Insulin-like growth factor-1 increases skeletal muscle dihydropyridine receptor α1S transcriptional activity by acting on the cAMP-response element-binding protein element of the promoter region

Abstract

Previous work from our laboratory has shown that insulin-like growth factor 1 (IGF-1) increases the expression of the skeletal muscle dihydropyridine receptor (DHPR) α1 subunit by regulating DHPR α1S nuclear transcription. In this study, we investigated the mechanism by which IGF-1 enhances expression of the DHPR α1S gene. To this end, the promoter region of the mouse DHPR α1S gene was recently cloned and sequenced and various promoter deletion-luciferase reporter constructs were used. These constructs were transfected into C2C12 cells and IGF-1 effects were measured by recording luciferase activity. IGF-1 significantly enhanced DHPR als transcription in those constructs carrying cAMP-response element-binding protein (CREB) binding site but not in CREB core binding site mutants. Gel mobility shift assay using a double stranded oligonucleotide for the CREB site in the promoter region, and competition experiments with excess unlabeled or mutated promoter oligonucleotide, and unlabeled consensus CREB oligonucleotide demonstrated that IGF-1 induces CREB binding to the DHPR α1S promoter. IGF-1-mediated enhancement in charge movement was prevented by incubating the cells with antisense but not with sense oligonucleotides against CREB. These results support the conclusion that IGF-1 regulates DHPR α1S transcription in muscle cells by acting on the CREB element of the promoter.