Juvenile hormone action through a defined enhancer motif to modulate ecdysteroid-activation of natural core promoters

We have established a model system of hormone action, in an Sf9 cell transfection system, using defined enhancer motifs and natural core promoters of metamorphosis-associated genes. The DR1 enhancer, that is an established DNA binding site for the ecdysone receptor/ultraspiracle heterodimer, was necessary for transcriptional activation by 20-OH ecdysone. For this activated transcription, a natural sequence closely 5' to the TATA box is necessary. Cotreatment with juvenile hormone III strongly suppressed the steroid activation of transcription. However, in the absence of the sequence located closely 5' to the TATA box, cotreatment with juvenile hormone instead increased transcription over that occurring due to 20-hydroxy-ecdysone alone. This sensitivity to activation by cotreatment with juvenile hormone could be transferred to a related, but otherwise unresponsive, hexamerin core promoter simply by transferring to the unresponsive promoter the five base transcription start site (ACAGT) from the responsive hexamerin gene. These are the first reports that the direction of JH action on 20-OH ecdysone-activated transcription can be reversed by removal of a sequence at the core promoter, and that modulatory action of juvenile hormone can be transferred to a different gene by transferring the transcription start site motif.