The Starfish Asterina pectinifera: Collection and Maintenance of Adults and Rearing and Metamorphosis of Larvae.

Here we describe methods for (a) collecting starfish during their breeding period; (b) maintaining adults with fully grown gonads in laboratory aquaria; (c) rearing fertilized eggs to brachiolaria larvae, and (d) inducing larvae to metamorphose into juveniles under laboratory conditions. Such protocols should facilitate various analyses of starfish development throughout the entire life cycle of these model organisms.

Heterochronic activation of VEGF signaling and the evolution of the skeleton in echinoderm pluteus larvae.

The evolution of the echinoderm larval skeleton was examined from the aspect of interactions between skeletogenic mesenchyme cells and surrounding epithelium. We focused on vascular endothelial growth factor (VEGF) signaling, which was reported to be essential for skeletogenesis in sea urchin larvae. Here, we examined the expression patterns of vegf and vegfr in starfish and brittle stars. During starfish embryogenesis, no expression of either vegfr or vegf was detected, which contrast with previous reports on the expression of starfish homologs of sea urchin skeletogenic genes, including Ets, Tbr, and Dri. In later stages, when adult skeletogenesis commenced, vegfr and vegf expression were upregulated in skeletogenic cells and in the adjacent epidermis, respectively. These expression patterns suggest that heterochronic activation of VEGF signaling is one of the key molecular evolutionary steps in the evolution of the larval skeleton. The absence of vegf or vegfr expression during early embryogenesis in starfish suggests that the evolution of the larval skeleton requires distinct evolutionary changes, both in mesoderm cells (activation of vegfr expression) and in epidermal cells (activation of vegf expression). In brittle stars, which have well-organized skeletons like the sea urchin, vegfr and vegf were expressed in the skeletogenic mesenchyme and the overlying epidermis, respectively, in the same manner as in sea urchins. Therefore, the distinct activation of vegfr and vegf may have occurred in two lineages, sea urchins and brittle stars.

Characterization of a scavenger receptor cysteine-rich-domain-containing protein of the starfish, Asterina pectinifera: ApSRCR1 acts as an opsonin in the larval and adult innate immune systems.

Proteins containing a scavenger receptor cysteine-rich (SRCR) domain (SRCR proteins) play an important role in the innate immune system of various metazoan animals. In the starfish Asterina pectinifera, mesenchyme cells and coelomocytes govern the two distinct innate immune systems of the larvae and adults, respectively. Here we identify a cDNA encoding a protein containing nine SRCR domains termed ApSRCR1, and present characterization of the molecular structure, expression, subcellular localization and function of ApSRCR1 protein during ontogenesis of this animal. ApSRCR1 protein is a membrane-type protein with a predicted molecular mass of approximately 120 kDa. During ontogenesis, ApSRCR1 protein is de novo synthesized and localizes to cytoplasmic vesicles in both mesenchyme cells and coelomocytes without translation of maternal mRNA; however, the net production and modification by N-glycosylation of ApSRCR1 protein differs in each cell type. In both types of cell, functional inhibition of ApSRCR1 protein leads to incompetent bacterial clearance and failure of aggregate formation. However, this inhibitory effect is weaker in the mesenchyme cells than in the coelomocytes. In the bacteria-sensitized adult, ApSRCR1 protein is up-regulated and digested to enable its secretion into the coelomic fluid. This secreted form of ApSRCR1 protein can apparently bind to bacteria. Overall, we show that ApSRCR1 protein is finely regulated for expression not only during development but also in a sensitive innate immunological situation, and thereupon acts as an opsonin for bacteria to different extents in the larvae and adults of A. pectinifera.

Uneven distribution pattern and increasing numbers of mesenchyme cells during development in the starfish, Asterina pectinifera.

During development, the embryos and larvae of the starfish Asterina pectinifera possess a single type of mesenchyme cell. The aim of this study was to determine the patterns of behavior of mesenchyme cells during the formation of various organs. To this end, we used a monoclonal antibody (mesenchyme cell marker) to identify the distribution patterns and numbers of mesenchyme cells. Our results revealed the following: (i) mesenchyme cell behavior differs in the formation of different organs, showing temporal variations and an uneven pattern of distribution; and (ii) mesenchyme cells continue to be generated throughout development, and their numbers are tightly regulated in proportion to total cell numbers.

Modularity of gene-regulatory networks revealed in sea-star development.

Evidence that conserved developmental gene-regulatory networks can change as a unit during deutersostome evolution emerges from a study published in BMC Biology. This shows that genes consistently expressed in anterior brain patterning in hemichordates and chordates are expressed in a similar spatial pattern in another deuterostome, an asteroid echinoderm (sea star), but in a completely different developmental context (the animal-vegetal axis). This observation has implications for hypotheses on the type of development present in the deuterostome common ancestor.

Uncoupling of complex regulatory patterning during evolution of larval development in echinoderms.

BACKGROUND: Conservation of orthologous regulatory gene expression domains, especially along the neuroectodermal anterior-posterior axis, in animals as disparate as flies and vertebrates suggests that common patterning mechanisms have been conserved since the base of Bilateria. The homology of axial patterning is far less clear for the many marine animals that undergo a radical transformation in body plan during metamorphosis. The embryos of these animals are microscopic, feeding within the plankton until they metamorphose into their adult forms. RESULTS: We describe here the localization of 14 transcription factors within the ectoderm during early embryogenesis in Patiria miniata, a sea star with an indirectly developing planktonic bipinnaria larva. We find that the animal-vegetal axis of this very simple embryo is surprisingly well patterned. Furthermore, the patterning that we observe throughout the ectoderm generally corresponds to that of "head/anterior brain" patterning known for hemichordates and vertebrates, which share a common ancestor with the sea star. While we suggest here that aspects of head/anterior brain patterning are generally conserved, we show that another suite of genes involved in retinal determination is absent from the ectoderm of these echinoderms and instead operates within the mesoderm. CONCLUSIONS: Our findings therefore extend, for the first time, evidence of a conserved axial pattering to echinoderm embryos exhibiting maximal indirect development. The dissociation of head/anterior brain patterning from "retinal specification" in echinoderm blastulae might reflect modular changes to a developmental gene regulatory network within the ectoderm that facilitates the evolution of these microscopic larvae.

Fibrous component of the blastocoelic extracellular matrix shapes epithelia in concert with mesenchyme cells in starfish embryos.

By using a monoclonal antibody (4H11 Mab), we have investigated morphogenetic functions of a fibrous component of the blastocoelic extracellular matrix in relation to cellular activities during early development of the starfish Asterina pectinifera. The 4H11 fibers fill the blastocoele from the late-cleavage to late-gastrula stage and contain the 370-kDa proteinaceous molecule secreted only by the epithelial cells. When 4H11 Mab is introduced into the blastocoele of blastulae, the embryos reveal three distinct morphological abnormalities after the mid-gastrula stage: (1) Distribution of mesenchyme cells confined near the tip of the archenteron, (2) swelling of the posterior ectoderm, and (3) suppressed growth of the mouth, esophagus, and coelomic pouches. These abnormalities occur together with alterations in the distribution of the 4H11 fibers. In embryos recovering from the effect of 4H11 Mab, the mesenchyme cells rearrange the 4H11 fibers. We propose that 4H11 fibers play direct roles in the morphogenesis of starfish embryos by providing a dynamic scaffold not only for the mesenchyme cells but also for the epithelial cells. Moreover, 4H11 fibers have a resist force from within, in concert with the mesenchyme cells, to counter the bulging force intrinsic to the epithelia and hold the epithelia in specific positions, once the positions have been decided.