Fig. 7

Figure 7. Transcriptomic heterogeneity and specialization in isl1-derived venous ECs constituting the vascular HSPC niche

(A) Single confocal plane of the tail of a TgBAC(isl1:TagRFP);Tg(dab2:EGFP) embryo (48 hpf). Isl1:TagRFP⁺/dab2:EGFP⁺ cells are mostly present in venous ECs of the CHT (arrows). Isl1:TagRFP expression is also detected in the vISVs (yellow arrowheads) and motor neurons (white arrowhead).

(B) Experimental design for scRNA-seq from the tail of TgBAC(isl1:TagRFP);Tg(dab2:EGFP) embryos.

(C) Uniform manifold approximation and projection (UMAP) visualization of tail venous ECs corresponding to the venous EC cluster in Figure S6. Reclustered cells are color-coded for the 5 identified sub-clusters.

(D) Heatmap illustrating the expression of the top-ranking marker genes enriched in each sub-cluster. Representative gene names are shown on top.

(E) Mapping of isl1:TagRFP⁻/dab2:EGFP⁺ cells (772 cells: green) and isl1:TagRFP⁺/dab2:EGFP⁺ cells (449 cells: pink) into the UMAP shown in (C).

(F) Percentage contribution of isl1:TagRFP⁻/dab2:EGFP⁺ cells and isl1:TagRFP⁺/dab2:EGFP⁺ cells to each venous EC sub-cluster.

(G) Heatmap illustrating the log-count expression, log₂(gene expression in isl1:TagRFP⁻ EGFP⁺/gene expression in isl1:TagRFP⁺ dab2:EGFP⁺), of the 36 differentially expressed genes between isl1:TagRFP⁻/dab2:EGFP⁺ cells and isl1:TagRFP⁺/dab2:EGFP⁺ cells (q value < 0.05 in at least one sub-cluster).

(H) Violin plots of the representative result of three genes; lyz, BX248121.1, and CT583728.24 in (G). The expression of lyz is dominant in isl1:TagRFP⁻/dab2:EGFP⁺ over isl1:TagRFP⁺ dab2:EGFP⁺ cells, whereas that of BX248121.1 and CT583728.24 is dominant in isl1:TagRFP⁺ dab2:EGFP⁺ cells over isl1:TagRFP⁻/dab2:EGFP⁺ cells.

(I) isl1:TagRFP⁻/dab2:EGFP⁺ cells and isl1:TagRFP⁺/dab2:EGFP⁺ cells were isolated from the tail of TgBAC(isl1:TagRFP);Tg(dab2:EGFP) embryos at 48 hpf by fluorescence-activated cell sorting (FACS). Relative expression of lyz mRNA, BX248121.1 mRNA, and CT583728.24 mRNA in FACS-sorted cells was analyzed by quantitative PCR. Data are normalized to the values in isl1:TagRFP⁻/dab2:EGFP⁺ cells (fold change = 2^(−ΔΔCt)). Data are mean ± SD. Each dot represents an individual experiment (n = 3 independent experiments).

Scale bars, 50 μm.

See also Figure S6 and Tables S1 and S2.