A GABP-binding element in the Robo4 promoter is necessary for endothelial expression in vivo.
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.
Okada
Yoshiaki
Y
Center for Vascular Biology Research and Division of Molecular and Vascular Medicine, Beth Israel Deaconess Medical Center, Boston, MA 02215, USA.
Jin
Enjing
E
Nikolova-Krstevski
Vesna
V
Yano
Kiichiro
K
Liu
Ju
J
Beeler
David
D
Spokes
Katherine
K
Kitayama
Mie
M
Funahashi
Nobuaki
N
Doi
Takefumi
T
Janes
Lauren
L
Minami
Takashi
T
Oettgen
Peter
P
Aird
William C
WC
D000818:Animals; D001665:Binding Sites / Q000235:genetics; D004622:Embryo, Mammalian; D004727:Endothelium / Q000378:metabolism*; D051268:GA-Binding Protein Transcription Factor / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation; D009374:Neoplasms, Experimental; D011401:Promoter Regions, Genetic; D011956:Receptors, Cell Surface / Q000235:genetics*; D014018:Tissue Distribution; D014183:Transplantation, Heterologous; D000097905:Roundabout Proteins
D051268:GA-Binding Protein Transcription Factor; D011956:Receptors, Cell Surface; C462951:ROBO4 protein, human; D000097905:Roundabout Proteins