Position-dependent plasticity of distinct progenitor types in the primitive streak
- University of Edinburgh, United Kingdom
Figures
Figure 1
Location of neuromesodermal and lateral/ventral mesoderm progenitors.
(A) Diagrams of the E8.5 (2–5 s) embryo showing the location of neuromesodermal and lateral/ventral-fated progenitors and the terminology used in this study. NSB, node-streak border; CLE, caudal …
Figure 2
The potency of NM-fated regions is restricted to neural and mesodermal lineages.
Grafts of primitive streak and tail bud regions to the kidney capsule. (A) Masson’s trichrome-stained tumour sections derived from the indicated embryonic tissue regions. (B) Percentage of tumours …
Figure 3 with 1 supplement
Sox2+T+cells coincide with NMP regions.
(A) In situ hybridisation for Sox2 and T. Abbreviations are the same as in Figure 1D. Noto, notochord; tbm, tail bud mesoderm. (B–F) Confocal sections of wholemount, immunostained embryos; …
Figure 3—figure supplement 1
Quantifying Sox2+T+cells during axis elongation.
(A) Double in situ hybridisation for Sox2 and T at the end of axis elongation (E13.5, 65s). (B) Workflow for Sox2/T quantification and 3D reconstruction. (C and F) Total cells analysed per embryonic …
Figure 4
Sox2+T+ NMPs peak at mid trunk formation.
(A) 3D reconstruction of the E8.5 caudal region. (Aa–b) Dorsal view. (Ac) Frontal view along the PS and CLE. Colours show different thresholded Sox2/T populations. (B–C) Quantification of different …
Figure 5
NMPs express low levels of T and Sox2.
(A–C) 3D analysis showing the relative levels of Sox2 and T protein in the E8.5–10.5 caudal region. Sox2+T+ cells express low-to-medium levels of both transcription factors (green cells in Ac, Ad, Bc…
Figure 6 with 4 supplements
Fate in the caudal lateral epiblast.
(A) Representative examples of homotopic CLE grafts (numbers identify individual embryos in B–D). Cell fate in the rostral CLE in rostral-to-caudal (Aa–l) and medial-to-lateral direction (Am–t). …
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Figure 6—source data 1
Presomitic mesoderm contamination in CLE grafts.
Additional information and analysis to show the majority of paraxial mesoderm in CLE grafts is derived from NMPs rather than contaminating PSM progenitors that were co-grafted.
- https://doi.org/10.7554/eLife.10042.011
Figure 6—figure supplement 1
Rostral CLE tissue contains Sox2+T+ cells.
Graft donor tissue was examined for the presence of Sox2/T coexpressing cells. L1-3 and L/St5 pieces were dissected as described above. The underlying presomitic mesoderm was manually removed in the …
Figure 6—figure supplement 2
Embryo numbers and section count in different grafting experiments.
(A) Schematic diagram summarising the different grafting experiments conducted in this work and the embryos series they correspond to. Diagrams represent the primitive streak region at E8.5, with …
Figure 6—figure supplement 3
Homotopic and heterotopic grafts incorporate well into host embryos.
(A–C) Immunohistochemical confirmation of differentiation markers in different graft series. Upper row, DAPI-counterstained images (grey). Boxes, region magnified in lower row. Lower row, …
Figure 6—figure supplement 4
Fate of the CLE progenitors in the paraxial mesoderm.
(A-B) Scoring of GFP+ cell contribution to the paraxial mesoderm in CLE homotopic grafts (embryo series 1 and 2). No PXM contribution was observed in L/St5 homotopic grafts. Graph format is the same …
Figure 7
Plasticity of L1-3 cells on heterotopic grafting to the NSB.
(A) Representative examples of heterotopic CLE grafts (numbers identify individual embryos in B). Three different contribution patterns were observed: mainly neural (Aa-d), neuromesodermal (Ae–h) …
Figure 8
Dorsolateral bias of L1-3 cells is reset upon grafting to the midline.
(A) Quantitative analysis of L1-3 graft contribution. Left, the percentage of neural (N) vs paraxial mesoderm (PXM) contribution in all scorable sections. Right, the percentage of embryos with …
Figure 9 with 3 supplements
Canonical Wnt signalling mediates neuromesodermal fate decisions.
(A) Representative examples of L1-3 to St1-3 grafts. (Aa and Af) Cells immediately after grafting. βcatCKO:sGFP grafted embryos were grown for 8 hr in the presence of 5µM 4-OHT after which the …
Figure 9—figure supplement 1
Obtaining conditional β-catenin knock out embryos.
(A) Schematic diagram of mouse cross to obtain βcatCKO:sGFP embryos. Mice, homozygous for both a silent GFP (Gilchrist et al., 2003) and floxed β-catenin alleles (Brault et al., 2001) were crossed …
Figure 9—figure supplement 2
Grafts using β-catenin knock out donor tissue.
(A–C) Grafting scheme of L1-3 (A) or L/St5 (B) to St1-3 heterotopic grafts. Either AGFP7 control or βcatCKO:sGFP donor tissue was used to transplant into WT host embryos. (C) After receiving the …
Figure 9—figure supplement 3
Levels of T in wildtype and 4-OHT-treated βcatCKO embryos.
(A) Boxplot for T intensity in notochord (noto), presomitic mesoderm (PSM) and T- (Neg) cells (n, number of nuclei sampled from wholemount stained embryos). The threshold for positivity was set at 7 …
Figure 10 with 2 supplements
The caudal tip of the CLE shows paraxial mesoderm but not neural potency.
(A) Representative examples of L/St5 heterotopic grafts (numbers in brackets identify individual embryos depicted in (B–C). (Aa, Af and Ak) Cells immediately after grafting. Arrowheads show grafted …
Figure 10—figure supplement 1
Overall lateral/ventral and tail bud mesoderm contribution.
(A) Percentage of total embryos and sections containing GFP+ cells in the lateral/ventral mesoderm. LVM contribution is very rare in L1 grafts. Note that the L2–3 grafts produced LVM at different …
Figure 10—figure supplement 2
Somite contribution in heterotopic LPMP grafts.
(A) Different L/St5 grafts performed. (B) Representative sections taken from grafts in A. Green, grafted cells; cyan, autofluorescence channel; arrows, notochord position; nt, neural tube; pxm, …
Figure 11
Position-dependent plasticity of primitive streak progenitors.
(A) NMPs and LPMPs coexist in the caudal progenitor zone. The NSB and L1 regions show NM fate. NM potency is broader, encompassing L2–3 regions and coincides with a Sox2+T+ phenotype. LPMPs reside …
Videos
Video 1
Sox2 and T expression domains during axis elongation.
3D reconstruction of the caudal progenitor region at different stages of axis elongation. (00:00) E8.5 (2-5s); (00:38) E9.5; (01:02) E10.5; (01:26) E11.5; (01:47) E12.5; (02:08) E13.5 (62-63s); …
Video 2
The number of T+ cells is affected upon β-catenin removal.
3D reconstruction of the caudal progenitor region. (00:00) E9.5 WT sample and (00:28) example of a βcatCKO embryo at E9.5, after it was cultured in vitro for 24 hr in the presence of 5µM 4-OHT.
