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Fig. 6. Expression of XER81 requires FGF signaling. (B) 500 pg of dominant-negative FGF receptor (XFD) mRNA were injected radially in the marginal zone at the 4 cell stage. The embryos were cultured until stage 10.5, fixed in Memfa and hybridized with a digoxigenin-labeled XER81 antisense RNA probe. XFD blocks the expression of XER81 at the sites of injection. (A) Unperturbed XER81 staining in the marginal zone is seen in uninjected control embryos. (C) Experimental design of the animal cap experiment. 200 pg of full length FGF receptor (XFR) or 400 pg of XFD were injected into the four animal blastomeres at the 4 cell stage. Animal caps of injected and not injected embryos were dissected at stage 8.5 and cultured in the presence of bFGF until stage 10.5. The explants were transferred to bFGF-free medium and cultured until stage 14. RNA was isolated and RT-PCR was performed to analyze the expression of XER81, Xbra as a marker and EF-1a as a loading control. 1/17 embryo- or 0.2 animal cap equivalents were used for each PCR reaction. (D) bFGF induces the expression of XER81 and, as expected, of Xbra (lanes 2, 3), but not in the presence of dominant-negative FGF receptor (lanes 4, 5). Addition of the full-length FGF receptor rescues the inducing activity of FGF in the presence of XFD (lane 6). |
