Supplementary Material for: The Chick Caudolateral Epiblast Acts as a Permissive Niche for Generating Neuromesodermal Progenitor Behaviours

Neuromesodermal progenitors (NMps) are a population of bipotent progenitors that maintain competence to generate both spinal cord and paraxial mesoderm throughout the elongation of the posterior body axis. Recent studies have generated populations of NMp-like cells in culture, which have been shown to differentiate to both neural and mesodermal cell fates when transplanted into either mouse or chick embryos. Here, we aim to compare the potential of mouse embryonic stem (ES) cell-derived progenitor populations to generate NMp behaviour against both undifferentiated and differentiated populations. We define NMp behaviour as the ability of cells to: (i) contribute to a significant proportion of the anterior-posterior body axis, (ii) enter into both posterior neural and somitic compartments, and (iii) retain a proportion of the progenitor population within the posterior growth zone. We compare previously identified ES cell-derived NMp-like populations to undifferentiated mouse ES cells and find that they all display similar potentials to generate NMp behaviour in vivo. To assess whether this competence is lost upon further differentiation, we generated anterior and posterior embryonic cell types through the generation of 3D gastruloids and show that NMp competence is lost within the anterior (Brachyury-negative) portion of the gastruloid. Together this suggests that in vitro-derived NMp-like cells maintain an ability to contribute to multiple germ layers that is also present within pluripotent ES cells, rather than acquiring a neuromesodermal competent state through differentiation.