No abstract

Although sea urchins are one of the oldest and most widely used marine model systems, few species have been routinely kept in culture through multiple generations. The workhorse of the field is the purple urchin Strongylocentrotus purpuratus. However, one disadvantage of S. purpuratus is its long generation time, making it impractical as a model for generating and maintaining transgenic lines. In an effort to develop a sea urchin that is suitable for transgenerational experiments and the generation of transgenic lines, we have focused on development of updated culturing methods and genomic resources for the painted sea urchin, Lytechinus pictus. Compared to S. purpuratus, L. pictus have relatively large eggs, develop into optically clear embryos, and the smaller adults can become gravid in under a year. Fifty years ago, Hinegardner developed culturing methods for raising L. pictus through metamorphosis. Here, we provide an updated protocol for establishing and maintaining L. pictus in the laboratory, and describe a new genome resource for this urchin. In our hands, L. pictus reach the 4-armed pluteus stage at 4 days; become competent to metamorphosis at 24 days; and are gravid by 6 months. Plutei and juveniles are fed on a diet of algae and di-atoms, and adults are fed on kelp. We also make available a L. pictus transcriptome generated from developmental stages (eggs to 2-day-old plutei) to support the annotation of our genome sequencing project, and to enhance the utility of this species for molecular studies and transgenesis.

The gastropod shell is a composite composed of minerals and shell matrix proteins (SMPs). SMPs have been identified by proteomics in many molluscs, but few have been studied in detail. Open questions include 1) what gene regulatory networks regulate SMP expression, 2) what roles individual SMPs play in biomineralization, and 3) how the complement of SMPs changes over development. These questions are best addressed in a species in which gene perturbation studies are available; one such species is the slipper snail, Crepidula fornicata. Here, SEM and pXRD analysis demonstrated that the adult shell of C. fornicata exhibits crossed lamellar microstructure and is composed of aragonite. Using high-throughput proteomics we identified 185 SMPs occluded within the adult shell. Over half of the proteins in the shell proteome have known biomineralization domains, while at least 10% have no homologs in public databases. Differential gene expression analysis identified 20 SMP genes that are up-regulated in the shell-producing mantle tissue. Over half of these 20 SMPs are expressed during development with two, CfSMP1 and CfSMP2, expressed exclusively in the shell gland. Together, the description of the shell microstructure and a list of SMPs now sets the stage for studying the consequences of SMP gene knockdowns in molluscs.