Characterisation of the transcriptional dynamics underpinning the function, fate, and migration of the mouse Anterior Visceral Endoderm

Abstract

During early post-implantation development of the mouse embryo, the Anterior Visceral Endoderm (AVE) differs from surrounding visceral endoderm (VE) in its migratory behaviour and ability to restrict primitive streak formation to the opposite side of the egg cylinder. In order to characterise the molecular basis for the unique properties of the AVE, we combined single-cell RNA-sequencing of the VE prior to and during AVE migration, with high-resolution imaging, short-term lineage labelling, phosphoproteomics and pharmacological intervention. This revealed the transient nature of the AVE, the emergence of heterogeneities in AVE transcriptional states relative to position of cells, and its prominence in establishing gene expression asymmetries within the spatial constraints of the embryo. We identified a previously unknown requirement of Ephrin- and Semaphorin-signalling for AVE migration. These findings point to a tight coupling of transcriptional state and position in the AVE and reveal molecular heterogeneities underpinning its migratory behaviour and function.

Tissue-intrinsic Wnt signals antagonize Nodal-driven AVE differentiation

“…To corroborate this finding, we integrated the transcriptomes of BELA cells from clusters 3 and 4 with an embryo data set that focused especially on the AVE differentiation from E5.5 to 6.25. 16 Consistent with the integration with the wholeembryo dataset above, the majority of cells from both cluster 3 and 4 mapped together with E6.25 embryo cells (79% and 76%, respectively, Figures 3D-3F). While 95% of cells from cluster 4 were labeled as AVE after integration, cells from cluster 3 were labeled both as AVE (39%) and Epi-VE (59%, Figure 3F).…”

Embryonic and extraembryonic tissues during mammalian development: shifting boundaries in time and space

“…Until gastrulation, the parietal endoderm and VE facilitate nutrient and waste exchange between the implanted conceptus and the maternal tissues. In murid rodents, where the peri-implantation conceptus goes on to acquire an atypical cup shape, the VE differentiates over the proximal and distal regions, with distinct morphological and molecular characteristics [63]. The proximal region, which lines the polar TE-derived extraembryonic ectoderm, is a cuboidal epithelium that is continuous with the parietal endoderm, and likewise contributes to the yolk sac (figure 1 a ).…”

“…With no precise reference frame in developmental time, it is not always straightforward to determine which in vivo state a cell generated in vitro best represents. Differentially expressed genes no doubt could play roles in demarcating and maintaining embryonic and extraembryonic boundaries, as shown by the differential expression of Ephrin/Eph family members in tissues of the mouse peri-implantation conceptus [63]. Nonetheless, single markers, while indicating the ‘state’ of a cell, themselves are not adequate for determining the fate of a cell.…”

“…Such snapshots, from multiple cells collected at different stages of development, can be used to infer the in vivo ontogeny of tissues [105]. In the mouse for example, as the AVE migrates, it becomes transcriptionally more distant from the embryonic VE from which it originates, than the embryonic VE is from the extraembryonic VE [63]. It would therefore be interesting to see how the fate of AVE cells differs from that of other VE cells contributing to the gut endoderm as a consequence of this inferred transient transcriptomic divergence.…”

The role of cell geometry and cell-cell communication in gradient sensing

“…In conclusion, here we provide a modelling framework that clarifies the role of the size and the geometry of epithelial cells in gradient sensing, and can be easily extended to include further molecular and geometrical details. We believe it will be particularly interesting to combine this model with the large-scale sequencing and imaging data that are becoming available, for example on AVE migration in mouse embryos [66][67][68], with the goal of identifying the underlying molecular mechanisms. Many algorithms, in fact, have been developed to extract from single-cell and spatial transcriptomic datasets candidate molecules that could be mediating cellular communication (e.g., [69][70][71]).…”