Fig. 3

Fig. 3 miR-128 function in ECs to regulate HSPC heterogeneity before EHT.

a, Schematic representation of the transgene used to express WT miR-128 gene in zebrafish ECs (via flia1) or hemECs (via gata2b). For gata2b expression, we used the Tg(gata2b:Gal4^sd32) line and created the UAS:miR-128 plasmid, while for fli1a expression we created fli1a:miR-128 plasmid. One-cell-state embryo was injected with tol2 mRNA and the indicated plasmids, and WISH was performed against gata1a, ikzf1 and lcp1 at 4.5 dpf, respectively. b–d, WISH of gata1a (n = 27 (WT), 28 (128^Δ/Δ), 27 (128^Δ/Δ + fli1a), 29 (128^Δ/Δ + gata2b) embryos) (b) and ikaros (n = 41 (WT); 31 (128^Δ/Δ), 28 (128^Δ/Δ + fli1a), 28 (128^Δ/Δ + gata2b) embryos) (c) and lcp1 (n = 32 (WT), 27 (128^Δ/Δ), 30 (128^Δ/Δ + fli1a), 23 (128^Δ/Δ + gata2b) embryos) (d) at 4.5 dpf and relative cells quantification as indicated. fli1a endothelial expression of miR-128 WT gene rescues to WT level the increase of erythroid and lymphoid progenitors of miR-128^Δ/Δ, while gata2b hemEC expression of miR-128 does not rescue the increase of erythroid and lymphoid progenitors (3 independent experiments, ordinary one-way ANOVA with Tukey’s multiple comparison). All quantifications are represented with mean ± s.e.m. NS, not significant; P > 0.05, ****P ≤ 0.0001.