A conserved germline multipotency program

Abstract

The germline of multicellular animals is segregated from somatic tissues, which is an essential developmental process for the next generation. Although certain ecdysozoans and chordates segregate their germline during embryogenesis, animals from other taxa segregate their germline after embryogenesis from multipotent progenitor cells. An overlapping set of genes, including vasa, nanos and piwi, operate in both multipotent precursors and in the germline. As we propose here, this conservation implies the existence of an underlying germline multipotency program in these cell types that has a previously underappreciated and conserved function in maintaining multipotency.

Fig. 1.

Germline segregation strategies. (A) In animals that segregate their germline during embryogenesis, primordial germ cells (PGCs) are specified in the embryo. PGCs give rise exclusively to either male or female germline stem cells (GSCs), which both self-renew (as indicated by the curved arrows) and give rise to a constant supply of gametes. Gametes are highly specialized cells, but when they fuse at fertilization they create a totipotent zygote. (B) In animals that segregate their germline after embryogenesis, a multipotent progenitor is established in the embryo from which the germline is segregated after embryogenesis is completed. We propose that the red cells in both panels operate a conserved germline multipotency program.

Fig. 2.

Metazoan phylogenetic tree. The evolutionary relationships among the major animal phyla. Animals mentioned in the text are noted in parentheses. Deuterostome phyla are shown in green, Ecdysozoa phyla in red and Lophotrochozoa phyla in blue. Drawings by S.Z.S. and Elizabeth Schroeder.

Fig. 3.

The juvenile sea urchin develops from embryonic multipotent cells. (A) Schematic of indirect development in sea urchin. During embryogenesis, cells are set aside for constructing the adult (the rudiment, shown in purple). The larva swims and feeds, providing protection and nutritional support to the developing adult structures. A S. purpuratus larva is competent to undergo metamorphosis after ∼6-8 weeks of feeding. (B) In the four-armed pluteus, the small micromere descendents are located in the left and right coelomic pouches (purple), where the adult rudiment will form. In the eight-armed pluteus, adult structures in the rudiment (purple) begin to form, such as the tube feet and spines. At metamorphosis, the juvenile emerges as an independent entity (purple) and larval tissues are lost.

Fig. 4.

Multipotent progenitor cells in the sea urchin embryo. (A) A schematic of the early stages of sea urchin development during which the small micromere lineage (purple) is set aside. The vegetal fourth cleavage division is unequal, thus giving rise to a 16-cell embryo with four micromeres (red). The micromeres divide unequally to give rise to four large micromeres (blue) and four small micromeres (purple). The small micromeres reside at the vegetal plate of the blastula where they divide once more. The eight small micromere descendents are carried at the tip of the invaginating gut during gastrulation and upon larva formation, are incorporated into the larval coelomic pouches, the site of adult rudiment formation. The small micromere lineage gives rise exclusively to tissues of the adult. (B) nanos, vasa and seawi (a sea urchin piwi ortholog) whole-mount in situ RNA hybridizations of S. purpuratus embryos and larva at the indicated stages of development. nanos, vasa and seawi, members of the germline multipotency program (GMP) gene set, are expressed in the sea urchin small micromere lineage. Images reproduced, with permission, from Juliano et al. (Juliano et al., 2006).

Fig. 5.

The multipotent small micromere program. (A) Images of S. purpuratus embryos at the four-cell, 16-cell and blastula stages of development, showing vasa whole-mount in situ RNA hybridizations (upper) (with vasa uniformly distributed throughout blastula formation) and Vasa protein (lower) enriched in the micromeres and small micromeres, respectively, on their formation [reproduced, with permission, from Juliano et al. (Juliano et al., 2006) and Voronina et al. (Voronina et al., 2008)]. (B) Control embryos produce larvae with well-formed adult rudiments around 4 weeks of age (broken white circle) that metamorphose around 6 weeks (middle panel). By contrast, when Nanos protein is knocked down in the embryo, the resultant larva fails to form an adult rudiment or undergo metamorphosis [reproduced, with permission, from Juliano et al. (Juliano et al., 2010)] (C). Vasa protein is post-transcriptionally enriched in the small micromere lineage (Voronina et al., 2008). nanos1 and nanos2 are expressed in this lineage at the 60-cell stage and are required for the continued enrichment of Vasa protein, for cell cycle repression and for the maintenance of the multipotent fate in this lineage (Juliano et al., 2010). Vasa protein enrichment precedes nanos transcription, but whether Vasa is required for nanos expression is unclear (question mark). Several similarities between the sea urchin small micromere germline multipotency program (GMP) and the Drosophila pole cell GMP are highlighted: Vasa protein (green) is selectively expressed in the pole cells as a result of regulated protein stability (Kugler et al., 2010; Styhler et al., 2002); nanos (blue) is required to maintain Vasa expression (Hayashi et al., 2004); and nanos (orange) is required to maintain cell fate by repression of the cell cycle and repression of apoptosis (Asaoka-Taguchi et al., 1999; Hayashi et al., 2004; Kadyrova et al., 2007).

Fig. 6.

Models of the origin of post-embryonic germline segregation. (A) Model 1: post-embryonic germline segregation is ancestral. The echinoderms and lophotrochozoans use the same post-embryonic mechanism (blue) and obtained their multipotency program from a common origin. In this scenario, the acquisition of embryonic germline segregation in some chordates (dark green) and ecdysozoans (light green) must have occurred independently. (B) Model 2: post-embryonic germline segregation is derived. In this model, the last common bilaterian ancestor used embryonic germline specification (green), which is still used by at least some chordates and ecdysozoans. Thus, the acquisition of post-embryonic germline segregation in echinoderms (red) and lophotrochozoans (orange) must have occurred independently. In this scenario, the last common bilaterian ancestor probably acquired embryonic germline segregation after splitting from the cnidarians. Germline segregation in hemichordates and platyzoans remains largely unexplored (black).