REGULATION OF ENDOTHELIN-1 GENE EXPRESSION

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Endothelin-1 (ET-1) is a 21-amino acid peptide synthesized in endothelial cells. It is both a potent vasoconstrictor and growth factor. The role of ET-1 in vascular physiology and its involvement in vascular disease is unknown. Aims of the proposed work include the use of genetic studies to better understand the function of ET-1, the use of this gene as a model system for the study of endothelial cell transcription, and the use of ET-1 regulatory sequences for endothelial cell targeting in transgenic animals. We will elucidate the cis-acting DNA elements of the ET-1 gene in vitro by transfection of ET-1 sequences linked to a reporter gene. These experiments will be complemented by analogous studies in vivo using transgenic mice which carry ET-1-lacZ transgenes. In vitro footprinting and gelshift analysis will allow characterization of trans-acting proteins which bind these sequences. ET-1 cis-acting sequences which appear to be specific for endothelial cell transcription will be employed in the purification and cloning of DNA binding proteins. Elements of the ET-1 promoter which provide high level endothelial cell expression in transgenic animals will be utilized to develop lines of transgenic mice which express either increased or attenuated levels of ET-1 in the blood vessel wall. These mice will be evaluated for alterations in blood pressure and predisposition for vascular disease. The availability of a clone for an endothelial cell-specific trans-acting factor will make possible studies investigating the development and phenotypic diversity of this important cell type. The availability of constructs which direct endothelial cell expression in transgenic mice will allow an approach to the generation of mouse models of vascular disease.