DEV02815 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure 1 -

  Fig. S1. Alignment of mouse and chick CYP1B1 protein sequences. Conserved residues are designated by the corresponding single letter amino acid code in blue. The partial cDNA sequence of ChEST107a9 (603130733F1; www.chick.umist.ac.uk) was translated in the appropriate frame and aligned with mouse CYP1B1 sequence (SwissProt Q64429); an 84% similarity was observed, confirming the identification of the chick CYP1B1 ortholog.

• Supplemental Figure 2 -

  Fig. S2. Cyp1B1 is not induced by FGF8 protein in the developing midbrain. (Aa-Ac) Lateral view of a chick embryo processed for Cyp1B1 expression, where an FGF8-coated bead was implanted into the midbrain (arrow) at HH10 and left for 16 hours. (Ba-Bc). Lateral view of a chick embryo processed for Cyp1B1 expression following similar treatment with a PBS-soaked bead in the midbrain. In either case, Cyp1B1 expression could not be induced (n=5).

• Supplemental Figure 3 -

  Fig. S3. Embryos electroporated with mCyp1B1-IRES-GFP and analysed for Shh and GFP expression. Embryos were electroporated with mCyp1B1-IRES-GFP at HH10, cultured for 16 hours and analysed by in situ hybridisation for Shh (Aa,Ba,Ca) and GFP expression (Ab,Bb,Cb); merged images are shown (Ac,Bc,Cc). The black lines indicate the region where Shh transcripts are downregulated in the floorplate and notochord. The white arrows show regions of high GFP expression demonstrating where Cyp1B1 is overexpressed in the neural tube. These data suggest that Cyp1B1 is producing a signal that can act non-cell-autonomously to repress the expression of Shh.