Origin of animal epithelia: insights from the genome of the demosponge Amphimedon queenslandica

Metazoan body plans are diverse but make use of a basic set of cell and tissue types conserved across the majority of animal phyla. Epithelial tissues represent one such tissue type. They provide a cohesive unit for the construction of body walls and organs, and allow for sealing between distinct bodily compartments and between the organism and the external environment. Traditionally, an epithelial grade of organization has been considered a synapomorphy for the Eumetazoa (Ctenophora, Cnidaria and Bilateria), with tissues found in the phyla, Porifera and Placozoa, observed to lack key structural characteristics of epithelia. A scenario in which the poriferan and placozoan lineages diverged from other metazoans prior to the acquisition of true epithelia in the eumetazoan stem is consistent with recent molecular phylogenomic analyses that place Porifera as the earliest-branching metazoan phyla with Placozoa basal amongst the Eumetazoa. The evolution of the epithelial tissue phenotype involved the innovation and integration of several cellular and extracellular features. As with the evolution of any complex trait, it is possible that some of these features preceded others, with the full epithelial phenotype appearing in the lineage leading to the Eumetazoa but making use of cellular and molecular characteristics acquired earlier in metazoan evolution. In particular, the observation that epithelia-like tissues of poriferans display some of the structural characteristics of eumetazoan epithelia suggests the possibility that the evolution of this tissue type proceeded step-wise. This study applies a comparative molecular approach to address this question. Studies in model bilaterians have shown that a conserved set of genes are necessary for the development and maintenance of key epithelial characteristics. These latter include aligned cellular apical-basal polarity, belt-form junctions of the stabilizing and occluding types and an underlying basal lamina. A survey of the genome of the demosponge, Amphimedon queenslandica, reveals that this sponge encodes a near-complete complement of proteins necessary for establishing apical-basal cell polarity and forming adherens junctions. By contrast, no direct orthologues of bilaterian tight junction, septate junction or basal lamina genes were found. To further explore the timing of origin of these genes, I extend genome surveys to the completed genomes of multiple species of fungi, the choanoflagellate, Monosiga brevicollis, the placozoan, Trichoplax adhaerens, and the cnidarian, Nematostella vectensis. These analyses reveal that the majority of polarity-determining and adherens junction genes were metazoan innovations, with only Par-1 and Discs large clearly antedating the choanoflagellate-metazoan split. By contrast, the majority of septate junction and basal lamina genes are present in placozoan and cnidarian genomes, suggesting that they originated in the eumetazoan or eumetazoan sensu stricto lineages. The phylogenetic distribution of domains and domain combinations for all surveyed genes shows that most were assembled from pre-existing domains, with the occasional incorporation of novel domains or motifs. Searches for basal lamina genes reveal that Amphimedon encodes a unique set of laminin-related proteins but none that could be considered directly orthologous to bilaterian laminins. The finding of laminins with novel domain architectures prompted a more thorough comparative genomic analysis, along with domain-specific phylogenies aimed at exploring the relationships between genes with distinct architectures. Sequence analysis of the Amphimedon laminin-related proteins suggests that they may be capable of forming heterotrimers similar to those that polymerize in the basal lamina of bilaterians. However, the differences in domain structure between Amphimedon and bilaterian laminins make it impossible to fully predict the properties of the sponge proteins. The comparative genomic analysis uncovers considerable diversity in the laminin gene complements of surveyed metazoans and indicates that laminin-like genes were a holozoan innovation with typical bilaterian chain types first appearing in the eumetazoan stem. The ciliated outer layer of the Amphimedon larva resembles an epithelial tissue, particularly in the display of aligned apical-basal polarity between constituent cells. In situ hybridisation of genes identified in the genome surveys described above reveals that the polarity-related genes, AmqPar-6, AmqPar-1, AmqMPP5/7 and AmqLgl1, and the adherens junction gene, Amqp120Catenin, are expressed in epithelial-like cells of the larvae from early in embryogenesis. AmqPar-3 and AmqaPKC are also expressed during development but it is unclear whether transcripts are present in outer layer cells. Fluorescent phallacidin staining of larvae indicates that actin microfilaments are arranged in a belt around the apical circumference of epithelial-like cells, possibly in conjunction with zonula adherens junctions, which have not previously been observed in Amphimedon by microscopy. In conclusion, the results presented here suggest that the molecular bases for key features of the epithelial tissue phenotype, namely apical-basal cell polarity and zonula adherens junctions, evolved in the metazoan lineage and are likely to be functional in poriferan tissues. The origin of genes coding for components of occluding junctions and basal lamina in eumetazoan and eumetazoan sensu stricto lineages allowed for the emergence of the full epithelial phenotype subsequent to the divergence of the Porifera