THE ROLE OF THE 5'-FLANKING REGION IN THE CELL-SPECIFIC TRANSCRIPTIONOF THE HUMAN VON-WILLEBRAND-FACTOR GENE

Transcriptional regulation of the human von Willebrand factor (vWF) ge ne was investigated in calf pulmonary artery endothelial (CPAE), HeLa, COS 7 and Hep G2 cells. Various lengths of flanking sequences extendi ng up to 2123 bp 5' of the transcription initiation site and containin g 19 bp of the first exon, were linked to the bacterial chloramphenico l acetyltransferase (CAT) gene and these constructs were assayed in tr ansient transfection assays. Sequences up to 89 bp upstream of the cap site showed transcriptional activity in all cell types. Sequences bet ween - 147 and - 419 bp markedly reduced CAT activity in CPAE cells an d abolished it in other cell lines. A domain from - 592 to - 810 bp ge nerated low levels of expression only in CPAE cells. This transcriptio nal activity was repressed with constructs containing 1041 to 1240 bp upstream of the cap site. Endothelial cell-specific transcription was restored by a construct that contained 1286 bp upstream of the cap sit e. The additional 46 bp upstream of the negative regulatory domain wer e within the 5' end of an inverse human Alu-family DNA repeat. RNAase- protection assays confirmed the correct transcriptional initiation. Th e sequence between - 89 and - 420 contained at least one negative regu latory element able to repress the CAT gene expression controlled by t he heterologous thymidine kinase promoter in all ce types. A construct that included the sequence from - 89 to - 1286 bp increased the trans criptional activity directed by the thymidine kinase promoter only in CPAE cells. These results indicate that negative and positive elements in the 5'-flanking region interact to regulate vWF gene expression.