with clearly recognizable synapses, in contrast to eumetazoan animals. Methodology/Principal Findings We show that the genome of the demosponge Amphimedon queenslandica possesses a nearly complete set of post-synaptic protein homologs whose conserved interaction motifs suggest assembly into a complex structure. In the critical synaptic scaffold gene, dlg, residues that make hydrogen bonds and van der Waals interactions with the PDZ ligand are 100% conserved between sponge and human, as is the motif organization of the scaffolds. Expression in Amphimedon of multiple post-synaptic gene homologs in larval flask cells further supports the existence of an assembled structure. Among the few post-synaptic genes absent from Amphimedon, but present in Eumetazoa, are receptor genes including the entire ionotropic glutamate receptor family. Conclusions/Significance Highly conserved protein interaction motifs and co-expression in sponges of multiple proteins whose homologs interact in eumetazoan synapses indicate that a complex protein scaffold was present at the origin of animals, perhaps predating nervous systems. A relatively small number of crucial innovations to this pre-existing structure may represent the founding changes that led to a post-synaptic element.

Origin and diversification of the basic helix-loop-helix gene family in metazoans: Insights from comparative genomics

BackgroundMolecular and genetic analyses conducted in model organisms such as Drosophila and vertebrates, have provided a wealth of information about how networks of transcription factors control the proper development of these species. Much less is known, however, about the evolutionary origin of these elaborated networks and their large-scale evolution. Here we report the first evolutionary analysis of a whole superfamily of transcription factors, the basic helix-loop-helix (bHLH) proteins, at the scale of the whole metazoan kingdom.ResultsWe identified in silico the putative full complement of bHLH genes in the sequenced genomes of 12 different species representative of the main metazoan lineages, including three non-bilaterian metazoans, the cnidarians Nematostella vectensis and Hydra magnipapillata and the demosponge Amphimedon queenslandica. We have performed extensive phylogenetic analyses of the 695 identified bHLHs, which has allowed us to allocate most of these bHLHs to defined evolutionary conserved groups of orthology.ConclusionThree main features in the history of the bHLH gene superfamily can be inferred from these analyses: (i) an initial diversification of the bHLHs has occurred in the pre-Cambrian, prior to metazoan cladogenesis; (ii) a second expansion of the bHLH superfamily occurred early in metazoan evolution before bilaterians and cnidarians diverged; and (iii) the bHLH complement during the evolution of the bilaterians has been remarkably stable. We suggest that these features may be extended to other developmental gene families and reflect a general trend in the evolution of the developmental gene repertoires of metazoans.

Genesis and Expansion of Metazoan Transcription Factor Gene Classes

We know little about the genomic events that led to the advent of a multicellular grade of organization in animals, one of the most dramatic transitions in evolution. Metazoan multicellularity is correlated with the evolution of embryogenesis, which presumably was underpinned by a gene regulatory network reliant on the differential activation of signaling pathways and transcription factors. Many transcription factor genes that play critical roles in bilaterian development largely appear to have evolved before the divergence of cnidarian and bilaterian lineages. In contrast, sponges seem to have a more limited suite of transcription factors, suggesting that the developmental regulatory gene repertoire changed markedly during early metazoan evolution. Using whole-genome information from the sponge Amphimedon queenslandica, a range of eumetazoans, and the choanoflagellate Monosiga brevicollis, we investigate the genesis and expansion of homeobox, Sox, T-box, and Fox transcription factor genes. Comparative analyses reveal that novel transcription factor domains (such as Paired, POU, and T-box) arose very early in metazoan evolution, prior to the separation of extant metazoan phyla but after the divergence of choanoflagellate and metazoan lineages. Phylogenetic analyses indicate that transcription factor classes then gradually expanded at the base of Metazoa before the bilaterian radiation, with each class following a different evolutionary trajectory. Based on the limited number of transcription factors in the Amphimedon genome, we infer that the genome of the metazoan last common ancestor included fewer gene members in each class than are present in extant eumetazoans. Transcription factor orthologues present in sponge, cnidarian, and bilaterian genomes may represent part of the core metazoan regulatory network underlying the origin of animal development and multicellularity.

Wnt and TGF-β Expression in the Sponge Amphimedon queenslandica and the Origin of Metazoan Embryonic Patterning

Background The origin of metazoan development and differentiation was contingent upon the evolution of cell adhesion, communication and cooperation mechanisms. While components of many of the major cell signalling pathways have been identified in a range of sponges (phylum Porifera), their roles in development have not been investigated and remain largely unknown. Here, we take the first steps toward reconstructing the developmental signalling systems used in the last common ancestor to living sponges and eumetazoans by studying the expression of genes encoding Wnt and TGF-β signalling ligands during the embryonic development of a sponge. Methodology/Principal Findings Using resources generated in the recent sponge Amphimedon queenslandica (Demospongiae) genome project, we have recovered genes encoding Wnt and TGF-β signalling ligands that are critical in patterning metazoan embryos. Both genes are expressed from the earliest stages of Amphimedon embryonic development in highly dynamic patterns. At the time when the Amphimedon embryos begin to display anterior-posterior polarity, Wnt expression becomes localised to the posterior pole and this expression continues until the swimming larva stage. In contrast, TGF-β expression is highest at the anterior pole. As in complex animals, sponge Wnt and TGF-β expression patterns intersect later in development during the patterning of a sub-community of cells that form a simple tissue-like structure, the pigment ring. Throughout development, Wnt and TGF-β are expressed radially along the anterior-posterior axis. Conclusions/Significance We infer from the expression of Wnt and TGF-β in Amphimedon that the ancestor that gave rise to sponges, cnidarians and bilaterians had already evolved the capacity to direct the formation of relatively sophisticated body plans, with axes and tissues. The radially symmetrical expression patterns of Wnt and TGF-β along the anterior-posterior axis of sponge embryos and larvae suggest that these signalling pathways contributed to establishing axial polarity in the very first metazoans.

Parallel Evolution of Nacre Building Gene Sets in Molluscs

The capacity to biomineralize is closely linked to the rapid expansion of animal life during the early Cambrian, with many skeletonized phyla first appearing in the fossil record at this time. The appearance of disparate molluscan forms during this period leaves open the possibility that shells evolved independently and in parallel in at least some groups. To test this proposition and gain insight into the evolution of structural genes that contribute to shell fabrication, we compared genes expressed in nacre (mother-of-pearl) forming cells in the mantle of the bivalve Pinctada maxima and the gastropod Haliotis asinina. Despite both species having highly lustrous nacre, we find extensive differences in these expressed gene sets. Following the removal of housekeeping genes, less than 10% of all gene clusters are shared between these molluscs, with some being conserved biomineralization genes that are also found in deuterostomes. These differences extend to secreted proteins that may localize to the organic shell matrix, with less than 15% of this secretome being shared. Despite these differences, H. asinina and P. maxima both secrete proteins with repetitive low-complexity domains (RLCDs). Pinctada maxima RLCD proteins-for example, the shematrins-are predominated by silk/fibroin-like domains, which are absent from the H. asinina data set. Comparisons of shematrin genes across three species of Pinctada indicate that this gene family has undergone extensive divergent evolution within pearl oysters. We also detect fundamental bivalve-gastropod differences in extracellular matrix proteins involved in mollusc-shell formation. Pinctada maxima expresses a chitin synthase at high levels and several chitin deacetylation genes, whereas only one protein involved in chitin interactions is present in the H. asinina data set, suggesting that the organic matrix on which calcification proceeds differs fundamentally between these species. Large-scale differences in genes expressed in nacre-forming cells of Pinctada and Haliotis are compatible with the hypothesis that gastropod and bivalve nacre is the result of convergent evolution. The expression of novel biomineralizing RLCD proteins in each of these two molluscs and, interestingly, sea urchins suggests that the evolution of such structural proteins has occurred independently multiple times in the Metazoa.

Developmental expression of transcription factor genes in a demosponge: insights into the origin of metazoan multicellularity.

SUMMARY Demosponges are considered part of the most basal evolutionary lineage in the animal kingdom. Although the sponge body plan fundamentally differs from that of other metazoans, their development includes many of the hallmarks of bilaterian and eumetazoan embryogenesis, namely fertilization followed by a period of cell division yielding distinct cell populations, which through a gastrulation‐like process become allocated into different cell layers and patterned within these layers. These observations suggest that the last common ancestor (LCA) to all living animals was developmentally more sophisticated than is widely appreciated and used asymmetric cell division and morphogen gradients to establish localized populations of specified cells within the embryo. Here we demonstrate that members of a range of transcription factor gene classes, many of which appear to be metazoan‐specific, are expressed during the development of the demosponge Reniera, including ANTP, Pax, POU, LIM‐HD, Sox, nuclear receptor, Fox (forkhead), T‐box, Mef2, and Ets genes. Phylogenetic analysis of these genes suggests that not only the origin but the diversification of some of the major developmental metazoan transcription factor classes took place before sponges diverged from the rest of the Metazoa. Their expression during demosponge development suggests that, as in todays sophisticated metazoans, these genes may have functioned in the regulatory network of the metazoan LCA to control cell specification and regionalized gene expression during embryogenesis.

Unexpected repertoire of metazoan transcription factors in the unicellular holozoan Capsaspora owczarzaki

How animals (metazoans) originated from their single-celled ancestors remains a major question in biology. As transcriptional regulation is crucial to animal development, deciphering the early evolution of associated transcription factors (TFs) is critical to understanding metazoan origins. In this study, we uncovered the repertoire of 17 metazoan TFs in the amoeboid holozoan Capsaspora owczarzaki, a representative of a unicellular lineage that is closely related to choanoflagellates and metazoans. Phylogenetic and comparative genomic analyses with the broadest possible taxonomic sampling allowed us to formulate new hypotheses regarding the origin and evolution of developmental metazoan TFs. We show that the complexity of the TF repertoire in C. owczarzaki is strikingly high, pushing back further the origin of some TFs formerly thought to be metazoan specific, such as T-box or Runx. Nonetheless, TF families whose beginnings antedate the origin of the animal kingdom, such as homeodomain or basic helix-loop-helix, underwent significant expansion and diversification along metazoan and eumetazoan stems.

Cytological basis of photoresponsive behavior in a sponge larva.

Ontogenetic changes in the photoresponse of larvae from the demosponge Reneira sp. were studied by analyzing the swimming paths of individual larvae exposed to diffuse white light. Larvae swam upward upon release from the adult, but were negatively phototactic until at least 12 hours after release. The larval photoreceptors are presumed to be a posterior ring of columnar monociliated epithelial cells that possess 120-μm-long cilia and pigment-filled protrusions. A sudden increase in light intensity caused these cilia to become rigidly straight. If the light intensity remained high, the cilia gradually bent over the pigmented vesicles in the adjacent cytoplasm, and thus covered one entire pole of the larva. The response was reversed upon a sudden decrease in light intensity. The ciliated cells were sensitive to changes in light intensity in larvae of all ages. This response is similar to the shadow response in tunicate larvae or the shading of the photoreceptor in Euglena and is postulated to allow the larvae to steer away from brighter light to darker areas, such as under coral rubble—the preferred site of the adult sponge on the reef flat. In the absence of a coordinating system in cellular sponges, the spatial organization and autonomous behavior of the pigmented posterior cells control the rapid responses to light shown by these larvae.