Figure 8. The gypsy insulators directly interact with the enhancer and promoter of the white gene.

(A) Eye phenotypes in flies from the transgenic line heterozygous (P/+) for the construct and in flies after deletion of either the proximal gypsy insulator, or both insulators, or introduction of the mod(mdg4)^u1 mutation. (B) The 3C profile of the E(Gy)^R _4.5W(Gy) line and its derivatives analyzed at the pupa stage for an anchor fragment located in the enhancer or promoter region or downstream of the distal gypsy insulator. The transgene and the surrounding genomic region are drawn to scale. Relative interaction frequencies between DpnII restriction fragments selected for this analysis (gray vertical bands) and anchor regions (purple vertical bands) are shown for the construct (blue-square line), and its derivatives with deletion of proximal gypsy (green-circle line), proximal gypsy at the mod(mdg4)^u1 background (yellow-triangle line), and both gypsy copies (red-cross line). Samples were normalized by qPCR relative to an undigested locus. Error bars show standard deviations from the means of three independent experiments. Transgene DNA is shown as blue line at the bottom of figure, and the boundaries of the transgenes are indicated by blue triangles. The white gene, eye enhancer, and gypsy insulator are represented, respectively, by yellow and green boxes and red pentagon. The lox and frt sites allowing the excision of insulators are indicated by vertical black brackets. Other designations are as in Figure 1.