title: Sp1 is involved in Akt-mediated induction of VEGF expression through an HIF-1-independent mechanism.
abstract: Increased expression of vascular endothelial growth factor (VEGF) contributes to the growth of many tumors by increasing angiogenesis. Although hypoxia is a potent inducer of VEGF, we previously showed that epidermal growth factor receptor amplification and loss of PTEN, both of which can increase phosphatidylinositol-3-kinase (PI3K) activity, increase VEGF expression. Using both adenoviral vectors and a cell line permanently expressing constitutively active myristoylated Akt (myrAkt), we show that activation of Akt, which is downstream of PI3K, increases VEGF expression in vitro and increases angiogenesis in a Matrigel plug assay. Transient transfection experiments using reporter constructs containing the VEGF promoter showed that up-regulation of VEGF by Akt is mediated through Sp1 binding sites located in the proximal promoter. Small interfering RNA directed against Sp1 prevented the induction of VEGF mRNA in response to myrAkt but not to hypoxia. Expression of myrAkt is associated with increased phosphorylation of Sp1 and its increased binding to a probe corresponding to the -88/-66 promoter region. In conclusion, our results indicate that Sp1 is required for transactivation of the VEGF by Akt. Others have proposed that the PI3K/Akt pathway can increase VEGF expression via the hypoxia-inducible factor 1 (HIF-1); however, our results suggest an alternative mechanism can also operate.
authors.affiliation: Department of Radiation Oncology, University of Pennsylvania School of Medicine, Philadelphia, PA 19004, USA.
authors.forename: Nabendu
authors.initials: N
authors.lastname: Pore
authors.forename: Shuang
authors.initials: S
authors.lastname: Liu
authors.forename: Hui-Kuo
authors.initials: HK
authors.lastname: Shu
authors.forename: Bin
authors.initials: B
authors.lastname: Li
authors.forename: Daphne
authors.initials: D
authors.lastname: Haas-Kogan
authors.forename: David
authors.initials: D
authors.lastname: Stokoe
authors.forename: Julie
authors.initials: J
authors.lastname: Milanini-Mongiat
authors.forename: Gilles
authors.initials: G
authors.lastname: Pages
authors.forename: Donald M
authors.initials: DM
authors.lastname: O'Rourke
authors.forename: Eric
authors.initials: E
authors.lastname: Bernhard
authors.forename: Amit
authors.initials: A
authors.lastname: Maity
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000469:Alkaline Phosphatase / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D015152:Blotting, Northern; D015153:Blotting, Western; D002460:Cell Line; D045744:Cell Line, Tumor; D003094:Collagen / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000096:biosynthesis*; D003720:Densitometry; D004338:Drug Combinations; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D000860:Hypoxia; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007797:Laminin / Q000494:pharmacology; D008969:Molecular Sequence Data; D009389:Neovascularization, Pathologic; D009687:Nuclear Proteins / Q000096:biosynthesis*; D010710:Phosphates / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D008926:Plicamycin / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011509:Proteoglycans / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016329:Sp1 Transcription Factor / Q000502:physiology*; D014157:Transcription Factors / Q000096:biosynthesis*; D015533:Transcriptional Activation; D014162:Transfection; D015854:Up-Regulation; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D004338:Drug Combinations; C497442:HIF1A protein, human; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007797:Laminin; D009687:Nuclear Proteins; D010710:Phosphates; D011509:Proteoglycans; D034741:RNA, Small Interfering; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; C059485:matrigel; D049452:Green Fluorescent Proteins; D003094:Collagen; D019869:Phosphatidylinositol 3-Kinases; D000469:Alkaline Phosphatase; D008926:Plicamycin