Mechanisms of specification, quiescence, and regeneration of primordial germ cells

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Project Summary We request support through this administrative supplement mechanism for a chilled marine aquarium unit to house our several echinoderm species, and our genetically manipulated adults derived from the experiments proposed in the parent application. Our 16-year-old main marine aquarium unit has failed, and our work has significantly expanded beyond its capacity and here we request support to replace it with a unit that will give us functionality and expanded capacity. The research supported by this request is focused on understanding the mechanisms of formation of primordial germ cells during embryogenesis, how they form during early development, and how they regenerate when the originals are removed. Our work leverages embryos from a sister group to chordates – the sea star and sea urchin. While not common organisms for biomedical research, these echinoderms have many strategic benefits for revealing unique perspectives in the biology of germline formation and regeneration. Millions of synchronous embryos from a single male/female cross allow biochemical and metabolic analysis of the germline, the resultant embryos have ideal transparency for in vivo longitudinal imaging, they develop rapidly, are easy to manipulate (single cell drop-mRNA-seq, optogenetics, cell and tissue transplantations) and they are well suited to complementary gene perturbation approaches (CRISPR/Cas9, morpholinoantisense oligonucleotides, MASO), and small molecule perturbations. The existing deep genomic and reagent resources for these animals, coupled with their tractable experimental characteristics, yields a unique system for understanding primordial germ cell biology with defined molecular and morphological endpoints, in live embryos with longitudinal analysis, distinct metrics of quantitation, and transgenerational evaluations.