Negative regulatory sequences in the Ela-inducible enhancer of the adenovirus-2 eariy EIIa promoter

The adenovirus type 2 early EIIa (EIIaE) transcriptional control region exhibits an EIa-dependent enhancer activity (Imperiale et al., 1985, Proc. Natl. Acad. Sci. USA 82, 381-385). We have determined the sequence requirements for this enhancer activity by analysing the enhancing capacity of the entire EIIa promoter region, or portions of it, when inserted approximately 400 bp upstream of the rabbit beta-globin gene. Globin-specific transcription efficiency from the resulting recombinants was measured after transfection into HeLa cells, both in the presence and absence of the EIa products. It was found that the minimal EIIa element with bidirectional, EIa-dependent enhancer activity extends between -111 and -27 relative to the EIIaE major startsite (+1). Furthermore an extensive deletion analysis revealed, within this element, three functionally distinct regions: a central region between about -90 and -70, corresponding to an essential EIIaE upstream promoter element, and two flanking control elements (about 20 bp each) which, in the absence of the EIa products, exert a negative effect on the enhancer activity. Deletion of either one of these control elements renders the EIIaE enhancer activity constitutive, suggesting that the EIa products stimulate the EIIaE enhancer by relieving the negative control mediated by these sequences.