Parathyroid hormone regulation of the rat collagenase-3 promoter by protein kinase A-dependent transactivation of core binding factor alpha 1

Previously we showed that the activator protein-1 site and the runt domain binding site in the collagenase-3 promoter act cooperatively in response to parathyroid hormone (PTH) in the rat osteoblastic osteosarcoma cell line, UMR 106-01, Our results of the expression pattern of core binding factor al pha 1 (Cbfa1), which binds to the runt domain site, indicated that there is no change in the levels of Cbfa1 protein or RNA under either control condi tions or after PTH treatment. The importance of posttranslational modificat ion of Cbfa1 in the signaling pathway for PTH-induced collagenase-3 promote r activity was analyzed. PTH stimulation of collagenase-3 promoter activity was completely abrogated by protein kinase A (PKA) inhibition. To determin e the role of PKA activity with respect to Cbfa1 activation (in addition to its known activity of phosphorylating cAMP-response element-binding protei n to enhance c-fos promoter activity), we utilized the heterologous Gal4 tr anscription system. PTH stimulated the transactivation of activation domain -3 in Cbfa1 through the PKA site. In vitro phosphorylation studies indicate d that the PKA site in the wild type activation domain-3 is a substrate for phosphorylation by PKA, Thus, we demonstrate that PTH induces a PKA-depend ent transactivation of Cbfa1, and this transactivation is required for coll agenase-3 promoter activity in UMR cells.