Fig. 2.
Rspo2 depletion promotes FGF signaling. (A,B) Two-cell embryos were injected animally with RspoMOATG (10 ng) or RspoMOSB (20 ng). Ectoderm explants were dissected at stage 8, treated with 25 ng/ml of FGF2 and cultured until stage 13. (A) Representative morphology of the embryos. (B) Quantification of the data in A, representative of two independent experiments. (C) RT-qPCR shows enhanced tbxt expression in FGF-stimulated ectodermal explants (stage 13) after Rspo2 depletion. (D,E) Enhanced cdx4 and msgn1 expression in the dorsal marginal zone (DMZ) explants depleted of Rspo2. RT-qPCR was carried out in stage 13 DMZ explants that were isolated at stage 10. Data are means±s.d. for triplicate samples. Graphs are representative of three independent experiments. (F) In situ hybridization with antisense cdx4 probes was carried out with stage ≥10 control embryos and embryos injected marginally four times with 10 ng of RspoMOATG. The number of embryos with the displayed phenotype and the total number of injected embryos are shown. (G) Two dorsal animal blastomeres of four-cell embryos were injected with RMOATG or RMOSB (10-20 ng each). Representative embryos are shown at stage 39. Arrowheads point to the eye (white) and the cement gland (black). The graph presents frequencies of embryos with head defects (missing eyes, cement gland and reduced facial structures). Numbers of embryos per group are shown at the top of each bar. Data are representative of three to four independent experiments. **P<0.01, *P<0.05, Student's t-test. Scale bar: 300 µm.