Fig. 5
ALK6 is required for neural crest development. a When compared to control MO injected embryos, ALK6 morphant embryos showed increased pigmentation of the head (arrow), ectopic melanocytes in the dorsal fin (arrowheads) and narrower heads reflected by less lateral protrusion of the gills and smaller distance between the eyes (double arrow). b Cranial cartilage from embryos injected as indicated was stained with Alcian Blue dye. Cartilage from ALK6 morphant embryos generally showed weaker staining, was smaller (double arrows in B’, B” and B”’) and showed partial or complete loss of cartilage elements (arrowheads in B’, B” and B”’) of the gills (g) or ceratohyal cartilage (c). The images in A and B show representative examples from one out of three independent experiments. c Embryos were injected with 1.6 pmol morpholino in one animal-dorsal blastomere at the 8-cell stage as indicated. A lacZ plasmid was co-injected to identify the injected side, embryos were cultured till stage 13 and stained for LacZ. Expression of msx2 was analyzed by whole-mount in situ hybridization. Representative images of embryos injected as indicated are shown. Asterisks indicate the injected side. The results from three independent experiments are summarized in the graph (d). Active BMP signaling at the neural plate border was shown by whole-mount immunostaining for phosphorylated Smad 1/5/8 (pSmad 1/5/8). e The graph shows a summary of pSmad1/5/8 staining results from two independent experiments. In (f) representative images of embryos injected with control MO, ALK3 MO 1 or ALK6 MO 1 are shown. Embryos were categorized according to decreased or enhanced msx2 expression or pSmad 1/5/8 staining respectively and differences between indicated groups were analyzed using the Χ 2 test. Statistically significant deviations (p-value < 0.01) are indicated by double asterisks; n.s.: not significant