Osteogenic protein-1 regulates insulin-like growth factor-I (IGF-I), IGF-II, and IGF-binding protein-5 (IGFBP-5) gene expression in fetal rat calvaria cells by different mechanisms.

Abstract

Osteogenic protein-1 (OP-1 or BMP-7) stimulates new bone formation in vivo and induces cell proliferation and differentiation of osteoblasts in vitro. Previous studies from our laboratory revealed that OP-1 led to a two- to threefold increase in steady-state insulin-like growth factor-I (IGF-I) and IGF-II mRNA levels and a fivefold decrease in IGF-binding protein-5 (IGFBP-5) mRNA levels in primary cultures of fetal rat calvaria (FRC) cells. In the present study, we determined whether the effects of OP-1 were at the transcriptional or posttranscriptional level. OP-1 increased the half-life of the IGF-I mRNA from 6 to 17 h without changing the level of IGF-I nuclear pre-mRNA. In transiently transfected FRC cells, the luciferase activity driven by the -1122/+362 or the -133/+362 IGF-I exon 1 promoter fragment was not changed by OP-1. Similar results were observed using the -1500/+44 or -362/+44 IGF-I exon 2 promoter constructs. Effects of OP-1 on IGF-I mRNA were independent of cell division, as they remained elevated in the presence of hydroxyurea. Cycloheximide inhibited moderately the OP-1-induced increase in IGF-I mRNA, suggesting partial dependency on protein synthesis. On the other hand, the IGF-II nuclear pre-mRNA levels were increased by OP-1 but the half-life of the mature IGF-II mRNA was not affected. Effects of OP-1 on IGF-II mRNA were also independent of cell division, but were dependent on protein synthesis. OP-1 caused a 43-50% reduction in the level of IGFBP-5 nuclear pre-mRNA transcripts and a 40% decrease in the IGFBP-5 promoter activity in FRC cells transfected with the -1278/+1 IGFBP-5 promoter fragment. The half-life of the mature IGFBP-5 mRNA was not affected by OP-1. Hydroxyurea did not prevent the OP-1-induced reduction in IGFBP-5 mRNA. The level of IGFBP-5 mRNA was barely detectable in the presence of cycloheximide, and further suppressive effect of OP-1 on IGFBP-5 mRNA could not be determined. In conclusion, OP-1 regulates IGF-I gene expression at the posttranscriptional level, but regulates IGF-II and IGFBP-5 gene expression at the transcriptional level.