NOVEL CIS-ACTING ELEMENTS IN THE HUMAN PLATELET-DERIVED GROWTH-FACTORB-CHAIN CORE PROMOTER THAT MEDIATE GENE-EXPRESSION IN CULTURED VASCULAR ENDOTHELIAL-CELLS

Platelet-derived growth factor (PDGF) is a potent mitogen and chemoatt ractant constitutively expressed by a variety of normal and transforme d cells. Transient transfection and deletion analysis of the human c-s is proto-oncogene in cultured vascular endothelial cells revealed a mi nimal core promoter region extending 82 base pairs upstream from the T ATA box. Two novel and functional cis-acting elements were identified within the core that share considerable sequence homology with consens us binding elements for transacting factors of the ETS class and those involved in AP-1 complexes. Deletion or mutation of either the ETS-li ke site or the AP-1-like site resulted in significant attenuation in t he ability of the core to drive transcription. Electrophoretic mobilit y shift assays revealed that proteins from bovine aortic and human umb ilical vein endothelial nuclear extracts bound to these elements in a specific manner and that both sites were essential for protein binding . Ferguson analysis predicted a combined molecular mass of 153 kDa for these proteins. In addition, transient transfection, gel shift, and D Nase I footprint analysis were used to identify a functional Sp1 bindi ng site downstream of these elements in the core promoter By localizin g the functional cis-acting elements in the PDGF-B promoter, it may be possible to elucidate the normal transcriptional control of the gene, as well as the mechanisms that activate it in pathologic settings.