Transcription regulation of the vegf gene by the BMP/Smad pathway in the angioblast of zebrafish embryos.
Vascular endothelial growth factor (VEGF) is a mitogen that is critically involved in vasculogenesis, angiogenesis, and hematopoiesis. However, what and how transcription factors participate in the regulation of vegf gene expression are not fully understood. Here we report the cloning and sequencing of the zebrafish vegf promoter which revealed that the promoter contains a number of bone morphogenetic protein (BMP)-activated Smad binding elements (SBE), implicating Smad1 and Smad5 in the regulation of BMP-induced expression of vegf. Electrophoretic mobility shift assays of adding recombinant Smad proteins to the SBE-containing DNA oligonucleotides that represent portions of zebrafish vegf promoter resulted in mobility shift of the oligonucleotides. These changes demonstrate potential interactions between Smad1/5 and the vegf promoter. Reporter activity assays using the wild-type or SBE-deleted vegf promoters to drive the luciferase reporter gene expression revealed that Smad1 stimulated while Smad5 repressed the vegf promoter activity in zebrafish embryos. These data indicate that the BMP/Smad signaling pathway is involved in the regulation of zebrafish vegf transcription. In addition, we demonstrate that transgenic expression of human BMP4 in zebrafish embryos induced an expansion of the posterior intermediate cell mass (ICM, also commonly called blood island), a population of cells containing endothelial and hematopoietic precursors. In the expanded ICM, vegf and VEGF receptor 2 (flk-1) were ectopically co-expressed, suggesting that an autocrine/paracrine regulation of vegf expression may exist and contribute to the BMP-induced hemangiogenic cell proliferation.
He
Chen
C
The Department of Biological Sciences, Ohio University, Athens, OH 45701, USA.
Chen
Xiaozhuo
X
D000818:Animals; D001483:Base Sequence; D019485:Bone Morphogenetic Proteins / Q000737:chemistry / Q000378:metabolism*; D049109:Cell Proliferation; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006801:Humans; D017403:In Situ Hybridization; D008156:Luciferases / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018919:Neovascularization, Physiologic; D009841:Oligonucleotides / Q000737:chemistry; D010750:Phosphoproteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry; D015398:Signal Transduction; D051785:Smad Proteins; D051898:Smad1 Protein; D051902:Smad5 Protein; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D014158:Transcription, Genetic; D019076:Transgenes; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000235:genetics*; D015027:Zebrafish; D029961:Zebrafish Proteins
D019485:Bone Morphogenetic Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D009841:Oligonucleotides; D010750:Phosphoproteins; D011994:Recombinant Proteins; C497399:SMAD1 protein, human; D051785:Smad Proteins; D051898:Smad1 Protein; D051902:Smad5 Protein; D015534:Trans-Activators; D042461:Vascular Endothelial Growth Factor A; D029961:Zebrafish Proteins; C497801:smad5 protein, zebrafish; D008156:Luciferases