Identification and characterization of high glucose and glucosamine responsive element in the rat osteopontin promoter

Abstract 

We have previously reported that high glucose stimulates osteopontin (OPN) expression via a protein kinase C-dependent pathway and a hexosamine pathway in cultured rat aortic smooth muscle cells (SMCs) [Biochem. Biophys. Res. Commun. 258 (1999) 722.]. In the present study, we carried out functional OPN promoter assays using the luciferase expression vector system in cultured rat aortic SMCs to determine a high glucose/glucosamine responsive element. An extensive deletion analysis of the 5′-flanking region of the rat OPN gene revealed that an element involved in high glucose and glucosamine responses was present within a region between −112 and −62 bp of the OPN promoter. This region is highly conserved in the rat, mouse, and human promoters and contains a number of consensus regions, including an E-box and a GC-rich region. Mutation of the E-box or the GC-rich region resulted in a significant loss of both high glucose and glucosamine responses. These results suggest that two cis-acting elements, the E-box and the GC-rich region, are involved at least partly in high glucose/glucosamine-stimulated transcription of the rat OPN gene.

Keywords:  Diabetic macroangiopathy, E-box, GC-rich region, High glucose/glucosamine responsive element, Osteopontin, Vascular smooth muscle cells

Abbreviations:  OPN, osteopontin, PAI-1, plasminogen activator inhibitor-1, SMCs, smooth muscle cells, USF, upstream stimulatory factor