IDENTIFICATION OF AN INSULIN-RESPONSIVE ELEMENT IN THE PROMOTER OF THE HUMAN GENE FOR INSULIN-LIKE GROWTH-FACTOR BINDING PROTEIN-1

Insulin inhibits the hepatic transcription of insulin-like growth fact or binding protein-1 (IGFBP-1). In the present studies, human HEP G2 h epatoma cells were transiently transfected with human IGFBP-1 gene pro moter constructs in order to identify cis elements and trans-acting fa ctors that confer the insulin effect. Transfections of IGFBP-1 promote r deletion constructs localized an insulin responsive element (IRE) be tween approximately 140- and approximately 103-base pair (bp) 5' to th e mRNA capsite. This region contains a 25-bp sequence which is 100% co nserved in the rat IGFBP-1 promoter and which has two AT-rich, 8-bp el ements exhibiting dyad symmetry. Site-directed mutagenesis of both ele ments in the same 1205-bp IGFBP-1 promoter construct abolished the inh ibitory effect of insulin on promoter activity. Also, the native but n ot the mutant IGFBP-1 IRE conferred the inhibitory effect of insulin t o the heterologous thymidine kinase promoter. Gel mobility shift assay s identified a DNA binding activity which specifically binds the nativ e IGFBP-1 IRE and which is not altered by prior insulin treatment. The IGFBP-1 IRE sequence is similar to those of functionally mapped IREs from other gene promoters, suggesting that this common IRE and the pro tein(s) which it binds confer the insulin effect to a number of insuli n-sensitive genes.