Regulation of insulin-like growth factor I receptor gene expression by Sp1: physical and functional interactions of Sp1 at GC boxes and at a CT element.
The insulin-like growth factor I (IGF-I) receptor mediates signal transduction by the IGFs and plays a critical role in growth and development. The proximal promoter region of the rat IGF-I receptor gene contains multiple Sp1 consensus-binding sites (GC boxes). Various promoter fragments fused to a luciferase reporter gene were transiently cotransfected together with an Sp1 expression vector into Drosophila Schneider cells, which lack endogenous Sp1. A proximal promoter fragment containing 476 nucleotides of 5'-flanking region and 640 nucleotides of 5'-untranslated region was strongly activated by Sp1 (an average of 116-fold), and progressive 5'-deletions of the promoter that sequentially removed GC boxes reduced Sp1 activation to 15-fold over basal promoter activity. DNase I footprinting studies with purified Sp1 protein revealed four GC boxes in the 5'-flanking region of the promoter and one homopurine/homopyrimidine motif (CT element) in the 5'-untranslated region that bound Sp1. Mutation of the CT element reduced Sp1 activation by 70%. Taken together, these results demonstrate that Sp1 can regulate expression of the IGF-I receptor promoter by acting both on GC boxes in the 5'-flanking region of the promoter and on a CT element in the 5'-untranslated region.
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D018983:DNA Footprinting; D003850:Deoxyribonuclease I; D005786:Gene Expression Regulation / Q000187:drug effects*; D008156:Luciferases / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017526:Receptor, IGF Type 1 / Q000235:genetics*; D011993:Recombinant Fusion Proteins; D012045:Regulatory Sequences, Nucleic Acid; D016329:Sp1 Transcription Factor / Q000378:metabolism* / Q000494:pharmacology
D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D016329:Sp1 Transcription Factor; D004247:DNA; D008156:Luciferases; D017526:Receptor, IGF Type 1; D003850:Deoxyribonuclease I