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. 2006 May 10;26(19):5037–5048. doi: 10.1523/JNEUROSCI.0715-06.2006

Figure 6.

Figure 6.

Sulf1 modifies the distribution of Shh at the surface of neural progenitors of the neuroepithelium and activates Shh signaling pathway in this tissue. A–E, Electroporation of Sulf1 and/or GFP-expressing vectors were performed in E4 embryonic spinal cord that was further dissected, opened at its dorsal aspect, and plated in culture with the neuroepithelial precursors up (A). The electroporated side is represented on the left in all pictures. The distribution of the Shh protein at the surface of neural progenitors was assessed 2 d later by immunodetection on living tissue using the 5E1 antibody (B, C). Electroporation of a control vector expressing GFP (green) does not modify the distribution of Shh, detected as a decreasing ventrodorsal gradient, compared with the control side (B). In marked contrast, coelectroporation with a Sulf1-expressing vector dramatically changes the pattern of Shh by extending the domain of Shh staining (C). The inset in C shows high magnification of Sulf1 electroporated cells that have concentrated Shh at their surface. The dashed lines (B, C) highlight the ventral midline of the floor plate. D, Quantification of staining intensity showing that the decreasing gradient of Shh observed in control explants is modified by Sulf1 overexpression, which leads to concentration of Shh at distance from the midline. E, The readout of Shh signaling was assessed by analyzing ptc expression on transverse section of electroporated explant after 2 d in culture. Double in situ hybridization experiments show strong induction of ptc (purple) through the entire dorsoventral axis of the neuroepithelium in the vicinity of sulf1-overexpressing neuroepithelial cells (brown). F, G, Control and Sulf1-expressing vectors (green) has been electroporated in E1.5 neural tube, and Nkx2.2 pattern of expression (red) was analyzed at E4.5. High magnification of ventral neuroepithelium showing that overexpression of a control vector does not modify the dorsoventral extension of the Nkx2.2-expressing domain (F), whereas this domain extends dorsally in Sulf1-overexpressing spinal cord (G). FP, Floor plate; V, ventral; D, dorsal; NE, neuroepithelium.