BINDING OF A MEMBER OF THE NF1 FAMILY OF TRANSCRIPTION FACTORS TO 2 DISTINCT CIS-ACTING ELEMENTS IN THE PROMOTER AND 5'-FLANKING REGION OF THE HUMAN CELLULAR RETINOL-BINDING PROTEIN-1 GENE

We studied the interaction of nuclear proteins with the 5'-flanking and promoter region of the human cellular retinol binding protein 1 (hCRBP1) gene and identified seven specific sequences that interacted with nuclear proteins from liver and prostate. Two of these sequences, footprint 1 (Fp1) and footprint 5 (Fp5), were highly homologous, sharing the core sequence GGCCAAC, which has a certain similarity to the consensus sequence for the NF1 binding site. Competition experiments in gel mobility shift assays and DNasel footprinting indicated that a common protein interacted with both elements. Immunological and biochemical data indicated that this protein belongs to the nuclear factor 1 (NF1) family of transcription factors. The ability of the Fp1 and Fp5 sequences to control gene expression was studied by transient transfections of several cell types. In the wild type promoter, both Fp1 and Fp5 acted as repressors of human (h) CRBP1 gene transcription. Once inserted upstream of the basal promoter from the heterologous p12 gene, the function of both Fp1 and Fp5 was reverted to that of transcriptional activators, although Fp5 exerted only moderate transcriptional activation of the chloramphenicol acetyl transferase (CAT) reporter gene. Hence, the position of these NF1 binding sites and the nature of the flanking sequences appear to direct their effect on transcription. Despite close sequence homology, a common core sequence, and a similar ability to bind nuclear proteins in vitro, these results indicate that Fp1 and Fp5 exert similar regulatory functions but to different levels in vivo. In conclusion, these results indicate that a member of the NF1 family plays a significant role in regulating CRBP1 gene expression.