Figure 1: FGF signalling is required for HSC specification during mid-somitogenesis stages.

(a–d) hsp70:dn-fgfr1 transgenic embryos, heat induced at 12 hpf (38 °C, 20 min) visualized by WISH for HSC markers runx1 (black brackets) (a,b) and cmyb (black brackets and yellow arrowheads in high magnification images) (c,d) in aortic haemogenic endothelium compared with wt embryos. (e) runx1 phenotype percentages in hsp70:dn-fgfr1 animals following heat-shock at the indicated stages (12 hpf, n=82; 15 hpf, n=26;17 hpf, n=38). (f) Quantification of 35 hpf cmyb⁺ cells (yellow arrowheads in c,d) in the DA of wt and hsp70:dn-fgfr1 induced at 12 hpf (wt, n=42; hsp70:dn-fgfr1, n=61; *P-value<0.00001, significantly different from wt, Student’s t-test), 15 hpf (wt, n=52; hsp70:dn-fgfr1, n=56), and 17 hpf (wt, n=48; hsp70:dn-fgfr1, n=54). Red lines indicate mean±s.e.m. (g,h) Confocal microscopy images of 48 hpf double-positive HSCs (white arrowheads) in cmyb:EGFP; kdrl;RFP; hsp70:dn-fgfr1 animals following heat-shock at 12 hpf. (i) Quantification of 48 hpf cmyb⁺ kdrl⁺ double-positive HSCs in the DA of wt and hsp70:dn-fgfr1 animals induced at 12 hpf (wt, n=15; hsp70:dn-fgfr1, n=13; **P-value<0.005, significantly different from wt, Student’s t-test). (j–m) Lateral and ventral view of WISH images using hsp70:dn-fgfr1 transgenic embryos heat-induced at 12 hpf for rag1 at 4 dpf (j,k) and cmyb at 5 dpf (l,m) in thymus (black arrows) compared with wt embryos. (n–p) Confocal microscopy images of hsp70:dn-fgfr1 embryos heat-shocked at 12 hpf. Expression of dominant-negative Fgfr1-EGFP at 13 hpf (n,o) and 15 hpf (p). (o) The magnified region boxed in yellow shown in (n). (q) WISH for the FGF target pea3 in wt (top row) and hsp70:dn-fgfr1 embryos (bottom row) at 13, 14, and 15 hpf after heat-shock at 12 hpf. Scale bar=100 μm.