BINDING OF NUCLEAR FACTORS TO FUNCTIONAL DOMAINS OF THE DUCK HEPATITIS-B VIRUS ENHANCER

We have analyzed the structures, relative organization, and activities of binding sites for nuclear factors in the duck hepatitis B virus (d uck HBV) enhancer. DNase I footprinting analysis and mobility shift as says demonstrate that this enhancer of 192 bp contains at least three binding sites for transcription factors: one for hepatocyte-adipocyte C/EBP, a second for the liver-specific transactivator hepatocyte nucle ar factor 1 HNF-1, and a third for a factor, called F3, which binds to a DNA sequence bearing some resemblance to that for the ubiquitous fa ctor EF-C. Analysis of transcriptional activity reveals that oligonucl eotides corresponding to the individual binding sites, inserted upstre am from a heterologous promoter, display very weak enhancer activity, whereas the enhancer encompassing these three sites displays very high activity. Analysis of duck HBV enhancer mutants indicates that the de letion of any of these sites leads to a modification of transcriptiona l enhancer activity. The hepatocyte nuclear factor 1 binding site is c rucial, since an internal deletion of 14 bp abolishes the activity. Th e C/EBP site can act as repressor, and the F3 site is required for ful l activity. Comparative analysis reveals that the nuclear factors are similar to those bound to the human HBV enhancer but that the organiza tion of their binding sites in the duck HBV enhancer is different.