Sally P. Leys

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.

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.

The Analysis of Eight Transcriptomes from All Poriferan Classes Reveals Surprising Genetic Complexity in Sponges

Sponges (Porifera) are among the earliest evolving metazoans. Their filter-feeding body plan based on choanocyte chambers organized into a complex aquiferous system is so unique among metazoans that it either reflects an early divergence from other animals prior to the evolution of features such as muscles and nerves, or that sponges lost these characters. Analyses of the Amphimedon and Oscarella genomes support this view of uniqueness-many key metazoan genes are absent in these sponges-but whether this is generally true of other sponges remains unknown. We studied the transcriptomes of eight sponge species in four classes (Hexactinellida, Demospongiae, Homoscleromorpha, and Calcarea) specifically seeking genes and pathways considered to be involved in animal complexity. For reference, we also sought these genes in transcriptomes and genomes of three unicellular opisthokonts, two sponges (A. queenslandica and O. carmela), and two bilaterian taxa. Our analyses showed that all sponge classes share an unexpectedly large complement of genes with other metazoans. Interestingly, hexactinellid, calcareous, and homoscleromorph sponges share more genes with bilaterians than with nonbilaterian metazoans. We were surprised to find representatives of most molecules involved in cell-cell communication, signaling, complex epithelia, immune recognition, and germ-lineage/sex, with only a few, but potentially key, absences. A noteworthy finding was that some important genes were absent from all demosponges (transcriptomes and the Amphimedon genome), which might reflect divergence from main-stem lineages including hexactinellids, calcareous sponges, and homoscleromorphs. Our results suggest that genetic complexity arose early in evolution as shown by the presence of these genes in most of the animal lineages, which suggests sponges either possess cryptic physiological and morphological complexity and/or have lost ancestral cell types or physiological processes.

Phylogenetic Position of the Hexactinellida Within the Phylum Porifera Based on the Amino Acid Sequence of the Protein Kinase C from Rhabdocalyptus dawsoni

Abstract. Recent analyses of genes encoding proteins typical for multicellularity, especially adhesion molecules and receptors, favor the conclusion that all metazoan phyla, including the phylum Porifera (sponges), are of monophyletic origin. However, none of these data includes cDNA encoding a protein from the sponge class Hexactinellida. We have now isolated and characterized the cDNA encoding a protein kinase C, belonging to the C subfamily (cPKC), from the hexactinellid sponge Rhabdocalyptus dawsoni. The two conserved regions, the regulatory part with the pseudosubstrate site, the two zinc fingers, and the C2 domain, as well as the catalytic domain were used for phylogenetic analyses. Sequence alignment and construction of a phylogenetic tree from the catalytic domains revealed that the yeast Saccharomyces cerevisiae and the protozoan Trypanosoma brucei are at the base of the tree, while the hexactinellid R. dawsoni branches off first among the metazoan sequences; the other two classes of the Porifera, the Calcarea (the sequence from Sycon raphanus was used) and the Demospongiae (sequences from Geodia cydonium and Suberites domuncula were used), branch off later. The statistically robust tree also shows that the two cPKC sequences from the higher invertebrates Drosophila melanogaster and Lytechinus pictus are most closely related to the calcareous sponge. This finding was also confirmed by comparing the regulatory part of the kinase gene. We suggest, that (i) within the phylum Porifera, the class Hexactinellida diverged first from a common ancestor to the Calcarea and the Demospongiae, which both appeared later, and (ii) the higher invertebrates are more closely related to the calcareous sponges.

The biology of glass sponges.

As the most ancient extant metazoans, glass sponges (Hexactinellida) have attracted recent attention in the areas of molecular evolution and the evolution of conduction systems but they are also interesting because of their unique histology: the greater part of their soft tissue consists of a single, multinucleate syncytium that ramifies throughout the sponge. This trabecular syncytium serves both for transport and as a pathway for propagation of action potentials that trigger flagellar arrests in the flagellated chambers. The present chapter is the first comprehensive modern account of this group and covers work going back to the earliest work dealing with taxonomy, gross morphology and histology as well as dealing with more recent studies. The structure of cellular and syncytial tissues and the formation of specialised intercellular junctions are described. Experimental work on reaggregation of dissociated tissues is also covered, a process during which histocompatibility, fusion and syncytialisation have been investigated, and where the role of the cytoskeleton in tissue architecture and transport processes has been studied in depth. The siliceous skeleton is given special attention, with an account of discrete spicules and fused silica networks, their diversity and distribution, their importance as taxonomic features and the process of silication. Studies on particle capture, transport of internalised food objects and disposal of indigestible wastes are reviewed, along with production and control of the feeding current. The electrophysiology of the conduction system coordinating flagellar arrests is described. The review covers salient features of hexactinellid ecology, including an account of habitats, distribution, abundance, growth, seasonal regression, predation, mortality, regeneration, recruitment and symbiotic associations with other organisms. Work on the recently discovered hexactinellid reefs of Canadas western continental shelf, analogues of long-extinct Jurassic sponge reefs, is given special attention. Reproductive biology is another area that has benefited from recent investigations. Seasonality, gametogenesis, embryogenesis, differentiation and larval biology are now understood in broad outline, at least for some species. The process whereby the cellular early larva becomes syncytial is described. A final section deals with the classification of recent and fossil glass sponges, phylogenetic relationships within the Hexactinellida and the phylogenetic position of the group within the Porifera. Palaeontological aspects are covered in so far as they are relevant to these topics.

Epithelia and integration in sponges.

An epithelium is important for integrity, homeostasis, communication and co-ordination, and its development must have been a fundamental step in the evolution of modern metazoan body plans. Sponges are metazoans that are often said to lack a true epithelium. We assess the properties of epithelia, and review the history of studies on sponge epithelia, focusing on their homology to bilaterian epithelia, their ultrastructure, and on their ability to seal. Electron micrographs show that adherens-type junctions are present in sponges but they can appear much slighter than equivalent junctions in other metazoans. Fine septae are seen in junctions of all sponge groups, but distinct septate junctions are only known from Calcarea. Similarly, all sponges can have collagenous sheets underlying their epithelia, but only homoscleromorphs are established to have a distinct basal lamina. The presence of most, but not all, gene families known to be involved in epithelial development and function also suggests that sponge epithelia function like, and are homologous to, bilaterian epithelia. However, physiological evidence that sponge epithelia regulate their internal environment is so far lacking. Given that up to six differentiated epithelia can be recognized in sponges, distinct physiological roles are expected. Recognition that sponges have epithelia challenges the perception that sponges are only loose associations of cells, and helps to relate the biology and physiology of the body plan of the adult sponge to the biology of other metazoans.

The Sponge Pump: The Role of Current Induced Flow in the Design of the Sponge Body Plan

Sponges are suspension feeders that use flagellated collar-cells (choanocytes) to actively filter a volume of water equivalent to many times their body volume each hour. Flow through sponges is thought to be enhanced by ambient current, which induces a pressure gradient across the sponge wall, but the underlying mechanism is still unknown. Studies of sponge filtration have estimated the energetic cost of pumping to be <1% of its total metabolism implying there is little adaptive value to reducing the cost of pumping by using “passive” flow induced by the ambient current. We quantified the pumping activity and respiration of the glass sponge Aphrocallistes vastus at a 150 m deep reef in situ and in a flow flume; we also modeled the glass sponge filtration system from measurements of the aquiferous system. Excurrent flow from the sponge osculum measured in situ and in the flume were positively correlated (r>0.75) with the ambient current velocity. During short bursts of high ambient current the sponges filtered two-thirds of the total volume of water they processed daily. Our model indicates that the head loss across the sponge collar filter is 10 times higher than previously estimated. The difference is due to the resistance created by a fine protein mesh that lines the collar, which demosponges also have, but was so far overlooked. Applying our model to the in situ measurements indicates that even modest pumping rates require an energetic expenditure of at least 28% of the total in situ respiration. We suggest that due to the high cost of pumping, current-induced flow is highly beneficial but may occur only in thin walled sponges living in high flow environments. Our results call for a new look at the mechanisms underlying current-induced flow and for reevaluation of the cost of biological pumping and its evolutionary role, especially in sponges.

Sponge Development and Antiquity of Animal Pattern Formation

Abstract The last common ancestor to all extant animals possessed features shared between the most basal metazoan lineage—Porifera—and the rest of the animal kingdom. To identify ancient and conserved developmental processes, we have been investigating embryogenesis and metamorphosis in the demosponge Reniera. Many of the cardinal features of eumetazoan development are displayed during Reniera embryogenesis. Specifically, after fertilization there is a period of cell division with little to no cell growth that results in two obvious cell populations distinguished by size as micromeres and macromeres, and by fate: the small cells differentiate into ciliated cells. This is followed by a period of differential cell activities that produces an embryo consisting of two then three layers, where at least 11 populations of differentiated cells are allocated into the different layers and patterned within these layers. This organization yields a swimming larva with the capacity to sense and respond to the surrounding environment, despite a lack of neurons and a coordinating system. During Reniera embryogenesis, the clearest example of cell patterning is the formation of a ring of pigment cells at the future posterior pole of the larva. Pigment cell pattern formation has two phases, both of which may require the movement of a large number of cells apparently in response to a morphogen gradient. First, pigmented cells, which initially cover the surface of the embryo, migrate to the future posterior end and form a dark spot. Second, the cells move outwards from the spot and rearrange into a ring. Numerous and diverse transcription factor genes are expressed during Reniera embryogenesis, most of which belong to metazoan-specific families and include members of POU, LIMHD, Pax, Bar, Prox2, NK-2, T-box, MEF-2, Fox, Sox, Ets, and nuclear hormone receptor families. In combination, these observations suggest that the last common ancestor to all extant metazoan lineages already possessed the basic regulatory genetic architecture to direct the specification, patterning and differentiation of multiple cell types. Some of these differentiated cells may have been arranged into localised functional units—i.e., simple tissues.

Evolutionary origins of sensation in metazoans: functional evidence for a new sensory organ in sponges.

BackgroundOne of the hallmarks of multicellular organisms is the ability of their cells to trigger responses to the environment in a coordinated manner. In recent years primary cilia have been shown to be present as ‘antennae’ on almost all animal cells, and are involved in cell-to-cell signaling in development and tissue homeostasis; how this sophisticated sensory system arose has been little-studied and its evolution is key to understanding how sensation arose in the Animal Kingdom. Sponges (Porifera), one of the earliest evolving phyla, lack conventional muscles and nerves and yet sense and respond to changes in their fluid environment. Here we demonstrate the presence of non-motile cilia in sponges and studied their role as flow sensors.ResultsDemosponges excrete wastes from their body with a stereotypic series of whole-body contractions using a structure called the osculum to regulate the water-flow through the body. In this study we show that short cilia line the inner epithelium of the sponge osculum. Ultrastructure of the cilia shows an absence of a central pair of microtubules and high speed imaging shows they are non-motile, suggesting they are not involved in generating flow. In other animals non-motile, ‘primary’, cilia are involved in sensation. Here we show that molecules known to block cationic ion channels in primary cilia and which inhibit sensory function in other organisms reduce or eliminate sponge contractions. Removal of the cilia using chloral hydrate, or removal of the whole osculum, also stops the contractions; in all instances the effect is reversible, suggesting that the cilia are involved in sensation. An analysis of sponge transcriptomes shows the presence of several transient receptor potential (TRP) channels including PKD channels known to be involved in sensing changes in flow in other animals. Together these data suggest that cilia in sponge oscula are involved in flow sensation and coordination of simple behaviour.ConclusionsThis is the first evidence of arrays of non-motile cilia in sponge oscula. Our findings provide support for the hypothesis that the cilia are sensory, and if true, the osculum may be considered a sensory organ that is used to coordinate whole animal responses in sponges. Arrays of primary cilia like these could represent the first step in the evolution of sensory and coordination systems in metazoans.

Wnt signaling and induction in the sponge aquiferous system: evidence for an ancient origin of the organizer

SUMMARY The importance of polarity—the possession of a primary body axis—is evident in the functional features of animals, such as feeding, and therefore must have arisen simultaneously with the evolution of multicellular animal body plans. Sponges are thought to represent the most ancient extant lineage of multicellular animals and whereas adult sponges do not possess obvious polarity, they are useful study organisms in which to examine the origin and evolution of body polarity. We tested the effect of pharmacological agents known to disrupt the polarity of a wide variety of animals on sponge organization during development. Lithium chloride and alsterpaullone, which mimic canonical Wnt signaling in other animals, caused formation of ectopic oscula and disrupted the ability of the sponge to feed. Transplanted oscula were able to attach to and induce canal reorganization in host sponges suggesting that the osulum has inductive capabilities. This work suggests that canonical Wnt signaling is responsible for setting up the aquiferous system, which acts as an organizing center polarizing the sponge.