Silicatein expression in Haliclona indistincta (Phylum Porifera, Order Haplosclerida) at different developmental stages.

Silicatein is the main protein responsible for the formation of spicules, tiny structures that constitute the silica skeleton of marine demosponges (Phylum Porifera). A unique innovation in Porifera that evolved from the cathepsin L family of proteins, it has been reported that two amino acids (S and H) are necessary to form the catalytic triad (SHN) to enable silica condensation. However, a diversity of silicatein sequence variants has since been reported with a variable pattern of presence/absence across sponge groups. Variants containing CHN or C/SQN at the active site appear more common in sponges from the Haplosclerida. Here, we report the expression levels of five silicatein variants through different developmental stages in the haplosclerid Haliclona indistincta. All five silicatein variants were expressed at low levels in the free-swimming larvae, which lack spicules and expression significantly increased at the two developmental phases in which spicules were visible. At these two phases, silicateins of CHN and C/SQN types were much more highly expressed than the SHN type indicating a possible ability of active sites with these alternative amino acids to condense silica and a more complex evolutionary story for spicule formation in marine demosponges than previously understood.

Evolution of the main skeleton-forming genes in sponges (phylum Porifera) with special focus on the marine Haplosclerida (class Demospongiae).

The skeletons of sponges (Phylum Porifera) are comprised of collagen, often embedded with small siliceous structures (spicules) arranged in various forms to provide strength and flexibility. The main proteins responsible for the formation of the spicules in demosponges are the silicateins, which are related to the cathepsins L of other animals. While the silicatein active site, necessary for the formation of biosilica crystals, is characterized by the amino acids SHN, different variants of the silicatein genes have been found, some that retain SHN at the active site and some that don't. As part of an effort to further understand skeleton formation in marine sponges of the order Haplosclerida, a search for all silicatein variants were made in Irish species representing the main clades of this large sponge group. For this task, transcriptomes were sequenced and de novo assembled from Haliclona oculata, H. simulans and H. indistincta. Silicatein genes were identified from these and all available genomes and transcriptomes from Porifera. These were analysed along with all complete silicateins from GenBank. Silicateins were only found in species belonging to the class Demospongiae but excluding Keratosa and Verongimorpha and there was significant duplication and diversity of these genes. Silicateins showing SHN at the active site were polyphyletic. Indeed silicatein sequences were divided into six major clades (CHNI, CHNII, CHNIII, SHNI, SHNII and C/SQN). In those clades where haplosclerids were well represented the silicatein phylogeny reflected previous ribosomal and mitochondrial topologies. The most basal silicatein clade (CHNI) contained sequences only from marine haplosclerids and freshwater sponges while one silicatein from H. indistincta was more related to cathepsins L (outgroup) than to the overall silicatein clade indicating the presence of an old silicatein or an intermediary form. This data could suggest that marine haplosclerids were one of the first groups of extant demosponges to acquire silicatein genes. Furthermore, we suggest that the paucity of spicule types in this group may be due to their single copy of SHNI variants, and the lack of a silintaphin gene.

Comparative Hox genes expression within the dimorphic annelid Streblospio benedicti reveals patterning variation during development.

Hox genes are transcriptional regulators that elicit cell positional identity along the anterior-posterior region of the body plan across different lineages of Metazoan. Comparison of Hox gene expression across distinct species reveals their evolutionary conservation; however, their gains and losses in different lineages can correlate with body plan modifications and morphological novelty. We compare the expression of 11 Hox genes found within Streblospio benedicti, a marine annelid that produces two types of offspring with distinct developmental and morphological features. For these two distinct larval types, we compare Hox gene expression through ontogeny using hybridization chain reaction (HCR) probes for in situ hybridization and RNA-seq data. We find that Hox gene expression patterning for both types is typically similar at equivalent developmental stages. However, some Hox genes have spatial or temporal differences between the larval types that are associated with morphological and life-history differences. This is the first comparison of developmental divergence in Hox gene expression within a single species and these changes reveal how body plan differences may arise in larval evolution.

Comparative Hox genes expression within the dimorphic annelid Streblospio benedicti reveals patterning variation during development.

Hox genes are transcriptional regulators that elicit cell positional identity along the anterior-posterior region of the body plan across different lineages of Metazoan. Comparison of Hox gene expression across distinct species reveals their evolutionary conservation, however their gains and losses in different lineages can correlate with body plan modifications and morphological novelty. We compare the expression of eleven Hox genes found within Streblospio benedicti, a marine annelid that produces two types of offspring with distinct developmental and morphological features. For these two distinct larval types, we compare Hox gene expression through ontogeny using HCR (hybridization chain reaction) probes for in-situ hybridization and RNA-seq data. We find that Hox gene expression patterning for both types is typically similar at equivalent developmental stages. However, some Hox genes have spatial or temporal differences between the larval types that are associated with morphological and life-history differences. This is the first comparison of developmental divergence in Hox genes expression within a single species and these changes reveal how body plan differences may arise in larval evolution.

Functional analysis in a model sea anemone reveals phylogenetic complexity and a role in cnidocyte discharge of DEG/ENaC ion channels.

Ion channels of the DEG/ENaC family share a similar structure but serve strikingly diverse biological functions, such as Na+ reabsorption, mechanosensing, proton-sensing, chemosensing and cell-cell communication via neuropeptides. This functional diversity raises the question of the ancient function of DEG/ENaCs. Using an extensive phylogenetic analysis across many different animal groups, we found a surprising diversity of DEG/ENaCs already in Cnidaria (corals, sea anemones, hydroids and jellyfish). Using a combination of gene expression analysis, electrophysiological and functional studies combined with pharmacological inhibition as well as genetic knockout in the model cnidarian Nematostella vectensis, we reveal an unanticipated role for a proton-sensitive DEG/ENaC in discharge of N. vectensis cnidocytes, the stinging cells typifying all cnidarians. Our study supports the view that DEG/ENaCs are versatile channels that have been co-opted for diverse functions since their early occurrence in animals and that respond to simple and ancient stimuli, such as omnipresent protons.