Structure and function of the mouse insulin-like growth factor binding protein 5 gene promoter.

The actions of insulin-like growth factors I and II (IGF-I and -II) are modulated by interactions with one or more of a family of secreted IGF binding proteins (IGFBPs). IGFBP-5, the most conserved of the six known IGFBPs, is a 252-amino-acid protein that has been shown both to potentiate and inhibit IGF action. In previous studies, we have cloned and characterized the mouse IGFBP-5 gene and demonstrated that it is expressed in a hierarchical pattern in different adult mouse tissues and during rodent embryonic development. In this report, we describe the initial analysis of the IGFBP-5 gene promoter. By transient gene transfer studies, we show the orientation-specific activity of DNA fragments containing from 31 to 4,100 bp from the 5'-flanking region of the mouse IGFBP-5 gene in directing expression of the heterologous reporter gene luciferase in Hep G2 cells. DNA fragments with only 156 bp of 5'-flanking sequence mediated over 60% of maximal promoter activity, and a segment containing the TATA box and the first 120 bp of exon 1 still conferred some promoter function. Within the highly active 156-bp region, we identified a 37-bp segment from -70 to -34 that exhibited specific binding in DNase I footprinting and gel-mobility shift experiments with Hep G2 nuclear protein extracts. The footprinted region, which is almost completely conserved in the rat and human IGFBP-5 genes, was responsible for at least 70% of the activity of the intact promoter, as evidenced by the deleterious consequences of small internal deletions within this sequence on promoter function.(ABSTRACT TRUNCATED AT 250 WORDS)