Human von Willebrand factor gene sequences target expression to a subpopulation of endothelial cells in transgenic mice.

Abstract

The present study was undertaken to define the 5' and 3' regulatory sequences of human von Willebrand factor gene that confer tissue-specific expression in vivo. Transgenic mice were generated bearing a chimeric construct that included 487 bp of 5' flanking sequence and the first exon fused in-frame to the Escherichia coli lacZ gene. In situ histochemical analyses in independent lines demonstrated that the von Willebrand factor promoter targeted expression of LacZ to a subpopulation of endothelial cells in the yolk sac and adult brain. LacZ activity was absent in the vascular beds of the spleen, lung, liver, kidney, testes, heart, and aorta, as well as in megakaryocytes. In contrast, in mice containing the lacZ gene targeted to the thrombomodulin locus, the 5-bromo-4-chloro-3-indolyl 38-D-galactopyranoside reaction product was detected throughout the vascular tree. These data highlight the existence of regional differences in endothelial cell gene regulation and suggest that the 733-bp von Willebrand factor promoter may be useful as a molecular marker to investigate endothelial cell diversity.The development of the cardiovascular system is dependent upon a tightly regulated program beginning with differentiation of endothelial cells from mesoderm, followed by their subsequent assembly into vascular channels, and ending with the establishment of organ-and region-specific endothelial cell properties (1, 2). Although the heterogeneity of endothelial cells has been defined at the level of cell structure, function, antigenic composition, and response to environmental stimuli (3-8), the molecular mechanisms that lead to this phenotypic diversity are poorly understood.The von Willebrand factor (VWF) is a multimeric glycoprotein that mediates adhesion of platelets to damaged endothelium and serves as a carrier for the antihemophilic factor (for reviews, see refs. 9 and 10). The cellular distribution of VWF is restricted to the endothelium, megakaryocytes, and platelets. Histochemical studies of mouse embryos show that the VWF gene is expressed early in vascular development in a limited subset of endothelial cells (2). In adult mammals, VWF is also heterogeneously distributed in different vascular beds and is associated with regional variations in mRNA levels (7,(11)(12)(13)

Ets transcription factors bind and transactivate the core promoter of the von Willebrand factor gene

“…Results from transgenesis studies have suggested that other unknown trans-acting positive regulatory elements should interact with the DNA in the regions external to the 7487 to +246 domain in order to promote the vWF gene transcription in vivo (Aird et al, 1995). The Ets proteins of the ternary complex might also operate synergistically with these unknown regulators.…”

“…The cis-acting element, required for expression, is a GATA transcription factor binding site, situated at position +220 in the ®rst exon. This 7487 to +246 fragment was fused to the E. coli LacZ gene and used to generate transgenic mice (Aird et al, 1995). Only a limited subpopulation of yolk sac and adult brain endothelial cells in the transgenic animals expressed the LacZ construct and b-galactosidase activity was also detected in brain cells that do not synthesize endogenous vWF.…”

Role of a 5′-enhancer in the transcriptional regulation of the human endothelial cell protein C receptor gene

“…Despite considerable progress in our knowledge of EC gene regulation and vascular bed specificity, [28][29][30][31] there has been incomplete understanding of the mechanisms targeting the expression of genes such as EPCR to large-vessel ECs. In a study by Gu et al, 14 transgenic mice were generated that expressed the green fluorescent protein under the control of mEPCR promoter fragments (extending up to Ϫ1080 bp).…”

Differential roles for ETS, CREB, and EGR binding sites in mediating VEGF receptor 1 expression in vivo

“…Genotyping was performed by PCR analysis. Whole-mount and tissue section LacZ and CD31 staining of embryonic and adult tissues, ␤-galactosidase activity assays, and tumor xenografts were carried out as previously described 6,7 (and supplemental Methods).…”