REGULATION OF RABBIT ALPHA(1)-ACID GLYCOPROTEIN GENE-EXPRESSION IN ACUTE-PHASE LIVER - IDENTIFICATION OF INDUCIBLE AND CONSTITUTIVE PROTEINS LIKE CCAAT-ENHANCER BINDING-PROTEIN THAT INTERACT WITH THE 5'-PROXIMAL PROMOTER ELEMENTS

To identify the cis-acting DNA sequences responsible for inducible transcription of rabbit alpha1-acid glycoprotein gene, 5'-flanking region containing 529 bp of this gene and its various 5'-deletions were linked to the reporter gene coding for the bacterial chloramphenicol acetyltransferase (CAT) and analyzed for their ability to confer cytokine-mediated inducibility to the reporter CAT gene in liver cells. Deletion analysis has identified a 151-bp region from the sequence -186 to -35, that contains the regulatory promoter element(s) responsible for stimulation mediated by cytokines present in the conditioned-medium. Using mobility shift assays, we have identified highly inducible nuclear factors in acute liver nuclear extract that interact with this regulatory promoter region. DNase I footprint analysis has revealed two adjacent nuclear factor binding sites and competition of DNA-binding activity has indicated that the distal element of these two sites has higher affinity for nuclear factors than the proximal one. Both of these two regions have been found to be capable of directing conditioned-medium-induced transcription. Studies on the characterization of nuclear factors binding to these elements have shown that they belong to a class of transcription factors called CCAAT-enhancer binding protein (C/EBP). Our results indicate that binding of C/EBP-like factors to the inducible promoter elements of rabbit alpha1-acid glycoprotein gene is highly specific and the induction of this gene under acute-phase conditions may involve their participation.