Fig. 2

Fig. 2

Regulatory interactions of pou5f1, fgf8a and bmp2b. Whole-mount in situ hybridization of wild-type (WT) embryos and MZspg mutants, injected with mRNAs as indicated. (A–D) WT embryos were left non-injected, or were injected with Pou5f1-VP16 (pou-vp) or Pou5f1-Engrailed (pou-en) encoding mRNAs, fixed at 40%–50% epiboly stage, and stained with the probes as indicated on the left sides of the panels. Animal views. Note the gap in the marginal ring of expression of no tail, ndr1 and fgf8a for Pou5f1-VP16 overexpressing embryos, and local thickening of the marginal expression ring for Pou5f1-Engrailed overexpressing embryos (B–D). (E, F) Sphere stage WT embryos and MZspg mutants probed with fgf8a. fgf8a is expanded in MZspg embryos. (C) Animal view, (D) lateral view. (G) WT embryos were left non-injected (left), or were injected pou5f1-En mRNA (43 pg; right) and probed with fgf8a. fgf8ais expanded in pou5f1-En mRNA injected embryos (7, n = 8). Animal view, sphere stage. (H) WT embryos were left non-injected (left), or were injected with fgf8a mRNA (3 pg/embryo; right) and probed with bmp2b. bmp2b expression is reduced in 12 out of 14 fgf8a mRNA injected WT embryos analyzed. Lateral view, 30% epiboly. (I, J) FGF signaling is functional in blastula MZspg embryos and can suppress bmp2b expression. MZspg embryos were left non-injected (left), or were injected with fgf8a mRNA (3 pg/embryo) and probed with bmp2b (G) or vox (H). (G) bmp2b expression is further reduced in fgf8a mRNA injected MZspg embryos (13, n = 14 embryos). (H) vox expression did not significantly change. Lateral views 30% epiboly. (K, L) Inhibition of FGF signaling restores bmp2b expression at sphere stage in MZspg embryos. To inhibit FGF signaling, WT (K) and MZspg (L) embryos were treated with 40 µM SU5402 (or controls with DMSO only) starting from 2-cell stage, fixed at sphere stage, and probed with bmp2b.