A GABP-binding element in the Robo4 promoter is necessary for endothelial expression in vivo

• Published in: Blood 2008-09, Vol.112 (6), p.2336-2339
• Main Authors: Okada, Yoshiaki, Jin, Enjing, Nikolova-Krstevski, Vesna, Yano, Kiichiro, Liu, Ju, Beeler, David, Spokes, Katherine, Kitayama, Mie, Funahashi, Nobuaki, Doi, Takefumi, Janes, Lauren, Minami, Takashi, Oettgen, Peter, Aird, William C.
• Format: Article
• Language: English
• Subjects: Animals; Binding Sites - genetics; Biological and medical sciences; Embryo, Mammalian; Endothelium - metabolism; GA-Binding Protein Transcription Factor - metabolism; Hematologic and hematopoietic diseases; Hemostasis, Thrombosis, and Vascular Biology; Humans; Medical sciences; Mice; Mice, Transgenic; Mutation; Neoplasms, Experimental; Promoter Regions, Genetic; Receptors, Cell Surface - genetics; Tissue Distribution; Transplantation, Heterologous
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood-2008-01-135079

We recently demonstrated that the 3-kb 5′-flanking region of the human ROBO4 gene directs endothelial cell–specific expression in vitro and in vivo. Moreover, a GA-binding protein (GABP)–binding motif at −119 was necessary for mediating promoter activity in vitro. The goal of the present study was to confirm the functional relevance of the −119 GABP-binding site in vivo. To that end, the Hprt locus of mice was targeted with a Robo4-LacZ transgenic cassette in which the GABP site was mutated. In other studies, the GABP mutation was introduced into the endogenous mouse Robo4 locus in which LacZ was knocked-in. Compared with their respective controls, the mutant promoters displayed a significant reduction in activity in embryoid bodies, embryos, and adult animals. Together, these data provide strong support for the role of the GABP-binding motif in mediating Robo4 expression in the intact endothelium.