Molecular mechanisms of the negative effect of insulin-like growth factor-I on growth hormone gene expression in MtT/S somatotroph cells

Although insulin-like growth factor-I (IGF-I) is shown to have a suppressiv e effect on GH gene expression at the pituitary level, its molecular mechan ism has not yet been clarified. To study the issue, we established a new in vitro system using MtT/S, a recently established rat somatotroph tumor cel l line that retains the basic characteristics of somatotroph function. Plas mids containing the GH 5' promoter (similar to 1.75 kb or shorter)-lucifera se fusion gene were transfected stably or transiently into the cells, and t he effect of IGF-I on the GH promoter activity was estimated by a luciferas e assay. The results showed that IGF-I inhibited GH promotor activity (more than 50% suppression) in a time- and dose-related manner. IGF-I also inhib ited GH secretion. A study using deletion mutants of the GH promoter reveal ed that the negative effect was maintained in the shortest construct (-80 t o +6), suggesting that IGF-I-related factor is acting at the region very cl ose to the minimal promoter, Interestingly, the negative effect was complet ely eliminated by a PI3 kinase inhibitor wortmannin (1 mu M), whereas a MAP kinase inhibitor PD98059 (20 mu M) or 86 kinase inhibitor rapamycin (10 nM ) did not influence the effect. Our results suggest that IGF-I suppresses G H gene expression at the transcriptional level and that the PI3 kinase-medi ated signaling pathway plays a major role in the negative effect of IGF-T. We believe that our system using MtT/S cells is an excellent experimental m odel system for studying the cellular and molecular mechanisms of the trans criptional regulation of GH in vitro.