Fig. 3: Interplay of EphB4, ephrin-B2 and SoxF factors.
From: Eph-ephrin signaling couples endothelial cell sorting and arterial specification
a–d Confocal images and quantitation of Sox17 immunostaining in retinal sprouts (b) and artery (b, d) (n = 5 control and 5 Ephb4iΔEC and n = 5 control and 5 Efnb2iΔEC) and percentage of cells with high Sox17 immunosignal vs total ECs (n = 3 control and 3 Ephb4iΔEC and n = 3 control and 3 Efnb2iΔEC) (b, d). e Violin plots of SOX7, SOX17 and SOX18 gene expression from scRNA-seq experiment comparing siControl and siEPHB4 cells. f SoxF factors reduce EPHB4 and increase EFNB2 transcripts (n = 3 experiments). g, h Immunoblot analysis and quantitation of EphB4 and ephrin-B2 protein upon SOX7 + SOX17 + SOX18 knockdown (n = 3 experiments). i Genome localization of Sox18 putative motifs in EFNB2 and EPHB4 human genes selected for further investigation. j Quantitation graphs for luciferase activity of putative binding regions of Sox18 on EFNB2 and EPHB4 showing direct regulation of EFNB2 expression via Sox18 in two regions (n = 3 experiments). k Increased luciferase activity upon EPHB4 KD compared to siControl for Sox18 binding regions on EFNB2 (n = 3 experiments). l Sox17 expression in ephrin-B2+ sprouts (single confocal z-plane). IB4, GFP and Sox17 staining in P6 Efnb2-H2B-GFP knock-in reporter retinal vasculature. m Graph indicating proportion of Sox17+ cells in ephrin-B2+ tip cells (n = 3 Efnb2-GFP mice). n Schematic representation of interactions between Sox17, ephrin-B2 and EphB4. P values were calculated using two-tailed unpaired t test (b, d, f, h) and one-way ANOVA (j, k). In vivo experiments were performed with tamoxifen injections at P1-P3 and analysis at P6 (a, c). Data are presented as mean values ± SEM. Source data are provided as a Source Data file.
