Fig. 3.

Differences in Hoxd13, Hoxd12, and Hoxd11 responses. (A) Shh ligand dose and Hoxd gene expression titration curves. Limb progenitors were treated for 24 h with varying levels of Shh ligand. Fgf8 ligand was supplied at a fixed dose of 120 ng/µL. (B) Fgf8 ligand dose and Hoxd gene expression titration curves. Limb progenitors were treated for 24 h with varying levels of Fgf8 ligand. Shh ligand was supplied at a fixed dose of 1 ng/μL. (C) Limb progenitors were treated in a two-step protocol. In the first step, progenitors were treated with Fgf8 and Shh for 4 h. In the second step, progenitors were treated for 10 h under one of four possible conditions: (i) Fgf8 and Shh, (ii) Fgf antagonist SU5402 and Shh, (iii) Fgf8 and Shh antagonist cyclopamine, or (iv) Fgf antagonist SU5402 and Shh antagonist cyclopamine. The expression levels of each gene were normalized to Actb and depicted as fold change relative to the baseline (no ligand) in A and B. Error bars are as for Fig. 1.