Extended Data Fig. 2: ddx21 is expressed in endothelium and genetically interacts with the Vegfc-Flt4 signalling pathway. | Nature Cell Biology

Extended Data Fig. 2: ddx21 is expressed in endothelium and genetically interacts with the Vegfc-Flt4 signalling pathway.

From: The RNA helicase Ddx21 controls Vegfc-driven developmental lymphangiogenesis by balancing endothelial cell ribosome biogenesis and p53 function

Extended Data Fig. 2

(a) in situ hybridisation for ddx21 showing expression at 4 cell-stage and 24 hpf (scale bars=100μm) and 48 hpf (bottom) lateral view (scale bar=100μm) and transverse section (zoom on right, n = 17, scale bars=20μm). Data shown represent 5 (4 cell), 6 (24 hpf), 16 (left, 48 hpf) and 17 (48 hpf) independent embryos analysed. (b,c) qPCR for ddx21 and endothelial genes on cDNA from sorted arterial endothelial cells (AECs, n = 4), lymphatic endothelial cells (LECs, n = 5 except for n = 4 for prox1a and ddx21) and venous endothelial cells (VECs, n = 4 for 60hpf and n = 5 for 3 dpf) at 60 hpf (b) and 3 dpf (c). Expression relative to kdrl for 60 hpf data and to rpl13 for 3 dpf data. n=biologically independent samples. (d) Confocal projections of TgBAC(ddx21:ddx21-Citrine) transgenics at 36 hpf and 5 dpf. Scale bars= 100μm. Data shown represent 15 independent embryos analysed. (e) Confocal projections showing nucleolar Ddx21-Citrine expression in haematopoietic stem cells (white arrows) in a 36 hpf TgBAC(ddx21:ddx21-Citrine);Tg(fli1a:H2BCherry) embryo (data shown represent 20 embryos analyzed). Image on the right shows TgBAC(ddx21:ddx21-Citrine) expression (grey). Scale bar= 25μm. (f) Confocal projections of an uninjected control sibling, ddx21uq20bh mutant, MO-dll4 injected sibling, and a ddx21uq20bhmutant at 7 dpf. Data shown represent n > 20 sibling control, n > 20 ddx21 mutant control, n = 70 MO-dll4 sibling and n = 67 MO-dll4 ddx21 mutant embryos analyzed in g). Arrows mark the ectopic branching and asterisk the absence. Scale bar= 50μm. (g) Quantification of the number of blood vessel branches per 10 somites in MO-dll4 injected siblings (n = 70) and ddx21uq20bh (n = 67) embryos. p = 0.0012. (h) Representative images for data shown in Fig. 3c. Confocal projections of siblings and ddx21uq20bh mutants in wild type or vegfr3+/- heterozygous backgrounds. Thoracic duct is false-coloured in magenta and asterisk indicate absent lymphatics. Scale bars= 50μm. DA = dorsal aorta, PCV = posterior cardinal vein. Transgenic labels are indicated. Mean values + /-SEM shown. Two-tailed Mann Whitney test for (g).

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