PRIMO lab interests
Multipotent cells
As embryogenesis proceeds from a totipotent zygote into a multicellular organism, diverse cell types are specified by a progressive loss of potency. By contrast, germ cells retain the potential for totipotency and give rise to the next generation (Figure 1).
The sea urchin exhibits maximal indirect development; embryogenesis culminates in the creation of a larva with the sole purpose of feeding and supporting the developing juvenile, which will emerge at metamorphosis (Figure 2). The juvenile sea urchin is patterned from cells that are set-aside during embryogenesis and shuttled into the larval coelomic pouches, the site of adult rudiment formation (Figure 3). Cells that contribute to the coelomic pouches come from various early embryonic lineages, including the small micromeres lineage. It is not known when the germ line is segregated during maximal indirect development. To address this question, we are testing the role of Vasa, Nanos, and Piwi, three conserved germ line markers, in sea urchin development. We find that Vasa, Nanos, and Piwi are selectively expressed in the small micromere lineage during embryogenesis. As larval development proceeds, the domain of Vasa expression expands to the entire developing adult rudiment. Thus, Vasa is not a strict germ line marker in the sea urchin, but may instead indicate multipotency. Functional analysis revealed that Nanos is required to maintain the identity of small micromeres as “set-aside” cells. Nanos-depleted small micromeres ectopically divide and are not incorporated into the larval coelomic pouches (Figure 4). In larvae formed from Nanos knockdown embryos, the adult rudiment cannot develop suggesting that the small micromeres contribute substantially to this structure (Figure 7). Thus, the small micromeres are likely multipotent cells that give rise to many tissues, which probably includes, but is not limited to the germ line. We hypothesize that the germline is segregated late in larval development, rather than during embryogenesis. Furthermore, we propose that the sea urchin uses a 2-step germline determination mechanism: first a long-term multipotent precursor is specified from which later the primordial germ cells (PGCs) are segregated. This strategy is used by Cnidarians and several Lophotrochozoans and therefore may be ancestral (Figure 4). Our work suggests a broader role for the traditional germline genes Vasa, Nanos, and Piwi in establishing and maintaining the fate of multipotent cells in 2-step germline determination.
Post-transcriptional regulation in small micromeres
Vasa, Nanos, and Piwi are proteins that accumulate in the sea urchin small micromeres. We have begun to test if post-transcriptional mechanisms are involved in this specific localization. These 3 proteins were also shown to be involved in translational control in different organisms: Nanos was shown to be a translational repressor, Vasa and Piwi were shown to be translational activators. We want to understand the translational functions of these 3 proteins in sea urchin small micromeres. We are now using a luciferase assay in sea urchin cell-free cap-dependent translation system to understand these mechanisms.