Platelet-derived growth factor (PDGF) is a potent mitogen and chemoattractant constitutively expressed by a variety of normal and transformed cells. Transient transfection and deletion analysis of the human c-sis proto-oncogene in cultured vascular endothelial cells revealed a minimal core promoter region extending 82 base pairs upstream from the TATA box. Two novel and functional cis-acting elements were identified within the core that share considerable sequence homology with consensus binding elements for transacting factors of the ETS class and those involved in AP-1 complexes. Deletion or mutation of either the ETS-like site or the AP-1-like site resulted in significant attenuation in the ability of the core to drive transcription. Electrophoretic mobility shift assays revealed that proteins from bovine aortic and human umbilical 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 DNase I footprint analysis were used to identify a functional Sp1 binding site downstream of these elements in the core promoter. By localizing 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.