E-BOX ACTIVATOR OF THE C4-PROMOTER IS RELATED TO BUT DISTINCT FROM THE TRANSCRIPTION FACTOR UPSTREAM STIMULATING FACTOR

The murine C4 promoter contains a motif (E-C4) active in transcription al activation whose structure complies with the E-box consensus sequen ce recognized by the helix-loop-helix transcription factors. This site is found also in human and rat C4 promoters and has the structure (-7 5) CACGTG (-70) characteristic of the class B subset of b-helix-loop-h elix-zipper proteins. We have challenged the hypothesis that the prote in factor responsible for the E-C4-mediated transcriptional activation is identical to one of the previously characterized nuclear factors. The molecular mass of the E-C4 factor is slightly bigger than that of Hela upstream stimulating factor (USF) (43/44 kDa). Moreover, the nucl eotides immediately adjoining the E-C4 core sequence contribute to the distinctive fine specificity of the E-C4 factor. Optimized USF and MY C DNA-binding sites, which differ in the nucleotides bordering the hex anucleotide box displace the E-C4 factor in competition assays but wit h lesser efficiency than the E-C4 site itself. Finally, the E-C4 facto r fails to exhibit the heat resistance characteristic of USF proteins. The results show that the E-C4 transcription factor has DNA binding p roperties overlapping those of other helix-loop-helix proteins but is structurally distinct from the factors so far described. Although C4 i s not the first liver gene endowed with an E-box-mediated activation, it affords the first example where such activation takes place in the context of a TATA-less promoter, and is functionally linked to an Init iator element-dependent transcription.