Primary Study Of Regulation Mechanism About Osterix's Promoter

Objective: To search for the potential BMP-2 stimulated regions upstream of Osterix gene, as the base for further research.Method: The possibility that BMP-2 stimulates Osterix expression had been detected at transcription level by Real-Time Q-PCR. As a first step toward localizing important control elements within the Osterix promoter, 9 different fragment of 5' deletion mutants were amplified by PCR form the BAC clone BAC-RP23-118K10. The mutant promoters were inserted upstream of the luciferase gene in luciferase report vector pGL3 Basic. Co-transformation the Osterix promoter/luciferase plasmid and internal control pRL-TK into C2C12, C3H10T1/2 and MC3T3-E1 cell lines. After transfection, cells were induced by BMP-2 for 48 h and then were detected by Dual-Luciferase Reporter Assay System.Result: At transcription level we detected that Osterix was stimulated by BMP-2. 9 different 5' deletion mutants Osterix promoter/luciferase reporter vectors were successfully constructed. Results of the luciferase activity measurement show that there are strong potential activating elements in the region of -100～+81,-760～ -560 and -1254～-1042, while strong potential repressive domains which related to BMP-2 are observed in the -100～+81 and -760～-560 region of the murine Osterix promoter.Conclusion: The core promoter region of murine Osterix has been primarily identified, and a platform for Osterix transcriptional regulation research has been constructed.