Cloning of the promoter region of human endoglin, the target gene for hereditary hemorrhagic telangiectasia type 1

Endoglin (CD105) is a cell surface component of the transforming growth fac tor-beta (TGF-beta) receptor complex highly expressed by endothelial cells. Mutations in the endoglin gene are responsible for the hereditary hemorrha gic telangiectasia type 1 (HHT1), also known as Osler-Weber-Rendu syndrome (OMIM 187300). This is an autosomal dominant vascular disorder probably cau sed by a haploinsufficiency mechanism displaying low levels of the normal p rotein. To understand the mechanisms underlying the regulated expression of endoglin, a genomic DNA clone containing 3.3 kb of the 5'-flanking sequenc e of the human endoglin gene has been isolated. The 5'-flanking region of t he endoglin gene lacks consensus TATA and CAAT boxes, but contains two GC-r ich regions and consensus motifs for Sp1, ets, GATA, AP-2, NF kappa B, and Mad, as well as TGF-beta-, glucocorticoid-, vitamin D-, and estrogen-respon sive elements. As determined by primer extension and 5' RACE experiments, a cluster of transcriptional start sites was found to be located 350 bp upst ream from the translation initiation codon. To analyze the endoglin promote r activity, the upstream -400/+341 fragment was fused to the luciferase gen e and transient transfections were conducted in several cell types. This co nstruct displayed a tissue-specific activity in human and bovine endothelia l cells. Analysis of various deletion constructs showed the existence of a basal promoter region within the -81/+350 fragment as well as major transcr iptional regulatory elements within the -400/-141 fragment. Electrophoretic mobility shift assays demonstrated the specific interaction of a member of the ets family with a consensus motif located at position -68. A promoter construct mutated at this ets sequence showed a much reduced activity as co mpared with the wild-type construct, supporting the involvement of this ets motif in the basal activity of the promoter. The endoglin promoter exhibit ed inducibility in the presence of TGF-beta 1, suggesting possible therapeu tic treatments in HHT1 patients, in which the expression level of the norma l endoglin allele might not reach the threshold required for its function. Isolation and characterization of the human endoglin promoter represents an initial step in elucidating the controlled expression of the endoglin gene . (C) 1998 by The American Society of Hematology.