Abstract 195: Characterization of DNA Modules in Regulating VWF Vascular Bed--Specific Expression

Abstract

Expression of von Willebrand Factor (vWF) in the endothelium is regulated by vascular bed-specific pathways. A region of the human vWF gene between -843 and -620 was previously shown to direct expression in the endothelium of capillaries and a subset of larger blood vessels in the heart and skeletal muscle. Here, our goal was to further delineate the responsible DNA sequences. A series of constructs containing 5’-deletions, internal deletions or point mutations of the vWF promoter coupled to LacZ were targeted to the Hprt locus of mice using homologous recombination, and the resulting animals were analyzed for reporter gene expression. The findings demonstrate that a three-base-pair fragment within vWF promoter is necessary for expression in capillary but not large vessel endothelium in heart and skeletal muscle. Although the sequence does not conform to established DNA binding sites, it bound to nuclear protein as determined by elecrophoretic mobility shift assay. Mass spectrometry of the DNA-protein complex revealed several potential binding transcription factors. The heart- and skeletal muscle-specific promoter region did not response to shear stress or mechanical stress. Bisulfite sequencing revealed differential methylation of the human vWF promoter in endothelial cells from expressing and non-expressing vascular beds. These differences were altered by the mutation at the short cis-element. Together, these data support a model of modular gene regulation in which distinct DNA sequences in the vWF promoter direct expression in different vascular beds.