Midline signalling distinct from Hedgehog is required for medial fast myoblast myod expression. In situ hybridisation for myod mRNA in embryos without (A,C,E,G) or with SU5402 (B,D,F,H) from yot (A,B,E,F) or flh (G,H) heterozygote crosses or wild type treated with cyclopamine (C,D). (A,B) Control 15 s embryos show reduction of lateral myod expression after SU5402 treatment (B, white arrowheads). (C,D) Cyclopamine prevents maintenance of myod mRNA in adaxial cells (C, arrow), without affecting lateral expression. Addition of both cyclopamine and SU5402 ablates myod mRNA in adaxial (arrow, D) and lateral (white arrowheads, D) cells, leaving medial myod expression. (E,F) In 15 s yot/yot mutant embryos, adaxial myod mRNA is lacking (arrows). Lateral myod expression is severely reduced by SU5402 treatment (F, white arrowheads). (G,H) In flh/flh embryos, myod mRNA is lost from adaxial cells (arrows) but expressed in stripes across the bilaterally fused somites (G, white arrowheads). Additional SU5402 treatment ablates essentially all myod expression in nascent somites (H, white arrowheads). Strong anterior myod expression originating at the midline (red line) may be in recovering slow muscle driven by floor-plate signals (Blagden et al., 1997). (I-M) At 26 s, control (I,K), SU5402-treated (J,L) and ace mutant (M) embryos show maintenance of eng1 mRNA (I,J) and engrailed protein (K-M; single confocal slices with 4D9 anti-engrailed red, nuclei revealed with DAPI in blue) in both MP (arrow, labelled strongly with A4.1025 anti-MyHC green) and MFF (arrowheads). ace mutants were identified by lack of her5PAC:egfp in mid-hindbrain (Tallafuss and Bally-Cuif, 2003).
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