Fig. 8. Kermit is required for neural crest induction by Xfz3. (A) Kermit morpholino antisense oligonucleotide (MO) directed against the 5'UTR of Kermit mRNA blocks translation of Kermit protein. Kermit mRNA was injected into embryos with increasing concentrations of Kermit MO or control MO and embryo lysates were analyzed by western blotting with anti-NIP antibodies (which crossreact with overexpressed Kermit). Translation of Kermit mRNA lacking the 5'UTR was not inhibited by the Kermit MO (20 ng MO; right panel). (B) Depletion of Kermit with antisense MO (4 ng; lane 6) reduces or eliminates neural crest induction by Xfz3 (lanes 4 and 6). Neural crest induction and RT-PCR were performed as in figure 7. Lane 1 is a no reverse transcriptase control; lane 2 shows uninjected control caps (un); lane 3 shows samples expressing chordin alone; lane 4 shows induction of neural crest makers by Xfz3 + chordin; lane 5 shows Xfz3 + chordin + control MO (C); and lane 6 shows Xfz3 + chordin + Kermit-MO (K). (C) Kermit mRNA lacking the 5'UTR rescues inhibition by Kermit MO. Neural crest induction assay was performed as above. Kermit MO (4 ng) was injected (lanes 3, 4) with (+) or without (-) Kermit mRNA (10 pg) lacking the 5'UTR.
