Disruption of promoter-enhancer communication leads to engrailed mutants

In Drosophila, the invected (inv) and engrailed (en) genes exist within a co-regulated complex and are expressed throughout early development. Although the inv/en promoters are separated by 54 kb, their expression is regulated by the same enhancers distributed across a 70 kb region, suggesting that enhancers can activate multiple promoters over long distances. Previous studies have identified a 2 kb regulatory fragment upstream of the en promotor, which may serve as a promoter tethering element by facilitating interactions between the en promoter and distant enhancers. We have generated a transgenic line containing the 2 kb regulatory fragment fused to a β-galactosidase reporter gene inserted near the en promoter. When coupled with a wild-type chromosome, transgenic organisms expressed β-galactosidase and En only in the posterior compartment of the wing imaginal discs, consistent with appropriate enhancer communication. However, in the absence of a wild-type chromosome, transgenic organisms exhibited impaired imaginal disc development in addition to a de-repression of β-galactosidase in the anterior compartment of the wing imaginal discs, suggesting the endogenous enhancers have been hijacked by the regulatory fragment within the transgene. Taken together, our data suggest that a specific regulatory fragment may serve as a promoter tethering element and is required to facilitate interactions between the en promoter and imaginal disc enhancers at discrete developmental stages.