Upregulation of rat Ccnd1 gene by exendin-4 in pancreatic beta cell line INS-1 : interaction of early growth response-1 with cis-regulatory element

The aim of this study was to investigate the effect of exendin-4 on the expression of cyclin D1 gene (Ccnd1), which is critical in regulating the progression of the cell cycle in INS-1 cells. INS-1 cells were stimulated with exendin-4 (10 nmol/l). Transient transfection and luciferase reporter assays were performed to measure promoter activities of rat Ccnd1. Electrophoretic mobility shift and chromatin immunoprecipitation assays were used to examine the binding of transcription factors to sites responsive to exendin-4 in vitro and in vivo, respectively. Exendin-4 increased both Ccnd1 mRNA and its protein levels in a time-dependent manner. The region from -174 to +130 of the promoter was found to contain cis-regulatory elements responsible for exendin-4-mediated gene induction. Early growth response-1 (EGR1) protein was bound to the region from -153 to -134, which includes the putative EGR1 binding site (5'-CACCCCCGC-3'). Moreover, exendin-4 recruited EGR1 protein to the promoter in vivo. These findings suggest that exendin-4 activates Ccnd1 transcription through induction of EGR1 binding to a cis-regulatory element between -153 and -134 on the rat Ccnd1 promoter. These results provide an important indication that exendin-4 is a growth factor regulating beta cell proliferation.