Spatiotemporal sequence of mesoderm and endoderm lineage segregation during mouse gastrulation

ABSTRACTAnterior mesoderm (AM) and definitive endoderm (DE) progenitors represent the earliest embryonic cell types that are specified during germ layer formation at the primitive streak (PS) of the mouse embryo. Genetic experiments indicate that both lineages segregate from Eomes-expressing progenitors in response to different Nodal signaling levels. However, the precise spatiotemporal pattern of the emergence of these cell types and molecular details of lineage segregation remain unexplored. We combined genetic fate labeling and imaging approaches with single-cell RNA sequencing (scRNA-seq) to follow the transcriptional identities and define lineage trajectories of Eomes-dependent cell types. Accordingly, all cells moving through the PS during the first day of gastrulation express Eomes. AM and DE specification occurs before cells leave the PS from Eomes-positive progenitors in a distinct spatiotemporal pattern. ScRNA-seq analysis further suggested the immediate and complete separation of AM and DE lineages from Eomes-expressing cells as last common bipotential progenitor.

Spatiotemporal sequence of mesoderm and endoderm lineage segregation during mouse gastrulation

Anterior mesoderm (AM) and definitive endoderm (DE) progenitors represent the earliest embryonic cell types that are specified during germ layer formation at the primitive streak (PS) of the mouse embryo. Genetic experiments indicate that both lineages segregate from Eomes expressing progenitors in response to different NODAL signaling levels. However, the precise spatiotemporal pattern of the emergence of these cell types and molecular details of lineage segregation remain unexplored. We combined genetic fate labeling and imaging approaches with scRNA-seq to follow the transcriptional identities and define lineage trajectories of Eomes dependent cell types. All cells moving through the PS during the first day of gastrulation express Eomes. AM and DE specification occurs before cells leave the PS from discrete progenitor populations that are generated in distinct spatiotemporal patterns. Importantly, we don’t find evidence for the existence of progenitors that co-express markers of both cell lineages suggesting an immediate and complete separation of AM and DE lineages.Summary statementCells lineages are specified in the mouse embryo already within the primitive streak where Mesp1+ mesoderm and Foxa2+ endoderm are generated in a spatial and temporal sequence from unbiased progenitors.

Molecular patterning during the development of Phoronopsis harmeri reveals similarities to rhynchonelliform brachiopods

BackgroundAnswering the question how conserved patterning systems are across evolutionary lineages requires a broad taxon sampling. Phoronid development has previously been studied using fate mapping and morphogenesis, yet molecular descriptions are missing. Here we report the expression patterns of the evolutionarily conserved anterior (otx, gsc, six3/6, nk2.1), posterior (cdx, bra) and endomesodermal (foxA, gata4/5/6, twist) markers in the phoronid Phoronopsis harmeri.ResultsThe transcription factors foxA, gata4/5/6 and cdx show conserved expression in patterning the development and regionalization of the phoronid embryonic gut, with foxA expressed in the presumptive foregut, gata4/5/6 demarcating the midgut and cdx confined to the hindgut. Surprisingly, brachyury, an evolutionary conserved transcription factor often associated with gastrulation movements and patterning of the mouth and hindgut, seems to be unrelated with gastrulation and mouth patterning in phoronids. Furthermore, six3/6, a well-conserved anterior marker, shows a remarkably dynamic expression, demarcating not only the apical organ and the oral ectoderm, but also clusters of cells of the developing midgut and the anterior mesoderm, similar to what has been reported for brachiopods, bryozoans and some deuterostome Bilateria.ConclusionsOur comparison of gene expression patterns with other studied Bilateria reveals that the timing of axis determination and cell fate distribution of the phoronid shows highest similarities to rhynchonelliform brachiopods. Despite these similarities, the phoronid P. harmeri shows also particularities in its development, which hint to divergences in the arrangement of gene regulatory networks responsible for germ layer formation and axis specification.

Single-cell mapping of gene expression landscapes and lineage in the zebrafish embryo

High-throughput mapping of cellular differentiation hierarchies from single-cell data promises to empower systematic interrogations of vertebrate development and disease. Here we applied single-cell RNA sequencing to >92,000 cells from zebrafish embryos during the first day of development. Using a graph-based approach, we mapped a cell-state landscape that describes axis patterning, germ layer formation, and organogenesis. We tested how clonally related cells traverse this landscape by developing a transposon-based barcoding approach (TracerSeq) for reconstructing single-cell lineage histories. Clonally related cells were often restricted by the state landscape, including a case in which two independent lineages converge on similar fates. Cell fates remained restricted to this landscape in embryos lacking the chordin gene. We provide web-based resources for further analysis of the single-cell data.

Gdf3 is required for robust Nodal signaling during germ layer formation and left-right patterning

Vertebrate embryonic patterning depends on signaling from Nodal, a TGFβ superfamily member. There are three Nodal orthologs in zebrafish; southpaw directs left-right asymmetries, while squint and cyclops function earlier to pattern mesendoderm. TGFβ member Vg1 is implicated in mesoderm formation but the role of the zebrafish ortholog, Growth differentiation factor 3 (Gdf3), has not been fully explored. We show that zygotic expression of gdf3 is dispensable for embryonic development, while maternally deposited gdf3 is required for mesendoderm formation and dorsal-ventral patterning. We further show that Gdf3 can affect left-right patterning at multiple stages, including proper development of regional cell morphology in Kupffer’s vesicle and the establishment of southpaw expression in the lateral plate mesoderm. Collectively, our data indicate that gdf3 is critical for robust Nodal signaling at multiple stages in zebrafish embryonic development.