Inhibition of endothelial cell activation by bHLH protein E2-2 and its impairment of angiogenesis.
E2-2 belongs to the basic helix-loop-helix (bHLH) family of transcription factors. E2-2 associates with inhibitor of DNA binding (Id) 1, which is involved in angiogenesis. In this paper, we demonstrate that E2-2 interacts with Id1 and provide evidence that this interaction potentiates angiogenesis. Mutational analysis revealed that the HLH domain of E2-2 is required for the interaction with Id1 and vice versa. In addition, Id1 interfered with E2-2-mediated effects on luciferase reporter activities. Interestingly, injection of E2-2-expressing adenoviruses into Matrigel plugs implanted under the skin blocked in vivo angiogenesis. In contrast, the injection of Id1-expressing adenoviruses rescued E2-2-mediated inhibition of in vivo angiogenic reaction. Consistent with the results of the Matrigel plug assay, E2-2 could inhibit endothelial cell (EC) migration, network formation, and proliferation. On the other hand, knockdown of E2-2 in ECs increased EC migration. The blockade of EC migration by E2-2 was relieved by exogenous expression of Id1. We also demonstrated that E2-2 can perturb VEGFR2 expression via inhibition of VEGFR2 promoter activity. This study suggests that E2-2 can maintain EC quiescence and that Id1 can counter this effect.
Department of Experimental Pathology, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba;
Aya
A
Tanaka
Fumiko
F
Itoh
Koichi
K
Nishiyama
Toshiaki
T
Takezawa
Hiroki
H
Kurihara
Susumu
S
Itoh
Mitsuyasu
M
Kato
D000818:Animals; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism; D004730:Endothelium, Vascular / Q000502:physiology*; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D047468:Immunoprecipitation; D051798:Inhibitor of Differentiation Protein 1 / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009389:Neovascularization, Pathologic / Q000378:metabolism / Q000517:prevention & control*; D011401:Promoter Regions, Genetic; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051820:TCF Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000073940:Transcription Factor 4; D058118:Transcription Factor 7-Like 2 Protein; D014158:Transcription, Genetic; D020798:Two-Hybrid System Techniques; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics / Q000378:metabolism
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; C497453:ID1 protein, human; C497455:Idb1 protein, mouse; D051798:Inhibitor of Differentiation Protein 1; D034741:RNA, Small Interfering; D051820:TCF Transcription Factors; C544894:TCF7L2 protein, human; C090743:Tcf4 protein, mouse; C544895:Tcf7l2 protein, mouse; D000073940:Transcription Factor 4; D058118:Transcription Factor 7-Like 2 Protein; D040301:Vascular Endothelial Growth Factor Receptor-2