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Fig. 3. Germ layer defects in Coco morphant embryos are caused by an increase in Xnr5/6 and Activin signalling. (A) Western blot analysis demonstrating an increase of P-Smad2 following CocoMO and Activin overexpression, compared with uninjected and ControlMO-injected embryos. A dorsoventral bias of P-Smad2 is detected in normal development (compare dorsal half with ventral half). (B) Coco depletion increases Nodal-induced transcription of a reporter gene. Error bars represent standard errors. Assays were performed in triplicate. (C-N) Embryos were injected with CocoMO, CocoMO+TGFβMO or TGFβMO and compared with uninjected control embryos at stage 9.5. (C-F) Injection of CocoMO causes a shift in Sox17β expression (D, arrows), which is rescued by co-injection with Xnr5/6MO (E). (F) Injection of Xnr5/6MO alone does not affect the Sox17β domain, but reduces the intensity of expression. (G-J′) Injection of CocoMO causes a reduction in Xbra staining in the marginal zone (H,H′), which is clearly rescued by co-injection with ActivinMO (I,I′), an effect different to that of ActivinMO alone (J,J′; compare black arrows in G-J and red arrowheads in G′,I′). (K-N) Injection of CocoMO causes a shift in Sox17β expression (L, arrows), which is also rescued by co-injection of ActivinMO (M, arrows). (N) ActivinMO alone caused a slight reduction in Sox17β. D, dorsal half; V, ventral half. |
