nr2f2 does not regulate Notch in the zebrafish vasculature. (A) Schematic diagram of a 1 dpf zebrafish (modified from Kimmel et al., 1995) with a red box showing the approximate position of images in B–V. (B–F) Whole mount in situ hybridization of 24 hpf zebrafish probed for lyve1. (G–K) Red fluorescent confocal micrographs of 24 hpf Tg(Tp1bglob:hmgb1-mCherry)h11 (“Tp1:mCherry”) transgenic zebrafish. Inset numbers show quantitative measurements of trunk vascular mCherry fluorescence in a single representative injected fish normalized to trunk neural mCherry fluorescence and to values in untreated controls (G). See Materials and Methods for additional information on measurements. (L–P) Whole mount in situ hybridization of 24 hpf zebrafish probed for efnb2a. Animals in image panels B–P were either untreated (B, G and L), injected with nr2f2 MO (C, H and M), injected with kdrl-CF (Full length nr2f2 under the control of the kdrl promoter; D, I and N), injected with kdrl-ECt (chimeric protein with nr2f2 lacking the transactivation domain fused to engrailed repressor domain; E, J and O), or injected with kdrl-VCt (chimeric protein with nr2f2 lacking the transactivation domain fused to VP16; F,K,P). The inset numbers in panels C–F and M–P show the number of in situ-stained embryos exhibiting the phenotype shown in the image panel over the total number of embryos examined. (Q) Quantitative RT-PCR measurement of gene expression in excised trunks of 24 hpf animals (see Materials and Methods). Values are all normalized to control gene expression levels, which are set equal to 1. Rostral is to the left and dorsal is up in all image panels. Scale bar, 100µm.
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