Research Articles Embryogenesis initiates with a totipotent fertilized egg cell, and subsequent development is characterized by progressive restrictions in cellular potential. At blastula stages, when the three primary germ layers are forming, chordate embryos still possess populations of pluripotent cells that are capable of differentiating into all somatic cell types. In mammals these are inner cell mass cells, whereas in Xenopus these cells are the deep/inner cells of the blastula roof, also termed animal pole cells (1). The pluripotency of blastula cells is transient; as embryogenesis proceeds into gastrulation, their potential becomes rapidly restricted into one of three cell types: ectoderm, mesoderm and endoderm. In all vertebrate species, a population of stem cell-like progenitors, called neural crest cells, represents an exception to this loss of potential. These cells arise from ectoderm positioned at the neural plate border, but in addition to ectodermal cell types can also differentiate into cartilage, bone, connective tissue, smooth muscle, pericytes and adipocytes, all of which are also formed by the mesoderm. Neural crest cells represent a major vertebrate innovation, collectively contributing to many of the features that distinguish vertebrates from non-vertebrate chordates, including much of the craniofacial skeleton, the chromaffin cells of the adrenal medulla and spinal nerve (dorsal root) ganglia. Because neural crest, despite its ectodermal origins, forms numerous cell types considered mesodermal, it has been likened to a fourth germ layer that renders vertebrates quadroblastic, and endows them with the potential to form a diversity of new cell types (2). Much effort has been di- rected toward determining the developmental mechanisms by which a classic embryonic induc- tion leads to the formation of the neural crest; cells that seemingly possess greater developmental potential than those from which they were derived embryological- ly or evolutionarily. Under cur- rent models these cells appear to defy the paradigm of progressive restriction in potential, and thus far no mechanism has been found to explain their apparent gain in potential. An alternative, more parsimonious, model for the origins of neural crest cells might be that they selectively retain the regulatory circuitry responsible for the pluripotency of their blastula precursors; a selective retention of earlier fea- tures. This model is supported by a shared requirement for Myc protein, and its transcrip- tional target Id3, in both neural crest cell genesis and ES cell pluripotency (3–6). We recently found that another neu- ral crest regulatory transcription factor, Sox5, is initially expressed in blastula cells where it functions as a BMP R- Smad co-factor (7), providing an additional link between neural crest cells and pluripotent blastula cells. Based on these observations, we decided to systematically test the alternative model in Xenopus. Neural crest shares regulatory circuitry with plu- ripotent blastula cells In mammals, Pou5F1 (Oct4), Sox2 and Nanog, constitute a core pluripotency network essential for maintaining the uncommitted state of blastula cells (8–13). In Xenopus, the Pou5F1 factors expressed in ectoderm are Pou5F3.1 (Oct91), Pou5F3.2 (Oct25) and Pou5F3.3 (Oct60) (14, 15). The func- tional role of Nanog in Xenopus is assumed by Ventx factors (Vent1/2) (16). These factors, along with Sox2, and the close- ly related Sox3, are expressed in blastula cells (17) (Fig. 1A). We wondered if other neural crest regulatory factors besides Myc, Id3 and Sox5 were co-expressed with the core pluripo- tency network in Xenopus blastula cells. We found that Id3, TF-AP2, Ets1, FoxD3 and Snail1 were co-expressed with the core pluripotency factors (Fig. 1B). FoxD3 and Snail1 are also expressed in murine ES cells (18, 19), providing further molecular links between neural crest factors and pluripo- Shared regulatory programs suggest retention of blastula-stage potential in neural crest cells Elsy Buitrago-Delgado, 1 * Kara Nordin, 1 * Anjali Rao, 1 Lauren Geary, 1 Carole LaBonne 1,2 † 1 Department of Molecular Biosciences, Northwestern University, Evanston, Il 60208, USA. 2 Robert H. Lurie Comprehensive Cancer Center, Northwestern University, Evanston, Il 60208, USA. *These authors contributed equally to this work. †Corresponding author: clabonne@northwestern.edu Neural crest cells, unique to vertebrates, arise in the ectoderm but can generate cell types typically categorized as mesodermal. This broad developmental potential persists past the time when most ectoderm- derived cells become lineage restricted. The ability of neural crest to contribute mesodermal derivatives to the bauplan has raised questions about how this apparent gain in potential is achieved. Here we describe shared molecular underpinnings of potency in neural crest and blastula cells. We show that in Xenopus, key neural crest regulatory factors are also expressed in blastula animal pole cells and promote pluripotency in both cell types. We suggest that neural crest cells may have evolved as a consequence of a subset of blastula cells retaining activity of the regulatory network underlying pluripotency. / sciencemag.org/content/early/recent / 30 April 2015 / Page 1 / 10.1126/science.aaa3655