FGF blocks erythroid gene expression by inhibiting its induction by BMP. (A) Erythroid gene expression is induced in caps when FGF signaling is blocked during gastrulation; 400 fg activin mRNA was injected into the animal pole of single cell embryos. At stage 8, animal caps were excised and cultured in the presence of 50 μM SU5402 or buffer from stage 8, 10, 11.5, or 12.5 until sibling embryos reached stage 18. Caps were processed for real-time RT-PCR probing for Scl (i), Runx1 (ii), Fli1 (iii), and Flk1 (iv). Ui- indicates uninjected; act, activin. Values represent the average of 3 experiments; error bars represent SD. (B) The induction of Scl and Runx1 by activin + XFD is BMP-dependent; 400 fg activin mRNA was injected with or without 1 ng tBR mRNA and 400 pg XFD mRNA. Animal caps were excised at stage 8 and cultured until sibling embryos reached stage 18. Explants were snap frozen and processed for real-time RT-PCR probing for Scl (i), Runx1 (ii), or Fli1 (iii). Ui indicates uninjected; tBR, truncated dominant negative BMP receptor; act, activin. Values represent the average of 3 experiments; error bars represent SD. (C) FGF expressed in presumptive ventral ectoderm late during gastrulation blocks Scl but not Fli1 expression in the anterior hemangioblast. Embryos were injected with 10 ng pCSKAefgf DNA either into both A4 blastomeres (ii,v) or both A1 blastomeres (iii,vi) at the 32-cell stage. Correctly targeted embryos were identified using coinjected GFP mRNA, and these embryos were grown to stage 17 alongside uninjected controls (i,iv) then probed for Scl (i,iii) or Fli1 (iv,vi) expression by whole-mount in situ hybridization. Scl, but not Fli1, expression was blocked when A4 was targeted (ii,v), whereas expression of both genes was unaffected when A1 was targeted (iii,vi). Numbers of embryos assayed is recorded in bottom right of each panel.