Identification of an insulin-responsive element in the rat insulin-like growth factor-binding protein-3 gene.
The hepatic expression and serum levels of insulin-like growth factor-binding protein-3 (IGFBP-3) are decreased in insulin-dependent and insulin-resistant diabetes. Insulin increases hepatic IGFBP-3 expression by enhancing gene transcription. This report identifies sequences within the IGFBP-3 promoter that are necessary and sufficient for the response to insulin in hepatic nonparenchymal cells. By transient transfection, we mapped the insulin response element to the -1150 to -1124 base pair (bp) region of the rat IGFBP-3 promoter. Three tandem repeats of the -1150 to -1117 bp region conferred insulin responses in a heterologous promoter. Gel shift analyses revealed a 3-fold increase in DNA-protein complex formation with nuclear extracts obtained from insulin-stimulated nonparenchymal cells compared with cells incubated without insulin and revealed 3-4-fold decrease in complex formation with nuclear extracts obtained from the livers of streptozotocin-diabetic rats compared with control rats. Mutational analysis of this 34-bp region showed a core sequence of 10 bp (-1148 to -1139) that is critical for interaction with insulin-induced trans-acting factors. Southwestern blotting revealed a approximately 90-kDa protein that was increased 2-3-fold by the addition of insulin. Thus, we have identified cis-acting DNA sequences that are responsible for regulation of IGFBP-3 transcription by insulin and essential for binding of insulin-responsive nuclear factors.
Villafuerte
B C
BC
Division of Endocrinology and Metabolism, Department of Medicine, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Zhao
W
W
Herington
A C
AC
Saffery
R
R
Phillips
L S
LS
D000818:Animals; D001665:Binding Sites / Q000235:genetics; D007328:Insulin / Q000378:metabolism*; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000235:genetics* / Q000378:metabolism; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D017422:Sequence Analysis, DNA
D007328:Insulin; D018972:Insulin-Like Growth Factor Binding Protein 3