Identification of a distal STAT5-binding DNA region that may mediate growth hormone regulation of insulin-like growth factor-I gene expression.
Growth hormone (GH) regulates insulin-like growth factor-I (IGF-I) gene expression through signal transducer and activator of transcription 5b (STAT5b) and STAT5a. The objective of this study was to identify the cis-regulatory DNA region involved in this process. By cotransfection analyses of shotgun DNA fragments of a bacterial artificial chromosome sequence containing the entire human IGF-I gene and a large 5'-flanking region, a approximately 700-bp DNA region approximately 75 kb 5' to the IGF-I gene was found to have the ability to enhance gene expression from both heterologous and homologous promoters in the presence of constitutively active STAT5a or STAT5b. This 700-bp DNA region contains two closely located consensus STAT5-binding sites, and its sequence appears to be evolutionarily conserved. Electrophoretic mobility shift assays verified the ability of the two putative STAT5-binding sites to bind to STAT5a and STAT5b. Cotransfection analyses confirmed that both STAT5-binding sites were necessary for the 700-bp DNA region to mediate STAT5a or STAT5b activation of gene transcription. Chromatin immunoprecipitation assays demonstrated that the chromosomal region containing these two STAT5-binding sites was bound by constitutively active STAT5b protein in HepG2 cells and that the binding was accompanied by increased expression of IGF-I mRNA. In reconstituted GH-responsive cells, this 700-bp DNA region was able to mediate GH-induced STAT5a or STAT5b activation of gene expression. These results together suggest that this STAT5-binding site-containing distal 5'-flanking region of IGF-I gene may be an enhancer mediating GH-induced STAT5 activation of IGF-I gene transcription.
Wang
Ying
Y
Department of Animal and Poultry Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, USA.
Jiang
Honglin
H
D024506:5' Flanking Region; D001483:Base Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects*; D013006:Growth Hormone / Q000494:pharmacology*; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000235:genetics*; D008894:Milk Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D010957:Plasmids; D050799:STAT5 Transcription Factor; D015534:Trans-Activators / Q000378:metabolism*; D025521:Tumor Suppressor Proteins
D004268:DNA-Binding Proteins; D008894:Milk Proteins; D050799:STAT5 Transcription Factor; C494092:STAT5A protein, human; C494095:STAT5B protein, human; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins; D007334:Insulin-Like Growth Factor I; D013006:Growth Hormone; D004247:DNA