|
|
|
|
|
|
|
||||
| Home Help Feedback Subscriptions Archive Search Table of Contents | ||||
| ||||||||||||||||||||
Files in this Data Supplement:
Fig. S1. X-TSK functions as a BMP antagonist in Xenopus embryos. (A-F) Dorsalization of ventral mesoderm by X-TSK. (A,D) Ventral marginal zone (VMZ) explants injected with X-TSK mRNA (1.6 ng) were dissected at stage 10 and cultured until stage 24. (B,E) Control VMZ explants. (C,F) Animal caps (ACs) injected with Xnr1 (50 pg; left) or Xnr1 together with X-TSK (0.5 ng; right). (E) The inset shows expression of cardiac actin in a normal embryo at stage 25. (D-F) In situ hybridization with cardiac actin. Explants injected with X-TSK showing extensive elongation and strong cardiac actin expression. (G-L) X-TSK functions as a BMP antagonist. (G,J) Control dorsal marginal zone (DMZ) explants. (H,K) DMZ explants injected with BMP4 (1 ng) dissected at stage 10 and cultured until stage 24. (I,L) DMZ explants injected with BMP4 (1 ng) + X-TSK (2 ng). (J-L) In situ hybridization with cardiac actin. Inhibition of cardiac actin expression by BMP4 was rescued by X-TSK.
Fig. S2. X-TSK induces anterior molecular markers in animal caps. (A-O) Results of the animal cap assays. (A,D,G,J,M) Expression of markers in normal embryos at stage 21/22. Animal caps (ACs) prepared from stage 8-9 embryos injected with 3 ng X-TSK mRNA (B,E,H,K,N) or controls (C,F,I,L,O). ACs were harvested at the time equivalent to stage 21/22 and hybridized with XAG-1 (A-C), Xotx5b (D-F), Xotx2 (G-I), Sox2 (J-L) and cardiac actin (M-O) probes. X-TSK induced the expression of cement gland and neural markers.
Fig. S3. X-TSK siRNA inhibits formation of the neural crest derivatives. (A-J) Melanocyte development in siRNA injected embryos. (A,B) Whole-mount in situ hybridization with a probe for Xenopus Trp-2, a marker of early melanocyte differentiation. (A) Trp-2 expression was not observed on the X-TSK-si injected side at stage 25. (B) Normal Trp-2 expression in the differentiating malenocytes was observed on the control side (arrows). (C,D) Pigmented melanocytes were observed from stage 34. (C) Pigmented cells were observed on the uninjected side (left) but not on the X-TSK-si injected side (right). (D) Pigmented melanocytes (arrows) are visible both on the uninjected (left) and C-TSK-si injected side (right). (E,F) Corresponding fluorescent images of C and D, showing the distribution of co-injected GFP mRNA. (G,H) Distributions of mature melanocytes at stage 42. (G) Melanocyte were observed on the uninjected side (left) but not on the X-TSK-si injected side (right). (H) Melanocytes (arrows) are visible both on the uninjected (left) and C-TSK-si injected side (right). (I,J) Corresponding fluorescent images of G,H showing the distribution of co-injected GFP mRNA. (K-R) The effect of siRNA on the cranial facial structures derived from cranial neural crest cells. (K,L) Whole-mount in situ hybridization analysis of Sox9 as a cranial neural crest marker in stage 25 embryos. (K) The expression of Sox9 in the mandibular segment was not observed on the X-TSK-si injected side. (L) Sox9 expression was detected in the mandibular (m, open arrow), anterior branchial segment (aB) and posterior branchial segment (pB) on the uninjected side. (M,N) Gill formation at stage 34. (M) Gills on the X-TSK-si injected side (left) were smaller than on the control side (right). (N) Gills were unaffected in C-TSK-si injected tadpoles. (O,P) Fluorescent images corresponding to showing the distribution of co-injected GFP mRNA (M,N). (Q,R) Alcian Blue/Alizarin Red staining of Xenopus tadpoles at stage 45. (Q) Branchial arch cartilages on the X-TSK-si injected side (left, inj.) were smaller than on the control side (right). (R) C-TSK-si did not affect the skeleton formation.
| ||||||||||||||||||||
