My long term career goal is to investigate the molecular mechanisms that mediate the development of blood vessels. Recent studies have shown that the expression of the endothelial cell-specific genes tie1, tie2, flk1 and flt1 is essential for normal vascular development in the mouse embryo. Unlike the hematopoietic and myogenic lineages where the identification of cell-specific transcription factors has provided insight into the early events of lineage determination, differentiation and tissue development, no endothelial-specific transcription factor has so far been identified and the transcriptional mechanisms that regulate specific expression in this cell type remain obscure. Guided by transcription studies of the tie2 promoter in transgenic mice, a cis- acting octamer element ATGCAAAT was found to exhibit an endothelial cell-specific pattern of binding in electrophoretic mobility shift assay. As expected, this element bound the ubiquitously expressed transcription factor Oct1. In addition, it formed a ternary complex with Oct1 and a novel cofactor, Ecco1, whose expression is restricted to endothelial cells. The octamer DNA element was also found in the regulatory regions of three additional endothelial cell-specific genes including tie1, flk1 and von Willebrand factor, all of which exhibited an identical and cell-specific pattern of binding. We hypothesize the Ecco1 is a master regulatory factor for the endothelial lineage, that it positively regulates the expression of endothelial cell genes and that it is essential for normal vascular development in the mouse embryo. The following specific aims are proposed: 1) To clone the mouse DNA encoding Ecco1. 2) To verify that Ecco1 is an interacting partner of Oct1 and capable of transactivating the promoters of several endothelial cell-specific genes. 3) To study the biochemical properties of Ecco1 by investigating its DNA binding and protein binding characteristics. 4) To examine the expression pattern of Ecco1 in different cell types and during vascular development in mouse embryonic tissues. 5) To examine the role of Ecco1 in the development of blood vessels through gain-of-function experiments in transgenic mice. Future experiments will be designed to address the developmental role of this factor using in vivo loss-of-function studies. Contributions from this work is likely to improve our knowledge of vascular development and may provide new therapeutic options to promote angiogenesis in conditions such as coronary and peripheral vascular disease.
