PURIFIED DROSOPHILA TRANSCRIPTION FACTOR, ADH DISTAL FACTOR-I (ADF-1), BINDS TO SITES IN SEVERAL DROSOPHILA PROMOTERS AND ACTIVATES TRANSCRIPTION

Adh distal factor-1 (Adf-1) is a sequence-specific DNA-binding activity originally identified in Drosophila tissue culture cells and embryos. Adf-1 binds to upstream recognition elements in each of the two promoters of the Drosophila alcohol dehydrogenase gene (Adh), and binding of Adf-1 to the Adh distal promoter site activates transcription. We have carried out a mutational analysis of the Adh distal promoter using both an in vitro transcription assay and a transient transfection assay in Drosophila tissue culture cells, and in both cases find that deletion of sequences required for Adf-1 binding leads to a 3-4-fold drop in transcription. We have purified Adf-1 and demonstrate by a sodium dodecyl sulfate-gel renaturation assay that it is a 34-kDa protein. Purified Adf-1 activates Adh distal promoter transcription in vitro in a binding site-dependent manner. DNase I footprint analysis shows that the purified protein binds not only to the two previously characterized sites in Adh but also to transcriptional regulatory elements in the dopa decarboxylase (Ddc) and Antennapedia (Antp) P1 promoters. Thus, it appears that Adf-1 may play an important role not only in the regulation of Adh expression but also in the transcription of other Drosophila genes as well.