Rapid activation of rat insulin-like growth factor-I gene transcription by growth hormone reveals no changes in deoxyribonucleic acid-protein interactions within the second promoter.
Insulin-like growth factor-I (IGF-I) is a highly conserved 70-residue circulating peptide that mediates many of the systemic growth-promoting effects of GH. This laboratory has found previously that GH rapidly stimulates hepatic IGF-I transcription in hypophysectomized (hypox) rats by activating promoter 1, the major rat IGF-I gene promoter. In this study, the hormonal regulation of IGF-I expression through promoter 2, a minor promoter in most tissues but active in the liver, was investigated. Through use of a sensitive RNase protection assay, GH was shown to rapidly induce the accumulation of correctly initiated transcripts directed by this promoter in hepatic nuclei. Using in vitro DNase-I footprinting, six DNA-protein interactions were identified within promoter 2 with hepatic nuclear extracts from juvenile male hypox rats given a single ip injection of GH or saline 60 min before death. These DNA-protein-binding complexes also were investigated for specificity and for regulation by GH by gel mobility shift assays. All DNA-protein interactions were detected in hepatic nuclear protein extracts from hypox rats and did not change within 15-120 min after GH treatment. These results thus identify and characterize a series of constitutive nuclear protein-binding sites within the second rat IGF-I promoter that may be involved in mediating its transcriptional activity.
Le Stunff
C
C
Department of Internal Medicine, Washington University School of Medicine, Saint Louis, Missouri 63110, USA.
Thomas
M J
MJ
Rotwein
P
P
D000818:Animals; D001483:Base Sequence; D004247:DNA / Q000378:metabolism*; D003850:Deoxyribonuclease I; D005091:Exons; D005786:Gene Expression Regulation / Q000187:drug effects*; D013006:Growth Hormone / Q000494:pharmacology*; D006801:Humans; D007016:Hypophysectomy; D007334:Insulin-Like Growth Factor I / Q000096:biosynthesis* / Q000235:genetics*; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D011401:Promoter Regions, Genetic; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000494:pharmacology; D015183:Restriction Mapping; D014158:Transcription, Genetic / Q000187:drug effects*
D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D013006:Growth Hormone; D004247:DNA; D003850:Deoxyribonuclease I