CLOTTING AND FUNCTIONAL-ANALYSIS OF THE PROMOTER FOR KDR/FLK-1, A RECEPTOR FOR VASCULAR ENDOTHELIAL GROWTH-FACTOR

KDR/flk-1 is one of two receptors for vascular endothelial growth factor, a potent angiogenic peptide, KDR/flk-1 is an early marker for endothelial cell progenitors, and its expression is restricted to endothelial cells in vivo, To investigate the molecular mechanisms regulating expression of KDR/flk-1, we cloned and characterized the promoter of the human KDR/flk-1 gene, The transcription start site was localized by primer extension and ribonuclease protection to a nucleotide 303 base pairs (bp) 5' of the initiation methionine codon, The 5'-flanking sequence is rich in G and C residues and contains five Spl elements but no TATA consensus sequence, By reporter gene transfection experiments, we found that similar to 4 kilobases of KDR/flk-1 5'-flanking sequence directed high level luciferase activity in bovine aortic endothelial cells; further deletion analysis revealed positive regulatory elements between bp -225 to -164, -95 to -77, -77 to -60, and +105 to +127. Mutation of an atypical GATA sequence between bp +105 and +127 did not affect promoter activity, suggesting that GATA elements are not essential for the high level promoter activity of this gene, Consistent with endothelial cell-restricted expression of KDR/flk-1 mRNA, we found that the 4-kilobase flanking sequence directed high level promoter activity in endothelial cells but not in other cell types, To our knowledge this is the first report characterizing the KDR/flk-1 promoter, Understanding the KDR/flk-1 promoter will allow us to investigate endothelial cell-specific gene regulation and to uncover methods for targeting gene delivery specifically to endothelial cells.