FGF Signaling Maintains a Sox9b- Expressing Population in Segment Centers (A–L) In situ hybridization for erm, fgfr2, neurog1, or neurod4 (red), followed by immunostaining with anti-Sox9 antibody (green). Images shown are a projection of confocal stacks. Scale bar, 50 μm. White arrowheads indicate segment centers; yellow arrowheads indicate colocalization of Sox9b with fgfr2 and erm. The staining in segment centers is due to Sox9b, as it is lost in Sox9b morphant embryos (not shown). (M and N) Whole-mount immunostaining of Sox9b in 36 hpf wt (M) or transgenic dominant-negative fgfr1 embryos (Tg(hsp70l:dnfgfr1-EGFP) [N]). White arrowheads indicate segment centers in wt embryos; open arrowheads in transgenic embryos point at centers where Sox9b expression is absent. Images shown are merged confocal stacks. Scale bar, 20 μm. (O–R) In situ hybridizations of 26 hpf wt embryos (left) or embryos expressing constitutively active FGFR1 (Tg(hsp70:ca-fgfr1)), using erm (O and P) or sox9b (Q and R) probes. Embryos were heat shocked at 24 hpf and fixed 2 hr later. Arrowheads indicate segment centers; red arrowheads indicate upregulation of erm or sox9b expression. Scale bar, 50 μm. See also Figure S2.
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