PARATHYROID-HORMONE VERSUS PHORBOL ESTER STIMULATION OF ACTIVATOR PROTEIN-1 GENE FAMILY MEMBERS IN RAT OSTEOSARCOMA CELLS

We have previously shown that in the rat osteoblastic osteosarcoma cel l line-UMR 106-01-PTH induces maximal collagenase mRNA levels at 4 hou rs. Since this response to PTH requires de novo protein synthesis, it may be mediated by the combined temporal expression of members of the activator protein-1 (AP-1) gene family. We have demonstrated that maxi mal mRNA levels of two of the members of this family, c-fos and c-jun, occur 30 min after stimulation by PTH. Phorbol myristate acetate (PMA ) elicits a similar increase in c-fos and c-jun mRNAs, but is unable t o stimulate transcription of collagenase in these cells. To investigat e further the involvement of the AP-1 gene family, we examined PTH and PMA stimulation of jun-B, jun-D, fos B, and fra-1 mRNAs in UMR 106-01 cells. The mRNA for jun-D was abundant under control conditions and s howed no variation in response to PTH (10(-8) M). The fos B transcript s were not detected under control conditions, whereas jun-B and fra-1 mRNAs were present at low basal levels. PTH caused an increase in fos B mRNA that reached a maximal 4- to 5-fold plateau between 45 and 60 m in. An increase in jun-B mRNA in response to PTH was detectable at 30 min, but reached a maximal 6- to 7-fold increase at 2 hours. After PTH stimulation, the fra-1 transcript showed a 10- to 11-fold peak at 4 h ours. PMA (2.6 x 10(-7) M) stimulated fos B mRNA to maximal abundance at 1 hour, similar to PTH. in contrast, PMA caused a maximal increase in jun-B mRNA at 30 min and fra-1 mRNA at 2 hours, which was earlier t han the response to PTH. To determine whether an increase in jun-B at the same time as c-fos and c-jun would inhibit collagenase gene transc ription, we cotransfected an expression vector for jun-B with a rat co llagenase promoter-reporter gene construct. This resulted in a decreas e in PTH-stimulation of promoter activity. Thus, it appears that the d ifferential temporal stimulation of the AP-1 genes by PTH and PMA, par ticularly an increase in jun-B at the same time as c-fos and c-jun, ex plains the difference seen in their ability to induce transcription of collagenase.