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Fig. 6 Bmp signaling is downstream of Wnt signaling in the maintenance of dorsal retinal markers. (A-F) Bmp4 can rescue dorsal retinal markers in the absence of Wnt signaling. hs:Dkk1 and control wild-type (WT) embryos at the one-cell stage were injected with a construct that expresses Bmp4 upon heat shock (pDestTol2pA2;hsp70l:bmp4-IRES-GFP), heat shocked at 12 hpf, and fixed at 24 hpf. (A,B) To illustrate transgene expression following heat shock, in situ hybridization was performed for gfp. Widespread clonal expression was observed in the retinas in 85% of embryos. (C-F) Expression of Bmp4 caused a clear expansion of tbx5 in embryos not expressing Dkk1 (D) and the rescue of tbx5 in embryos expressing Dkk1 (F). (G-L) The activation of Wnt signaling does not rescue dorsal markers in the absence of Bmp signaling. Embryos heterozygous for the Tg(hsp70l:nog3)fr14 transgene, which express the Bmp pathway inhibitor Noggin upon heat shock, were outcrossed to TL strain fish and placed in 200 mM LiCl at 18 hpf. A 2-hour heat shock was performed at 18 hpf, and embryos were fixed at 24 hpf. To illustrate Wnt pathway activation, TOP:dGFP embryos were similarly treated with LiCl from 18-24 hpf and gfp detected by in situ hybridization (G,H). For hs:Noggin embryos untreated with LiCl, 55% of 49 embryos lost expression of tbx5 (J) and, for embryos treated with LiCl, 57% of 122 embryos lost expression of tbx5 (L), showing that the activation of Wnt signaling cannot rescue tbx5 in the absence of Bmp signaling. Lateral views, dorsal up, anterior left.