INDUCTION IN HUMAN OSTEOBLASTIC CELLS (SAOS(2)) OF THE EARLY RESPONSEGENES FOS, JUN, AND MYC BY THE AMINO-TERMINAL FRAGMENT (ATF) OF UROKINASE

Previous studies have demonstrated that overexpression of urinary plas minogen activator (uPA) in rat prostate cancer cells results in increa sed skeletal metastases, which are primarily of the osteoblastic varie ty. The osseous activation induced by the metastases appears to be med iated through the amino terminal fragment (ATF) of uPA, which lacks th e catalytic domain and can act as a growth factor for osteoblasts. To explore further the mechanism of action of uPA in bone cells, we evalu ated the effects of ATF on modulating the expression of various protoo ncogenes. Human-osteoblast-derived osteosarcoma cells, SaOS(2), were t reated with graded doses of ATF for 10-120 min, and effects on early r esponse protooncogenes were monitored. ATF increased c-myc, c-jun, and c-fos gene expression in a time-dependent manner for up to 60 min, af ter which mRNA levels fell. The maximum induction was seen in c-fos ge ne expression, which was found to be dose dependent. This effect of AT F was localized to its growth-factorlike domain. Examination of the ha lf life of these transcripts in the presence of the transcriptional in hibitor actinomycin D demonstrated that ATF does not alter the stabili ty of c-fos mRNA in these bone cells. Nuclear run-off assays indicated that ATF effects were due to stimulation of c-fos gene transcription. An increase in c-fos protein levels was correlated with the augmentat ion of its mRNA in ATF-treated SaOS(2) cells. Pretreatment of SaOS(2) cells with the protein tyrosine kinase inhibitor herbimycin and recomb inant soluble uPA receptor (uPAR) caused a significant reduction in th e ability of ATF to induce c-fos expression. These results demonstrate a novel role for uPA in activating early response proto-oncogenes, in particular c-fos, which plays an important role in bone cell growth a nd differentiation and may be a key factor in the signal transduction pathway of ATF. (C) 1997 Wiley-Liss, Inc.