Fig. 1. (A) Insulating activity within the Hoxd13 to Evx2 intergenic region. The posterior extremity of the Hoxd complex is shown, as well as the position of the Evx2 gene. The expression patterns of both Evx2 and Hoxd13 are depicted above to illustrate enhancer sharing in developing digits (right), whereas expression in the central nervous system (CNS) and spinal cord is detected only for Evx2 (left). The transcriptional orientation of this latter gene is opposite to that of all Hoxd genes (arrows). Enhancer sequences driving Evx2 in various domains of the developing CNS are located downstream the gene, i.e. 5' to the Hoxd cluster; hence, an insulating property is expected to lie between the two promoters (red bar). This was further supported by the relocation of a Hoxd9/lacZ transgene at different positions upstream the cluster (B). When relocated between Evx2 and Hoxd13 (right panel) the transgene was expressed in distal limbs but not in CNS. By contrast, when relocated downstream Evx2 (left panel), the transgene was expressed in both distal limbs and CNS, in a way much related to the Evx2 pattern, demonstrating that Hox promoters can indeed respond to these controls, if placed at an appropriate position.

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