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Loss of <italic>gata2a</italic> expression in the endothelium of <italic>gata2a</italic><sup>Δi4/Δi4</sup> mutants leads to decreased levels of <italic>runx1</italic> and <italic>gata2b</italic> in the HE.a, b In situ hybridization for runx1 expression in the HE of wild-type and gata2aΔi4/Δi4 embryos at 28hpf. c Quantification of the runx1 in situ hybridization signal from wild-type (blue), heterozygous gata2a+/Δi4 (het, yellow) and gata2aΔi4/Δi4 (red) siblings at 28hpf shows significant decrease in runx1 pixel intensity in the DA in the homozygous mutants compared to wild-type (µwt = 34.8, µmut = 25.3; F = 4.956, d.f. = 2, 58; ANOVA),**p < 0.01. n = 14, wild-type; n = 25, het; n = 23, gata2aΔi4/Δi4. Error bars: mean ± SD. d qRT-PCR on RNA isolated from the sorted kdrl:GFP+ cells shows decreased levels of runx1 mRNA in the endothelium of gata2aΔi4/Δi4 mutants at 23hpf (t = 2.585, d.f. = 5, p < 0.05) but not at 30hpf (t = 1.326, d.f. = 4, p > 0.2), compared to wild-type. N = 4 for gata2aΔi4/Δi4 at 23hpf, N = 3 for other samples. Note different scales of expression levels. *p < 0.05. e, fGata2b expression in the HE of wild-type and gata2aΔi4/Δi4 embryos at 28hpf. g Quantification of the gata2b mRNA signal, detected by in situ hybridization, from wild-type (blue), heterozygous gata2a+/Δi4 (het; yellow) and gata2aΔi4/Δi4 (red) siblings at 28hpf shows significant decrease in gata2b pixel intensity in the DA in the homozygous mutants compared to wild-type (µwt = 39, µmut = 30.1; F = 5.05, d.f. = 2, 54; ANOVA), *p < 0.05. n = 22, wild-type; n = 24, het; n = 11, gata2aΔi4/Δi4. Error bars: mean ± SD. h qRT-PCR in sorted kdrl:GFP+ cells showed decreased levels of gata2b mRNA in the endothelium of gata2aΔi4/Δi4 mutants at 23hpf (t = 3.334, d.f. = 5, p < 0.05) but not at 30hpf (t = 0.373, d.f. = 4, p > 0.7), compared to wild-type. N = 4 for gata2aΔi4/Δi4 at 23hpf, N = 3 for other samples. *p < 0.05. See also Supplementary Figs. 3 and 4.
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