RAPID ACTIVATION OF RAT INSULIN-LIKE GROWTH-FACTOR-I GENE-TRANSCRIPTION BY GROWTH-HORMONE REVEALS NO ALTERATIONS IN DEOXYRIBONUCLEIC-ACID PROTEIN INTERACTIONS WITHIN THE MAJOR PROMOTER

Insulin-like growth factor-I (IGF-I) is an important mediator of prena tal and postnatal growth, but little is known about the control of IGF -I gene expression. Previously, we demonstrated that GH rapidly stimul ates hepatic IGF-I transcription in vivo in hypophysectomized (hyper) rats. In this study, we show that GH induces IGF-I gene transcription through the major promoter, promoter 1, and identify and characterize DNA-protein interactions throughout the promoter. In vitro deoxyribonu clease-I footprinting was used to analyze 1711 nucleotides of promoter 1 and the entire 328-nucleotide 5'-untranslated region of exon 1, usi ng hepatic nuclear protein extracts from male juvenile hypox rats give n a single ip injection of GH or saline 60 min before death. Fourteen DNA-protein binding sites were identified, with 6 located in the highl y conserved 5'-untranslated region of exon 1. These latter sites were further characterized for specificity and regulation by GH, using gel mobility shift assays. Two of these DNA-protein interactions were also detected by in vivo dimethylsulfate footprinting. All DNA-protein bin ding was seen using hepatic nuclear protein extracts from hyper rats a nd did not change within 15, 30, 60, or 120 min after treatment with G H. Our results thus define a series of constitutive DNA-protein intera ctions within the major rat IGF-I gene promoter that may be involved i n mediating GH-activated nuclear signals to initiate IGF-I transcripti on.