Figure 2

Phosphorylated forms of Akt and ERK1/2 are present in the caudal hindbrain. (a) Western Blot analysis of protein extracts of whole embryos (left column) and isolated caudal hindbrain (right column). Phosphorylated (activated) forms of both Akt and ERK1/2 were present in protein extracts from the caudal hindbrain. Total forms of Akt and ERK1/2 were used as loading controls. pERK1/2 immunodetection is shown in red (b, d-e, h-k) or in green fluorescence in (f). In situ hybridization with vHnf1-Hoxb1 (c) and vHnf1 (g) to position the caudal hindbrain. (i-k) Control experiments to show the specificity of pErk staining: embryos incubated with DMSO (i) or with SU5402 (j), and assayed for pErk (note that specific pErk staining in the MHB and in the caudal hindbrain disappears upon SU5402 treatment); (k) embryos treated as in (i) but without pErk-Ab incubation, as negative control. Whole-mount embryos (b-d, f-g, i-k), or flat-mounted hindbrains (e, h) at indicated stages. ANR, anterior neural ridge; cHB, caudal hindbrain; FB, forebrain; HB, hindbrain; im, intermediate mesoderm; MHB, midbrain-hindbrain boundary; pm, precardiac mesoderm; ps: primitive strike; psm, presomitic mesoderm. Anterior is at the top.