Fig. 4. Two Hox binding sites within Box2 are required for XC enhancer activity. Functional requirement of Hox6/7 sites for XC or XC([Hox/Pbx2-3-4]) enhancer activity visualized by in situ hybridization to lacZ transcripts. All embryos have been processed under the same conditions, with identical staining times. All panels show magnifications of lateral views of the midgut of stage 14 embryos. The embryo in A bears a wild-type copy of the XC enhancer and serves as a reference for the activity of XC variants. The deletion of Box2 (B), or the mutation of the two Hox binding sites (Hox6/7) found in Box2 (C), results in a strong diminution of XC enhancer activity. The activity of the XC([Hox/Pbx2-3-4]) enhancer (D) is stronger than that of the full-length XC (A), and is also significantly diminished upon mutation of the Hox6/7 sites (E).