Transcriptional regulation of osteopontin production in rat osteosarcoma cells by type beta transforming growth factor.

Abstract

Type beta transforming growth factor (TGF beta) was shown to regulate the production of several extracellular matrix proteins. Osteopontin (OP) is a recently discovered bone matrix protein which was shown to promote the attachment of osteoblastic rat osteosarcoma ROS 17/2.8 cells to their substrate. We examined the effects of TGF beta on OP production and OP mRNA in ROS 17/2.8 cells. Four-day treatment with 4 ng/ml TGF beta 1 increased substantially the level of osteopontin in the cell culture media, as estimated by immunoblotting. Metabolic labeling showed that this effect was associated with a 3-4-fold increase in OP biosynthesis. TGF beta 1 also increased, in a dose-dependent manner starting at 0.4 ng/ml, the steady-state level of OP mRNA. The increase in OP mRNA was first detected 48 h after the addition of TGF beta 1 and lasted at least until 120 h. The half-life of OP mRNA, estimated in the presence of 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole, was about 10 h and was not altered by TGF beta 1. On the other hand, the increase in OP mRNA was blocked by actinomycin D. Nuclear run-on assays indicated that TGF beta 1 increased the rate of transcription of the OP gene. Examination of hormonal interactions showed that TGF beta 1 opposed or compensated for the reduction in OP mRNA produced by dexamethasone and that TGF beta 1 did not further augment OP mRNA levels which had been increased by 1,25-dihydroxyvitamin D3 treatment. TGF beta 2 had similar effects on OP gene expression as TGF beta 1. In conclusion, TGF beta promotes the production of osteopontin in the osteoblastic osteosarcoma cells through a pathway which is at least in part mediated by transcriptional events.